Matthew P. Scott

All Publications

• Common Regulatory Targets of NFIA, NFIX and NFIB during Postnatal Cerebellar Development. Cerebellum (London, England) Fraser, J. n., Essebier, A. n., Brown, A. S., Davila, R. A., Harkins, D. n., Zalucki, O. n., Shapiro, L. P., Penzes, P. n., Wainwright, B. J., Scott, M. P., Gronostajski, R. M., Bodén, M. n., Piper, M. n., Harvey, T. J. 2019

  Abstract

  Transcriptional regulation plays a central role in controlling neural stem and progenitor cell proliferation and differentiation during neurogenesis. For instance, transcription factors from the nuclear factor I (NFI) family have been shown to co-ordinate neural stem and progenitor cell differentiation within multiple regions of the embryonic nervous system, including the neocortex, hippocampus, spinal cord and cerebellum. Knockout of individual Nfi genes culminates in similar phenotypes, suggestive of common target genes for these transcription factors. However, whether or not the NFI family regulates common suites of genes remains poorly defined. Here, we use granule neuron precursors (GNPs) of the postnatal murine cerebellum as a model system to analyse regulatory targets of three members of the NFI family: NFIA, NFIB and NFIX. By integrating transcriptomic profiling (RNA-seq) of Nfia- and Nfix-deficient GNPs with epigenomic profiling (ChIP-seq against NFIA, NFIB and NFIX, and DNase I hypersensitivity assays), we reveal that these transcription factors share a large set of potential transcriptional targets, suggestive of complementary roles for these NFI family members in promoting neural development.

  View details for DOI 10.1007/s12311-019-01089-3

  View details for PubMedID 31838646

• Granule neuron precursor cell proliferation is regulated by NFIX and intersectin 1 during postnatal cerebellar development. Brain structure & function Fraser, J., Essebier, A., Brown, A. S., Davila, R. A., Sengar, A. S., Tu, Y., Ensbey, K. S., Day, B. W., Scott, M. P., Gronostajski, R. M., Wainwright, B. J., Boden, M., Harvey, T. J., Piper, M. 2018

  Abstract

  Cerebellar granule neurons are the most numerous neuronal subtype in the central nervous system. Within the developing cerebellum, these neurons are derived from a population of progenitor cells found within the external granule layer of the cerebellar anlage, namely the cerebellar granule neuron precursors (GNPs). The timely proliferation and differentiation of these precursor cells, which, in rodents occurs predominantly in the postnatal period, is tightly controlled to ensure the normal morphogenesis of the cerebellum. Despite this, our understanding of the factors mediating how GNP differentiation is controlled remains limited. Here, we reveal that the transcription factor nuclear factor I X (NFIX) plays an important role in this process. Mice lacking Nfix exhibit reduced numbers of GNPs during early postnatal development, but elevated numbers of these cells at postnatal day 15. Moreover, Nfix-/- GNPs exhibit increased proliferation when cultured in vitro, suggestive of a role for NFIX in promoting GNP differentiation. At a mechanistic level, profiling analyses using both ChIP-seq and RNA-seq identified the actin-associated factor intersectin 1 as a downstream target of NFIX during cerebellar development. In support of this, mice lacking intersectin 1 also displayed delayed GNP differentiation. Collectively, these findings highlight a key role for NFIX and intersectin 1 in the regulation of cerebellar development.

  View details for PubMedID 30511336

• Developmental phosphoproteomics identifies the kinase CK2 as a driver of Hedgehog signaling and a therapeutic target in medulloblastoma. Science signaling Purzner, T., Purzner, J., Buckstaff, T., Cozza, G., Gholamin, S., Rusert, J. M., Hartl, T. A., Sanders, J., Conley, N., Ge, X., Langan, M., Ramaswamy, V., Ellis, L., Litzenburger, U., Bolin, S., Theruvath, J., Nitta, R., Qi, L., Li, X., Li, G., Taylor, M. D., Wechsler-Reya, R. J., Pinna, L. A., Cho, Y., Fuller, M. T., Elias, J. E., Scott, M. P. 2018; 11 (547)

  Abstract

  A major limitation of targeted cancer therapy is the rapid emergence of drug resistance, which often arises through mutations at or downstream of the drug target or through intrinsic resistance of subpopulations of tumor cells. Medulloblastoma (MB), the most common pediatric brain tumor, is no exception, and MBs that are driven by sonic hedgehog (SHH) signaling are particularly aggressive and drug-resistant. To find new drug targets and therapeutics for MB that may be less susceptible to common resistance mechanisms, we used a developmental phosphoproteomics approach in murine granule neuron precursors (GNPs), the developmental cell of origin of MB. The protein kinase CK2 emerged as a driver of hundreds of phosphorylation events during the proliferative, MB-like stage of GNP growth, including the phosphorylation of three of the eight proteins commonly amplified in MB. CK2 was critical to the stabilization and activity of the transcription factor GLI2, a late downstream effector in SHH signaling. CK2 inhibitors decreased the viability of primary SHH-type MB patient cells in culture and blocked the growth of murine MB tumors that were resistant to currently available Hh inhibitors, thereby extending the survival of tumor-bearing mice. Because of structural interactions, one CK2 inhibitor (CX-4945) inhibited both wild-type and mutant CK2, indicating that this drug may avoid at least one common mode of acquired resistance. These findings suggest that CK2 inhibitors may be effective for treating patients with MB and show how phosphoproteomics may be used to gain insight into developmental biology and pathology.

  View details for PubMedID 30206138

• Noncanonical hedgehog pathway activation through SRF-MKL1 promotes drug resistance in basal cell carcinomas. Nature medicine Whitson, R. J., Lee, A. n., Urman, N. M., Mirza, A. n., Yao, C. Y., Brown, A. S., Li, J. R., Shankar, G. n., Fry, M. A., Atwood, S. X., Lee, E. Y., Hollmig, S. T., Aasi, S. Z., Sarin, K. Y., Scott, M. P., Epstein, E. H., Tang, J. Y., Oro, A. E. 2018; 24 (3): 271–81

  Abstract

  Hedgehog pathway-dependent cancers can escape Smoothened (SMO) inhibition through mutations in genes encoding canonical hedgehog pathway components; however, around 50% of drug-resistant basal cell carcinomas (BCCs) lack additional variants of these genes. Here we use multidimensional genomics analysis of human and mouse drug-resistant BCCs to identify a noncanonical hedgehog activation pathway driven by the transcription factor serum response factor (SRF). Active SRF along with its coactivator megakaryoblastic leukemia 1 (MKL1) binds DNA near hedgehog target genes and forms a previously unknown protein complex with the hedgehog transcription factor glioma-associated oncogene family zinc finger-1 (GLI1), causing amplification of GLI1 transcriptional activity. We show that cytoskeletal activation through Rho and the formin family member Diaphanous (mDia) is required for SRF-MKL-driven GLI1 activation and for tumor cell viability. Remarkably, nuclear MKL1 staining served as a biomarker in tumors from mice and human subjects to predict tumor responsiveness to MKL inhibitors, highlighting the therapeutic potential of targeting this pathway. Thus, our study illuminates, for the first time, cytoskeletal-activation-driven transcription as a personalized therapeutic target for combatting drug-resistant malignancies.

  View details for PubMedID 29400712

  View details for PubMedCentralID PMC5839965

• Homeodomains, Hedgehogs, and Happiness ESSAYS ON DEVELOPMENTAL BIOLOGY, PT B Scott, M. P. 2016; 117: 331-337

  Abstract

  Developmental biologists have had a spectacular quarter century of discoveries, building on many decades of work earlier, discovering molecular, cellular, and genetic mechanisms that underlie the magical process by which an egg becomes a plant or animal. Among the discoveries were homeodomains, DNA-binding domains that allow transcription factors to recognize their target genes, and the Hedgehog signaling pathway, which is used in many organs and tissues for communication among cells. The experience of unveiling the mechanisms and molecules connected to both of these findings has been remarkable, joyful, difficult, and a time of great teamwork and collaboration within and between laboratory groups. More than ever it is possible to discern the evolutionary processes, and their mechanisms, that led to the diversity of life on earth. A huge amount of work remains to be done to obtain a broad understanding of what happened and how development works.

  View details for DOI 10.1016/bs.ctdb.2015.11.034

  View details for Web of Science ID 000376937700020

  View details for PubMedID 26969987

• Multiple Surface Regions on the Niemann-Pick C2 Protein Facilitate Intracellular Cholesterol Transport JOURNAL OF BIOLOGICAL CHEMISTRY McCauliff, L. A., Xu, Z., Li, R., Kodukula, S., Ko, D. C., Scott, M. P., Kahn, P. C., Storch, J. 2015; 290 (45): 27321-27331

  Abstract

  The cholesterol storage disorder Niemann-Pick type C (NPC) disease is caused by defects in either of two late endosomal/lysosomal proteins, NPC1 and NPC2. NPC2 is a 16-kDa soluble protein that binds cholesterol in a 1:1 stoichiometry and can transfer cholesterol between membranes by a mechanism that involves protein-membrane interactions. To examine the structural basis of NPC2 function in cholesterol trafficking, a series of point mutations were generated across the surface of the protein. Several NPC2 mutants exhibited deficient sterol transport properties in a set of fluorescence-based assays. Notably, these mutants were also unable to promote egress of accumulated intracellular cholesterol from npc2(-/-) fibroblasts. The mutations mapped to several regions on the protein surface, suggesting that NPC2 can bind to more than one membrane simultaneously. Indeed, we have previously demonstrated that WT NPC2 promotes vesicle-vesicle interactions. These interactions were abrogated, however, by mutations causing defective sterol transfer properties. Molecular modeling shows that NPC2 is highly plastic, with several intense positively charged regions across the surface that could interact favorably with negatively charged membrane phospholipids. The point mutations generated in this study caused changes in NPC2 surface charge distribution with minimal conformational changes. The plasticity, coupled with membrane flexibility, probably allows for multiple cholesterol transfer routes. Thus, we hypothesize that, in part, NPC2 rapidly traffics cholesterol between closely appositioned membranes within the multilamellar interior of late endosomal/lysosomal proteins, ultimately effecting cholesterol egress from this compartment.

  View details for DOI 10.1074/jbc.M115.667469

  View details for Web of Science ID 000364794000039

  View details for PubMedID 26296895

  View details for PubMedCentralID PMC4646368

• AMP-Activated Protein Kinase Directly Phosphorylates and Destabilizes Hedgehog Pathway Transcription Factor GLI1 in Medulloblastoma. Cell reports Li, Y., Luo, J., Mosley, Y. C., Hedrick, V. E., Paul, L. N., Chang, J., Zhang, G., Wang, Y., Banko, M. R., Brunet, A., Kuang, S., Wu, J., Chang, C., Scott, M. P., Yang, J. 2015; 12 (4): 599-609

  Abstract

  The Hedgehog (Hh) pathway regulates cell differentiation and proliferation during development by controlling the Gli transcription factors. Cell fate decisions and progression toward organ and tissue maturity must be coordinated, and how an energy sensor regulates the Hh pathway is not clear. AMP-activated protein kinase (AMPK) is an important sensor of energy stores and controls protein synthesis and other energy-intensive processes. AMPK is directly responsive to intracellular AMP levels, inhibiting a wide range of cell activities if ATP is low and AMP is high. Thus, AMPK can affect development by influencing protein synthesis and other processes needed for growth and differentiation. Activation of AMPK reduces GLI1 protein levels and stability, thus blocking Sonic-hedgehog-induced transcriptional activity. AMPK phosphorylates GLI1 at serines 102 and 408 and threonine 1074. Mutation of these three sites into alanine prevents phosphorylation by AMPK. This leads to increased GLI1 protein stability, transcriptional activity, and oncogenic potency.

  View details for DOI 10.1016/j.celrep.2015.06.054

  View details for PubMedID 26190112

• Single-molecule imaging of Hedgehog pathway protein Smoothened in primary cilia reveals binding events regulated by Patched1. Proceedings of the National Academy of Sciences of the United States of America Milenkovic, L., Weiss, L. E., Yoon, J., Roth, T. L., Su, Y. S., Sahl, S. J., Scott, M. P., Moerner, W. E. 2015; 112 (27): 8320-8325

  Abstract

  Accumulation of the signaling protein Smoothened (Smo) in the membrane of primary cilia is an essential step in Hedgehog (Hh) signal transduction, yet the molecular mechanisms of Smo movement and localization are poorly understood. Using ultrasensitive single-molecule tracking with high spatial/temporal precision (30 nm/10 ms), we discovered that binding events disrupt the primarily diffusive movement of Smo in cilia at an array of sites near the base. The affinity of Smo for these binding sites was modulated by the Hh pathway activation state. Activation, by either a ligand or genetic loss of the negatively acting Hh receptor Patched-1 (Ptch), reduced the affinity and frequency of Smo binding at the base. Our findings quantify activation-dependent changes in Smo dynamics in cilia and highlight a previously unknown step in Hh pathway activation.

  View details for DOI 10.1073/pnas.1510094112

  View details for PubMedID 26100903

  View details for PubMedCentralID PMC4500289

• The Eya1 Phosphatase Promotes Shh Signaling during Hindbrain Development and Oncogenesis DEVELOPMENTAL CELL Eisner, A., Pazyra-Murphy, M. F., Durresi, E., Zhou, P., Zhao, X., Chadwick, E. C., Xu, P., Hillman, R. T., Scott, M. P., Greenberg, M. E., Segal, R. A. 2015; 33 (1): 22-35

  Abstract

  Sonic hedgehog (Shh) signaling is critical in development and oncogenesis, but the mechanisms regulating this pathway remain unclear. Although protein phosphorylation clearly affects Shh signaling, little is known about phosphatases governing the pathway. Here, we conducted a small hairpin RNA (shRNA) screen of the phosphatome and identified Eya1 as a positive regulator of Shh signaling. We find that the catalytically active phosphatase Eya1 cooperates with the DNA-binding protein Six1 to promote gene induction in response to Shh and that Eya1/Six1 together regulate Gli transcriptional activators. We show that Eya1, which is mutated in a human deafness disorder, branchio-oto-renal syndrome, is critical for Shh-dependent hindbrain growth and development. Moreover, Eya1 drives the growth of medulloblastoma, a Shh-dependent hindbrain tumor. Together, these results identify Eya1 and Six1 as key components of the Shh transcriptional network in normal development and in oncogenesis.

  View details for DOI 10.1016/j.devcel.2015.01.033

  View details for Web of Science ID 000352454200004

  View details for PubMedID 25816987

  View details for PubMedCentralID PMC4418443

• Phosphodiesterase 4D acts downstream of Neuropilin to control Hedgehog signal transduction and the growth of medulloblastoma. eLife Ge, X., Milenkovic, L., Suyama, K., Hartl, T., Purzner, T., Winans, A., Meyer, T., Scott, M. P. 2015; 4

  Abstract

  Alterations in Hedgehog (Hh) signaling lead to birth defects and cancers including medulloblastoma, the most common pediatric brain tumor. Although inhibitors targeting the membrane protein Smoothened suppress Hh signaling, acquired drug resistance and tumor relapse call for additional therapeutic targets. Here we show that phosphodiesterase 4D (PDE4D) acts downstream of Neuropilins to control Hh transduction and medulloblastoma growth. PDE4D interacts directly with Neuropilins, positive regulators of Hh pathway. The Neuropilin ligand Semaphorin3 enhances this interaction, promoting PDE4D translocation to the plasma membrane and cAMP degradation. The consequent inhibition of protein kinase A (PKA) enhances Hh transduction. In the developing cerebellum, genetic removal of Neuropilins reduces Hh signaling activity and suppresses proliferation of granule neuron precursors. In mouse medulloblastoma allografts, PDE4D inhibitors suppress Hh transduction and inhibit tumor growth. Our findings reveal a new regulatory mechanism of Hh transduction, and highlight PDE4D as a promising target to treat Hh-related tumors.

  View details for DOI 10.7554/eLife.07068

  View details for PubMedID 26371509

  View details for PubMedCentralID PMC4569902

• Phosphodiesterase 4D acts downstream of Neuropilin to control Hedgehog signal transduction and the growth of medulloblastoma. eLife Ge, X., Milenkovic, L., Suyama, K., Hartl, T., Purzner, T., Winans, A., Meyer, T., Scott, M. P. 2015; 4

  Abstract

  Alterations in Hedgehog (Hh) signaling lead to birth defects and cancers including medulloblastoma, the most common pediatric brain tumor. Although inhibitors targeting the membrane protein Smoothened suppress Hh signaling, acquired drug resistance and tumor relapse call for additional therapeutic targets. Here we show that phosphodiesterase 4D (PDE4D) acts downstream of Neuropilins to control Hh transduction and medulloblastoma growth. PDE4D interacts directly with Neuropilins, positive regulators of Hh pathway. The Neuropilin ligand Semaphorin3 enhances this interaction, promoting PDE4D translocation to the plasma membrane and cAMP degradation. The consequent inhibition of protein kinase A (PKA) enhances Hh transduction. In the developing cerebellum, genetic removal of Neuropilins reduces Hh signaling activity and suppresses proliferation of granule neuron precursors. In mouse medulloblastoma allografts, PDE4D inhibitors suppress Hh transduction and inhibit tumor growth. Our findings reveal a new regulatory mechanism of Hh transduction, and highlight PDE4D as a promising target to treat Hh-related tumors.

  View details for DOI 10.7554/eLife.07068

  View details for PubMedID 26371509

  View details for PubMedCentralID PMC4569902

• A Rapid and Simple Method for DNA Engineering Using Cycled Ligation Assembly PLOS ONE Roth, T. L., Milenkovic, L., Scott, M. P. 2014; 9 (9)

  Abstract

  DNA assembly techniques have developed rapidly, enabling efficient construction of complex constructs that would be prohibitively difficult using traditional restriction-digest based methods. Most of the recent methods for assembling multiple DNA fragments in vitro suffer from high costs, complex set-ups, and diminishing efficiency when used for more than a few DNA segments. Here we present a cycled ligation-based DNA assembly protocol that is simple, cheap, efficient, and powerful. The method employs a thermostable ligase and short Scaffold Oligonucleotide Connectors (SOCs) that are homologous to the ends and beginnings of two adjacent DNA sequences. These SOCs direct an exponential increase in the amount of correctly assembled product during a reaction that cycles between denaturing and annealing/ligating temperatures. Products of early cycles serve as templates for later cycles, allowing the assembly of many sequences in a single reaction. To demonstrate the method's utility, we directed the assembly of twelve inserts, in one reaction, into a transformable plasmid. All the joints were precise, and assembly was scarless in the sense that no nucleotides were added or missing at junctions. Simple, efficient, and low-cost cycled ligation assemblies will facilitate wider use of complex genetic constructs in biomedical research.

  View details for DOI 10.1371/journal.pone.0107329

  View details for Web of Science ID 000344317700045

  View details for PubMedID 25226397

  View details for PubMedCentralID PMC4167330

• Wing tips: The wing disc as a platform for studying Hedgehog signaling. Methods Hartl, T. A., Scott, M. P. 2014; 68 (1): 199-206

  Abstract

  Hedgehog (Hh) signal transduction is necessary for the development of most mammalian tissues and can go awry and cause birth defects or cancer. Hh signaling was initially described in Drosophila, and much of what we know today about mammalian Hh signaling was directly guided by discoveries in the fly. Indeed, Hh signaling is a wonderful example of the use of non-vertebrate model organisms to make basic discoveries that lead to new disease treatment. The first pharmaceutical to treat hyperactive Hh signaling in Basal Cell Carcinoma was released in 2012, approximately 30years after the isolation of Hh mutants in Drosophila. The study of Hh signaling has been greatly facilitated by the imaginal wing disc, a tissue with terrific experimental advantages. Studies using the wing disc have led to an understanding of Hh ligand processing, packaging into particles for transmission, secretion, reception, signal transduction, target gene activation, and tissue patterning. Here we describe the imaginal wing disc, how Hh patterns this tissue, and provide methods to use wing discs to study Hh signaling in Drosophila. The tools and approaches we highlight form the cornerstone of research efforts in many laboratories that use Drosophila to study Hh signaling, and are essential for ongoing discoveries.

  View details for DOI 10.1016/j.ymeth.2014.02.002

  View details for PubMedID 24556557

• Insight into insulin secretion from transcriptome and genetic analysis of insulin-producing cells of Drosophila. Genetics Cao, J., Ni, J., Ma, W., Shiu, V., Milla, L. A., Park, S., Spletter, M. L., Tang, S., Zhang, J., Wei, X., Kim, S. K., Scott, M. P. 2014; 197 (1): 175-192

  Abstract

  Insulin-producing cells (IPCs) in the Drosophila brain produce and release insulin-like peptides (ILPs) to the hemolymph. ILPs are crucial for growth and regulation of metabolic activity in flies, functions analogous to those of mammalian insulin and insulin-like growth factors (IGFs). To identify components functioning in IPCs to control ILP production, we employed genomic and candidate gene approaches. We used laser microdissection and messenger RNA sequencing to characterize the transcriptome of larval IPCs. IPCs highly express many genes homologous to genes active in insulin-producing β-cells of the mammalian pancreas. The genes in common encode ILPs and proteins that control insulin metabolism, storage, secretion, β-cell proliferation, and some not previously linked to insulin production or β-cell function. Among these novelties is unc-104, a kinesin 3 family gene, which is more highly expressed in IPCs compared to most other neurons. Knockdown of unc-104 in IPCs impaired ILP secretion and reduced peripheral insulin signaling. Unc-104 appears to transport ILPs along axons. As a complementary approach, we tested dominant-negative Rab genes to find Rab proteins required in IPCs for ILP production or secretion. Rab1 was identified as crucial for ILP trafficking in IPCs. Inhibition of Rab1 in IPCs increased circulating sugar levels, delayed development, and lowered weight and body size. Immunofluorescence labeling of Rab1 showed its tight association with ILP2 in the Golgi of IPCs. Unc-104 and Rab1 join other proteins required for ILP transport in IPCs.

  View details for DOI 10.1534/genetics.113.160663

  View details for PubMedID 24558258

  View details for PubMedCentralID PMC4012477

• Single-Molecule Tracking of Smoothened in the Primary Cilium Weiss, L. E., Milenkovic, L., Sahl, S. J., Roth, T. L., Scott, M. P., Moerner, W. E. CELL PRESS. 2014: 20A

  View details for DOI 10.1016/j.bpj.2013.11.166

  View details for Web of Science ID 000337000400103

• Neuropilin-2 contributes to tumorigenicity in a mouse model of Hedgehog pathway medulloblastoma JOURNAL OF NEURO-ONCOLOGY Gephart, M. G., Su, Y. S., Bandara, S., Tsai, F., Hong, J., Conley, N., Rayburn, H., Milenkovic, L., Meyer, T., Scott, M. P. 2013; 115 (2): 161-168

  Abstract

  The Hedgehog (Hh) signaling pathway has been implicated in the most common childhood brain tumor, medulloblastoma (MB). Given the toxicity of post-surgical treatments for MB, continued need exists for new, targeted therapies. Based upon our finding that Neuropilin (Nrp) transmembrane proteins are required for Hh signal transduction, we investigated the role of Nrp in MB cells. Cultured cells derived from a mouse Ptch (+/-) ;LacZ MB (Med1-MB), effectively modeled the Hh pathway-related subcategory of human MBs in vitro. Med1-MB cells maintained constitutively active Hh target gene transcription, and consistently formed tumors within one month after injection into mouse cerebella. The proliferation rate of Med1-MBs in culture was dependent upon Nrp2, while reducing Nrp1 function had little effect. Knockdown of Nrp2 prior to cell implantation significantly increased mouse survival, compared to transfection with a non-targeting siRNA. Knocking down Nrp2 specifically in MB cells avoided any direct effect on tumor vascularization. Nrp2 should be further investigated as a potential target for adjuvant therapy in patients with MB.

  View details for DOI 10.1007/s11060-013-1216-1

  View details for Web of Science ID 000325821900004

• AN ENGINEERED PEPTIDE FOR NON-INVASIVE OPTICAL IMAGING OF BRAIN TUMORS Moore, S., Hayden-Gephart, M., Bergen, J., Su, Y., Rayburn, H., Edwards, M., Scott, M., Cochran, J. OXFORD UNIV PRESS INC. 2013: 40–41

  View details for Web of Science ID 000327456200170

• Neuropilin-2 contributes to tumorigenicity in a mouse model of Hedgehog pathway medulloblastoma. Journal of neuro-oncology Hayden Gephart, M. G., Su, Y. S., Bandara, S., Tsai, F., Hong, J., Conley, N., Rayburn, H., Milenkovic, L., Meyer, T., Scott, M. P. 2013; 115 (2): 161-168

  Abstract

  The Hedgehog (Hh) signaling pathway has been implicated in the most common childhood brain tumor, medulloblastoma (MB). Given the toxicity of post-surgical treatments for MB, continued need exists for new, targeted therapies. Based upon our finding that Neuropilin (Nrp) transmembrane proteins are required for Hh signal transduction, we investigated the role of Nrp in MB cells. Cultured cells derived from a mouse Ptch (+/-) ;LacZ MB (Med1-MB), effectively modeled the Hh pathway-related subcategory of human MBs in vitro. Med1-MB cells maintained constitutively active Hh target gene transcription, and consistently formed tumors within one month after injection into mouse cerebella. The proliferation rate of Med1-MBs in culture was dependent upon Nrp2, while reducing Nrp1 function had little effect. Knockdown of Nrp2 prior to cell implantation significantly increased mouse survival, compared to transfection with a non-targeting siRNA. Knocking down Nrp2 specifically in MB cells avoided any direct effect on tumor vascularization. Nrp2 should be further investigated as a potential target for adjuvant therapy in patients with MB.

  View details for DOI 10.1007/s11060-013-1216-1

  View details for PubMedID 24026530

• Engineered knottin peptide enables noninvasive optical imaging of intracranial medulloblastoma. Proceedings of the National Academy of Sciences of the United States of America Moore, S. J., Hayden Gephart, M. G., Bergen, J. M., Su, Y. S., Rayburn, H., Scott, M. P., Cochran, J. R. 2013; 110 (36): 14598-14603

  Abstract

  Central nervous system tumors carry grave clinical prognoses due to limited effectiveness of surgical resection, radiation, and chemotherapy. Thus, improved strategies for brain tumor visualization and targeted treatment are critically needed. We demonstrate that mouse cerebellar medulloblastoma (MB) can be targeted and illuminated with a fluorescent, engineered cystine knot (knottin) peptide that binds with high affinity to αvβ3, αvβ5, and α5β1 integrin receptors. This integrin-binding knottin peptide, denoted EETI 2.5F, was evaluated as a molecular imaging probe in both orthotopic and genetic models of MB. Following tail vein injection, fluorescence arising from dye-conjugated EETI 2.5F was localized to the tumor compared with the normal surrounding brain tissue, as measured by optical imaging. The imaging signal intensity correlated with tumor volume. Due to its unique ability to bind to α5β1 integrin, EETI 2.5F showed superior in vivo and ex vivo brain tumor imaging contrast compared with other engineered integrin-binding knottin peptides and with c(RGDfK), a well-studied integrin-binding peptidomimetic. Next, EETI 2.5F was fused to an antibody fragment crystallizable (Fc) domain (EETI 2.5F-Fc) to determine if a larger integrin-binding protein could also target intracranial brain tumors. EETI 2.5F-Fc, conjugated to a fluorescent dye, illuminated MB following i.v. injection and was able to distribute throughout the tumor parenchyma. In contrast, brain tumor imaging signals were not detected in mice injected with EETI 2.5F proteins containing a scrambled integrin-binding sequence, demonstrating the importance of target specificity. These results highlight the potential of using EETI 2.5F and EETI 2.5-Fc as targeted molecular probes for brain tumor imaging.

  View details for DOI 10.1073/pnas.1311333110

  View details for PubMedID 23950221

  View details for PubMedCentralID PMC3767496

• Engineered knottin peptide enables noninvasive optical imaging of intracranial medulloblastoma. Proceedings of the National Academy of Sciences of the United States of America Moore, S. J., Hayden Gephart, M. G., Bergen, J. M., Su, Y. S., Rayburn, H., Scott, M. P., Cochran, J. R. 2013; 110 (36): 14598-14603

  Abstract

  Central nervous system tumors carry grave clinical prognoses due to limited effectiveness of surgical resection, radiation, and chemotherapy. Thus, improved strategies for brain tumor visualization and targeted treatment are critically needed. We demonstrate that mouse cerebellar medulloblastoma (MB) can be targeted and illuminated with a fluorescent, engineered cystine knot (knottin) peptide that binds with high affinity to αvβ3, αvβ5, and α5β1 integrin receptors. This integrin-binding knottin peptide, denoted EETI 2.5F, was evaluated as a molecular imaging probe in both orthotopic and genetic models of MB. Following tail vein injection, fluorescence arising from dye-conjugated EETI 2.5F was localized to the tumor compared with the normal surrounding brain tissue, as measured by optical imaging. The imaging signal intensity correlated with tumor volume. Due to its unique ability to bind to α5β1 integrin, EETI 2.5F showed superior in vivo and ex vivo brain tumor imaging contrast compared with other engineered integrin-binding knottin peptides and with c(RGDfK), a well-studied integrin-binding peptidomimetic. Next, EETI 2.5F was fused to an antibody fragment crystallizable (Fc) domain (EETI 2.5F-Fc) to determine if a larger integrin-binding protein could also target intracranial brain tumors. EETI 2.5F-Fc, conjugated to a fluorescent dye, illuminated MB following i.v. injection and was able to distribute throughout the tumor parenchyma. In contrast, brain tumor imaging signals were not detected in mice injected with EETI 2.5F proteins containing a scrambled integrin-binding sequence, demonstrating the importance of target specificity. These results highlight the potential of using EETI 2.5F and EETI 2.5-Fc as targeted molecular probes for brain tumor imaging.

  View details for DOI 10.1073/pnas.1311333110

  View details for PubMedID 23950221

  View details for PubMedCentralID PMC3767496

• Genetic dissection of a cell-autonomous neurodegenerative disorder: lessons learned from mouse models of Niemann-Pick disease type C DISEASE MODELS & MECHANISMS Lopez, M. E., Scott, M. P. 2013; 6 (5): 1089-1100

  Abstract

  Understanding neurodegenerative disease progression and its treatment requires the systematic characterization and manipulation of relevant cell types and molecular pathways. The neurodegenerative lysosomal storage disorder Niemann-Pick disease type C (NPC) is highly amenable to genetic approaches that allow exploration of the disease biology at the organismal, cellular and molecular level. Although NPC is a rare disease, genetic analysis of the associated neuropathology promises to provide insight into the logic of disease neural circuitry, selective neuron vulnerability and neural-glial interactions. The ability to control the disorder cell-autonomously and in naturally occurring spontaneous animal models that recapitulate many aspects of the human disease allows for an unparalleled dissection of the disease neurobiology in vivo. Here, we review progress in mouse-model-based studies of NPC disease, specifically focusing on the subtype that is caused by a deficiency in NPC1, a sterol-binding late endosomal membrane protein involved in lipid trafficking. We also discuss recent findings and future directions in NPC disease research that are pertinent to understanding the cellular and molecular mechanisms underlying neurodegeneration in general.

  View details for DOI 10.1242/dmm.012385

  View details for Web of Science ID 000325789600005

  View details for PubMedID 23907005

  View details for PubMedCentralID PMC3759329

• Kif3a is necessary for initiation and maintenance of medulloblastoma. Carcinogenesis Barakat, M. T., Humke, E. W., Scott, M. P. 2013; 34 (6): 1382-1392

  Abstract

  Medulloblastoma (MB) cells arise from granule neuron precursors (GNPs) that have lost growth control. During normal development, GNPs divide in response to Sonic hedgehog (SHH), a ligand that binds to the patched (PTCH) receptor on GNPs. If one copy of the Ptch gene is lost, as in human Gorlin's syndrome and in Ptch(+/-) mice, MBs may form. Proper transduction of the SHH signal critically depends on primary cilia. Loss of primary cilia results in improper signal reception and failure to properly activate SHH target genes. KIF3a, part of a kinesin motor, is required for formation of primary cilia. Here, we use tamoxifen-induced ablation of Kif3a in GNPs of postnatal Ptch(+/-) mouse cerebella to show that KIF3a is necessary for MB formation. To investigate the importance of primary cilia in established tumors, we deleted Kif3a from cultured cells and from tumor cell grafts. The loss of Kif3a from established tumors led to their growth arrest and regression. MBs behave as if they are addicted to the presence of primary cilia. These results underscore the potential utility of agents that disrupt cilia for the treatment of Hh pathway-related MBs.

  View details for DOI 10.1093/carcin/bgt041

  View details for PubMedID 23389290

  View details for PubMedCentralID PMC3670253

• Regulation of ribosome biogenesis by nucleostemin 3 promotes local and systemic growth in Drosophila. Genetics Hartl, T. A., Ni, J., Cao, J., Suyama, K. L., Patchett, S., Bussiere, C., Gui, D. Y., Tang, S., Kaplan, D. D., Fish, M., Johnson, A. W., Scott, M. P. 2013; 194 (1): 101-115

  Abstract

  Nucleostemin 3 (NS3) is an evolutionarily conserved protein with profound roles in cell growth and viability. Here we analyze cell-autonomous and non-cell-autonomous growth control roles of NS3 in Drosophila and demonstrate its GTPase activity using genetic and biochemical assays. Two null alleles of ns3, and RNAi, demonstrate the necessity of NS3 for cell autonomous growth. A hypomorphic allele highlights the hypersensitivity of neurons to lowered NS3 function. We propose that NS3 is the functional ortholog of yeast and human Lsg1, which promotes release of the nuclear export adapter from the large ribosomal subunit. Release of the adapter and its recycling to the nucleus are essential for sustained production of ribosomes. The ribosome biogenesis role of NS3 is essential for proper rates of translation in all tissues and is necessary for functions of growth-promoting neurons.

  View details for DOI 10.1534/genetics.112.149104

  View details for PubMedID 23436180

  View details for PubMedCentralID PMC3632458

• Novel method for the scarless single-step assembly of many small DNA sequences Roth, T., Milenkovic, L., Scott, M. FEDERATION AMER SOC EXP BIOL. 2013

  View details for Web of Science ID 000319883501419

• The chromatin remodeling factor Chd1l is required in the preimplantation embryo. Biology open Snider, A. C., Leong, D., Wang, Q. T., Wysocka, J., Yao, M. W., Scott, M. P. 2013; 2 (2): 121-131

  Abstract

  During preimplantation development, the embryo must establish totipotency and enact the earliest differentiation choices, processes that involve extensive chromatin modification. To identify novel developmental regulators, we screened for genes that are preferentially transcribed in the pluripotent inner cell mass (ICM) of the mouse blastocyst. Genes that encode chromatin remodeling factors were prominently represented in the ICM, including Chd1l, a member of the Snf2 gene family. Chd1l is developmentally regulated and expressed in embryonic stem (ES) cells, but its role in development has not been investigated. Here we show that inhibiting Chd1l protein production by microinjection of antisense morpholinos causes arrest prior to the blastocyst stage. Despite this important function in vivo, Chd1l is non-essential for cultured ES cell survival, pluripotency, or differentiation, suggesting that Chd1l is vital for events in embryos that are distinct from events in ES cells. Our data reveal a novel role for the chromatin remodeling factor Chd1l in the earliest cell divisions of mammalian development.

  View details for DOI 10.1242/bio.20122949

  View details for PubMedID 23429299

  View details for PubMedCentralID PMC3575647

• The chromatin remodeling factor Chd1l is required in the preimplantation embryo BIOLOGY OPEN Snider, A. C., Leong, D., Wang, Q. T., Wysocka, J., Yao, M. W., Scott, M. P. 2013; 2 (2): 121-131

  Abstract

  During preimplantation development, the embryo must establish totipotency and enact the earliest differentiation choices, processes that involve extensive chromatin modification. To identify novel developmental regulators, we screened for genes that are preferentially transcribed in the pluripotent inner cell mass (ICM) of the mouse blastocyst. Genes that encode chromatin remodeling factors were prominently represented in the ICM, including Chd1l, a member of the Snf2 gene family. Chd1l is developmentally regulated and expressed in embryonic stem (ES) cells, but its role in development has not been investigated. Here we show that inhibiting Chd1l protein production by microinjection of antisense morpholinos causes arrest prior to the blastocyst stage. Despite this important function in vivo, Chd1l is non-essential for cultured ES cell survival, pluripotency, or differentiation, suggesting that Chd1l is vital for events in embryos that are distinct from events in ES cells. Our data reveal a novel role for the chromatin remodeling factor Chd1l in the earliest cell divisions of mammalian development.

  View details for DOI 10.1242/bio.20122949

  View details for Web of Science ID 000209205900003

  View details for PubMedCentralID PMC3575647

• Genetic dissection of a cell-autonomous neurodegenerative disorder: Lessons learned from mouse models of Niemann-Pick disease type C Disease Models & Mechanisms Lopez, M. E., Scott, M. P. 2013; 6: 1-12
• Microtubules Enable the Planar Cell Polarity of Airway Cilia CURRENT BIOLOGY Vladar, E. K., Bayly, R. D., Sangoram, A. M., Scott, M. P., Axelrod, J. D. 2012; 22 (23): 2203-2212

  Abstract

  Airway cilia must be physically oriented along the longitudinal tissue axis for concerted, directional motility that is essential for proper mucociliary clearance.We show that planar cell polarity (PCP) signaling specifies directionality and orients respiratory cilia. Within all airway epithelial cells, a conserved set of PCP proteins shows interdependent, asymmetric junctional localization; nonautonomous signaling coordinates polarization between cells; and a polarized microtubule (MT) network is likely required for asymmetric PCP protein localization. We find that basal bodies dock after polarity of PCP proteins is established and are polarized nearly simultaneously, and that refinement of basal body/cilium orientation continues during airway epithelial development. Unique to mature multiciliated cells, we identify PCP-regulated, planar polarized MTs that originate from basal bodies and interact, via their plus ends, with membrane domains associated with the PCP proteins Frizzled and Dishevelled. Disruption of MTs leads to misoriented cilia.A conserved PCP pathway orients airway cilia by communicating polarity information from asymmetric membrane domains at the apical junctions, through MTs, to orient the MT and actin-based network of ciliary basal bodies below the apical surface.

  View details for DOI 10.1016/j.cub.2012.09.046

  View details for Web of Science ID 000312115300016

  View details for PubMedID 23122850

  View details for PubMedCentralID PMC3518597

• Complement is dispensable for neurodegeneration in Niemann-Pick disease type C JOURNAL OF NEUROINFLAMMATION Lopez, M. E., Klein, A. D., Scott, M. P. 2012; 9

  Abstract

  The immune system has been implicated in neurodegeneration during development and disease. In various studies, the absence of complement (that is, C1q deficiency) impeded the elimination of apoptotic neurons, allowing survival. In the genetic lysosomal storage disease Niemann-Pick C (NPC), caused by loss of NPC1 function, the expression of complement system components, C1q especially, is elevated in degenerating brain regions of Npc1-/- mice. Here we test whether complement is mediating neurodegeneration in NPC disease.In normal mature mice, C1q mRNA was found in neurons, particularly cerebellar Purkinje neurons (PNs). In Npc1-/- mice, C1q mRNA was additionally found in activated microglia, which accumulate during disease progression and PN loss. Interestingly, C1q was not enriched on or near degenerating neurons. Instead, C1q was concentrated in other brain regions, where it partially co-localized with a potential C1q inhibitor, chondroitin sulfate proteoglycan (CSPG). Genetic deletion of C1q, or of the downstream complement pathway component C3, did not significantly alter patterned neuron loss or disease progression. Deletion of other immune response factors, a Toll-like receptor, a matrix metalloprotease, or the apoptosis facilitator BIM, also failed to alter neuron loss.We conclude that complement is not involved in the death and clearance of neurons in NPC disease. This study supports a view of neuroinflammation as a secondary response with non-causal relationship to neuron injury in the disease. This disease model may prove useful for understanding the conditions in which complement and immunity do contribute to neurodegeneration in other disorders.

  View details for DOI 10.1186/1742-2094-9-216

  View details for Web of Science ID 000311733400001

  View details for PubMedID 22985423

  View details for PubMedCentralID PMC3511250

• Neuronal and epithelial cell rescue resolves chronic systemic inflammation in the lipid storage disorder Niemann-Pick C HUMAN MOLECULAR GENETICS Lopez, M. E., Klein, A. D., Hong, J., Dimbil, U. J., Scott, M. P. 2012; 21 (13): 2946-2960

  Abstract

  Chronic systemic inflammation is thought to be a major contributor to metabolic and neurodegenerative diseases. Since inflammatory components are shared among different disorders, targeting inflammation is an attractive option for mitigating disease. To test the significance of inflammation in the lipid storage disorder (LSD) Niemann-Pick C (NPC), we deleted the macrophage inflammatory gene Mip1a/Ccl3 from NPC diseased mice. Deletion of Ccl3 had been reported to delay neuronal loss in Sandhoff LSD mice by inhibiting macrophage infiltration. For NPC mice, in contrast, deleting Ccl3 did not retard neurodegeneration and worsened the clinical outcome. Depletion of visceral tissue macrophages also did not alter central nervous system (CNS) pathology and instead increased liver injury, suggesting a limited macrophage infiltration response into the CNS and a beneficial role of macrophage activity in visceral tissue. Prevention of neuron loss or liver injury, even at late stages in the disease, was achieved through specific rescue of NPC disease in neurons or in liver epithelial cells, respectively. Local epithelial cell correction was also sufficient to reduce the macrophage-associated pathology in lung tissue. These results demonstrate that elevated inflammation and macrophage activity does not necessarily contribute to neurodegeneration and tissue injury, and LSD defects in immune cells may not preclude an appropriate inflammatory response. We conclude that inflammation remains secondary to neuronal and epithelial cell dysfunction and does not irreversibly contribute to the pathogenic cascade in NPC disease. Without further exploration of possible beneficial roles of inflammatory mediators, targeting inflammation may not be therapeutically effective at ameliorating disease severity.

  View details for DOI 10.1093/hmg/dds126

  View details for Web of Science ID 000305457700010

  View details for PubMedID 22493001

  View details for PubMedCentralID PMC3373242

• Interactions between Type III receptor tyrosine phosphatases and growth factor receptor tyrosine kinases regulate tracheal tube formation in Drosophila. Biology open Jeon, M., Scott, M. P., Zinn, K. 2012; 1 (6): 548-558

  Abstract

  The respiratory (tracheal) system of the Drosophila melanogaster larva is an intricate branched network of air-filled tubes. Its developmental logic is similar in some ways to that of the vertebrate vascular system. We previously described a unique embryonic tracheal tubulogenesis phenotype caused by loss of both of the Type III receptor tyrosine phosphatases (RPTPs), Ptp4E and Ptp10D. In Ptp4E Ptp10D double mutants, the linear tubes in unicellular and terminal tracheal branches are converted into bubble-like cysts that incorporate apical cell surface markers. This tube geometry phenotype is modulated by changes in the activity or expression of the epidermal growth factor receptor (Egfr) tyrosine kinase (TK). Ptp10D physically interacts with Egfr. Here we demonstrate that the Ptp4E Ptp10D phenotype is the consequence of the loss of negative regulation by the RPTPs of three growth factor receptor TKs: Egfr, Breathless and Pvr. Reducing the activity of any of the three kinases by tracheal expression of dominant-negative mutants suppresses cyst formation. By competing dominant-negative and constitutively active kinase mutants against each other, we show that the three RTKs have partially interchangeable activities, so that increasing the activity of one kinase can compensate for the effects of reducing the activity of another. This implies that SH2-domain downstream effectors that are required for the phenotype are likely to be able to interact with phosphotyrosine sites on all three receptor TKs. We also show that the phenotype involves increases in signaling through the MAP kinase and Rho GTPase pathways.

  View details for DOI 10.1242/bio.2012471

  View details for PubMedID 23213447

  View details for PubMedCentralID PMC3509443

• Interactions between Type III receptor tyrosine phosphatases and growth factor receptor tyrosine kinases regulate tracheal tube formation in Drosophila BIOLOGY OPEN Jeon, M., Scott, M. P., Zinn, K. 2012; 1 (6): 548-558

  Abstract

  The respiratory (tracheal) system of the Drosophila melanogaster larva is an intricate branched network of air-filled tubes. Its developmental logic is similar in some ways to that of the vertebrate vascular system. We previously described a unique embryonic tracheal tubulogenesis phenotype caused by loss of both of the Type III receptor tyrosine phosphatases (RPTPs), Ptp4E and Ptp10D. In Ptp4E Ptp10D double mutants, the linear tubes in unicellular and terminal tracheal branches are converted into bubble-like cysts that incorporate apical cell surface markers. This tube geometry phenotype is modulated by changes in the activity or expression of the epidermal growth factor receptor (Egfr) tyrosine kinase (TK). Ptp10D physically interacts with Egfr. Here we demonstrate that the Ptp4E Ptp10D phenotype is the consequence of the loss of negative regulation by the RPTPs of three growth factor receptor TKs: Egfr, Breathless and Pvr. Reducing the activity of any of the three kinases by tracheal expression of dominant-negative mutants suppresses cyst formation. By competing dominant-negative and constitutively active kinase mutants against each other, we show that the three RTKs have partially interchangeable activities, so that increasing the activity of one kinase can compensate for the effects of reducing the activity of another. This implies that SH2-domain downstream effectors that are required for the phenotype are likely to be able to interact with phosphotyrosine sites on all three receptor TKs. We also show that the phenotype involves increases in signaling through the MAP kinase and Rho GTPase pathways.

  View details for DOI 10.1242/bio.2012471

  View details for Web of Science ID 000209205100005

  View details for PubMedCentralID PMC3509443

• Molecular Cell Biology, 7th Edition Lodish, H.F., A.J. Berk, C.A. Kaiser, M. Krieger, A. Bretscher, H.L. Ploegh, A. Amon, M.P. Scott 2012
• Neuropilins are positive regulators of Hedgehog signal transduction GENES & DEVELOPMENT Hillman, R. T., Feng, B. Y., Ni, J., Woo, W., Milenkovic, L., Gephart, M. G., Teruel, M. N., Oro, A. E., Chen, J. K., Scott, M. P. 2011; 25 (22): 2333-2346

  Abstract

  The Hedgehog (Hh) pathway is essential for vertebrate embryogenesis, and excessive Hh target gene activation can cause cancer in humans. Here we show that Neuropilin 1 (Nrp1) and Nrp2, transmembrane proteins with roles in axon guidance and vascular endothelial growth factor (VEGF) signaling, are important positive regulators of Hh signal transduction. Nrps are expressed at times and locations of active Hh signal transduction during mouse development. Using cell lines lacking key Hh pathway components, we show that Nrps mediate Hh transduction between activated Smoothened (Smo) protein and the negative regulator Suppressor of Fused (SuFu). Nrp1 transcription is induced by Hh signaling, and Nrp1 overexpression increases maximal Hh target gene activation, indicating the existence of a positive feedback circuit. The regulation of Hh signal transduction by Nrps is conserved between mammals and bony fish, as we show that morpholinos targeting the Nrp zebrafish ortholog nrp1a produce a specific and highly penetrant Hh pathway loss-of-function phenotype. These findings enhance our knowledge of Hh pathway regulation and provide evidence for a conserved nexus between Nrps and this important developmental signaling system.

  View details for DOI 10.1101/gad.173054.111

  View details for PubMedID 22051878

• CANCER PATHWAY DISCOVERY IN A MOUSE MODEL OF MEDULLOBLASTOMA 16th Annual Scientific Meeting of the Society-for-Neuro-Oncology (SNO)/AANS/CNS Section on Tumors Gephart, M. H., Lee, E., Kyriazopoulou-Panagiotopoulou, S., Milenkovic, L., Xun, X., Hou, Y., Kui, W., Edwards, M., Batzoglou, S., Jun, W., Scott, M. OXFORD UNIV PRESS INC. 2011: 78–78

  View details for Web of Science ID 000297026600315

• Full p53 transcriptional activation potential is dispensable for tumor suppression in diverse lineages PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Jiang, D., Brady, C. A., Johnson, T. M., Lee, E. Y., Park, E. J., Scott, M. P., Attardi, L. D. 2011; 108 (41): 17123-17128

  Abstract

  Over half of all human cancers, of a wide variety of types, sustain mutations in the p53 tumor suppressor gene. Although p53 limits tumorigenesis through the induction of apoptosis or cell cycle arrest, its molecular mechanism of action in tumor suppression has been elusive. The best-characterized p53 activity in vitro is as a transcriptional activator, but the identification of numerous additional p53 biochemical activities in vitro has made it unclear which mechanism accounts for tumor suppression. Here, we assess the importance of transcriptional activation for p53 tumor suppression function in vivo in several tissues, using a knock-in mouse strain expressing a p53 mutant compromised for transcriptional activation, p53(25,26). p53(25,26) is severely impaired for the transactivation of numerous classical p53 target genes, including p21, Noxa, and Puma, but it retains the ability to activate a small subset of p53 target genes, including Bax. Surprisingly, p53(25,26) can nonetheless suppress tumor growth in cancers derived from the epithelial, mesenchymal, central nervous system, and lymphoid lineages. Therefore, full transactivation of most p53 target genes is dispensable for p53 tumor suppressor function in a range of tissue types. In contrast, a transcriptional activation mutant that is completely defective for transactivation, p53(25,26,53,54), fails to suppress tumor development. These findings demonstrate that transcriptional activation is indeed broadly critical for p53 tumor suppressor function, although this requirement reflects the limited transcriptional activity observed with p53(25,26) rather than robust transactivation of a full complement of p53 target genes.

  View details for DOI 10.1073/pnas.1111245108

  View details for Web of Science ID 000295973800051

  View details for PubMedID 21969549

  View details for PubMedCentralID PMC3193184

• Mechanisms of hedgehog target gene selection in development and cancer Yang, J., Lee, E. Y., Scott, M. P. AMER ASSOC CANCER RESEARCH. 2011

  View details for DOI 10.1158/1538-7445.AM2011-932

  View details for Web of Science ID 000209701305339

• Anatomically Defined Neuron-Based Rescue of Neurodegenerative Niemann-Pick Type C Disorder JOURNAL OF NEUROSCIENCE Lopez, M. E., Klein, A. D., Dimbil, U. J., Scott, M. P. 2011; 31 (12): 4367-4378

  Abstract

  Niemann-Pick type C disease is a fatal lysosomal storage disorder caused by loss of NPC1 function. The disorder severely affects multiple body systems, particularly the nervous system. To test whether rescue of NPC1 activity in neurons, astrocytes, or other cell types can correct the neurological defects, a Tet-inducible Npc1-YFP transgene was introduced into Npc1(-/-) mice for the cell type-specific rescue of NPC1 loss. NPC1-YFP produced in neurons prevented neuron degeneration, slowed reactive glial activity, and ameliorated the disease. NPC1-YFP produced in astrocytes or in cells of visceral tissue did not. These results suggest that loss of NPC1 activity from neurons is the primary cause of the neuropathology and that rescue of NPC1 function in neurons is sufficient to mitigate the disease. The ability of neurons to survive and function in a cell-autonomous fashion allowed the use of this newly engineered rescue system to further define the brain regions or neuron populations required to ameliorate a neurological symptom. NPC1-YFP produced specifically in cerebellar Purkinje neurons reduced ataxia, increased weight, and prolonged life, but it did not prevent the eventual decline and premature death of Npc1(-/-) mice. Significant increase in lifespan correlated with sustained reduction of inflammation in the thalamus. Neuron rescue of other forebrain areas provided little benefit. Future work targeting increasingly discrete neuronal networks should reveal which CNS areas are critical for survival. This work may have broad implications for understanding the anatomical and cellular basis of neurological signs and symptoms of other neurodegenerative and lysosomal disorders.

  View details for DOI 10.1523/JNEUROSCI.5981-10.2011

  View details for Web of Science ID 000288750700004

  View details for PubMedID 21430138

  View details for PubMedCentralID PMC3071647

• The cholesterol trafficking protein NPC1 is required for Drosophila spermatogenesis DEVELOPMENTAL BIOLOGY Wang, C., Ma, Z., Scott, M. P., Huang, X. 2011; 351 (1): 146-155

  Abstract

  Niemann-Pick C (NPC) disease is a lethal neurodegenerative disorder affecting cellular sterol trafficking. Besides neurodegeneration, NPC patients also exhibit other pleiotropic conditions, indicating that NPC protein is required for other physiological processes. Previous studies indicated that a sterol shortage that in turn leads to a shortage of steroid hormones (for example, ecdysone in Drosophila) is likely to be the cause of NPC disease pathology. We have shown that mutations in Drosophila npc1, one of the two NPC disease-related genes, leads to larval lethal and male infertility. Here, we reported that npc1 mutants are defective in spermatogenesis and in particular in the membrane-remodeling individualization process. Interestingly, we found that ecdysone, the steroid hormone responsible for the larval lethal phenotype in npc1 mutants, is not required for individualization. However, supplying 7-dehydrocholesterol can partially rescue the male infertility of npc1 mutants, suggesting that a sterol shortage is responsible for the spermatogenesis defects. In addition, the individualization defects of npc1 mutants were enhanced at high temperature, suggesting that the sterol shortage may lead to temperature-sensitive defects in the membrane-remodeling process. Together, our study reveals a sterol-dependent, ecdysone-independent mechanism of NPC1 function in Drosophila spermatogenesis.

  View details for DOI 10.1016/j.ydbio.2010.12.042

  View details for Web of Science ID 000287566100015

  View details for PubMedID 21215267

• Mutations in Prickle Orthologs Cause Seizures in Flies, Mice, and Humans AMERICAN JOURNAL OF HUMAN GENETICS Tao, H., Manak, J. R., Sowers, L., Mei, X., Kiyonari, H., Abe, T., Dandaleh, N. S., Yang, T., Wu, S., Chen, S., Fox, M. H., Gurnett, C., Montine, T., Bird, T., Shaffer, L. G., Rosenfeld, J. A., McConne, J., Madan-Khetarpal, S., Berry-Kravis, E., Griesbach, H., Saneto, R. P., Scott, M. P., Antic, D., Reed, J., Boland, R., Ehaideb, S. N., El-Shanti, H., Mahajan, V. B., Ferguson, P. J., Axelrod, J. D., Lehesjoki, A., Fritzsch, B., Slusarski, D. C., Wemmie, J., Ueno, N., Bassuk, A. G. 2011; 88 (2): 138-149

  Abstract

  Epilepsy is heritable, yet few causative gene mutations have been identified, and thus far no human epilepsy gene mutations have been found to produce seizures in invertebrates. Here we show that mutations in prickle genes are associated with seizures in humans, mice, and flies. We identified human epilepsy patients with heterozygous mutations in either PRICKLE1 or PRICKLE2. In overexpression assays in zebrafish, prickle mutations resulted in aberrant prickle function. A seizure phenotype was present in the Prickle1-null mutant mouse, two Prickle1 point mutant (missense and nonsense) mice, and a Prickle2-null mutant mouse. Drosophila with prickle mutations displayed seizures that were responsive to anti-epileptic medication, and homozygous mutant embryos showed neuronal defects. These results suggest that prickle mutations have caused seizures throughout evolution.

  View details for DOI 10.1016/j.ajhg.2010.12.012

  View details for Web of Science ID 000287684100002

  View details for PubMedID 21276947

  View details for PubMedCentralID PMC3035715

• Learning from Jekyll to control Hyde: Hedgehog signaling in development and cancer TRENDS IN MOLECULAR MEDICINE Barakat, M. T., Humke, E. W., Scott, M. P. 2010; 16 (8): 337-348

  Abstract

  The Hedgehog (Hh) cascade controls cell proliferation, differentiation and patterning of tissues during embryogenesis but is largely suppressed in the adult. The Hh pathway can become reactivated in cancer. Here, we assimilate data from recent studies to understand how and when the Hh pathway is turned on to aid the neoplastic process. Hh signaling is now known to have a role in established tumors, enabling categorization of tumors based on the role Hh signaling plays in their growth. This categorization has relevance for prognosis and targeted therapeutics. In the first category, abnormal Hh signaling initiates the tumor. In the second category, Hh signaling helps maintain the tumor. In the third category, Hh signaling is implicated but its role is not yet defined.

  View details for DOI 10.1016/j.molmed.2010.05.003

  View details for Web of Science ID 000281273500001

  View details for PubMedID 20696410

  View details for PubMedCentralID PMC3651907

• A Septin Diffusion Barrier at the Base of the Primary Cilium Maintains Ciliary Membrane Protein Distribution SCIENCE Hu, Q., Milenkovic, L., Jin, H., Scott, M. P., Nachury, M. V., Spiliotis, E. T., Nelson, W. J. 2010; 329 (5990): 436-439

  Abstract

  In animal cells, the primary cilium transduces extracellular signals through signaling receptors localized in the ciliary membrane, but how these ciliary membrane proteins are retained in the cilium is unknown. We found that ciliary membrane proteins were highly mobile, but their diffusion was impeded at the base of the cilium by a diffusion barrier. Septin 2 (SEPT2), a member of the septin family of guanosine triphosphatases that form a diffusion barrier in budding yeast, localized at the base of the ciliary membrane. SEPT2 depletion resulted in loss of ciliary membrane protein localization and Sonic hedgehog signal transduction, and inhibited ciliogenesis. Thus, SEPT2 is part of a diffusion barrier at the base of the ciliary membrane and is essential for retaining receptor-signaling pathways in the primary cilium.

  View details for DOI 10.1126/science.1191054

  View details for Web of Science ID 000280196500036

  View details for PubMedID 20558667

  View details for PubMedCentralID PMC3092790

• Role of Lipid Metabolism in Smoothened Derepression in Hedgehog Signaling DEVELOPMENTAL CELL Yavan, A., Nagaraj, R., Owusu-Ansah, E., Folick, A., Ngo, K., Hillman, T., Call, G., Rohatgi, R., Scott, M. P., Banerjee, U. 2010; 19 (1): 54-65

  Abstract

  The binding of Hedgehog (Hh) to its receptor Patched causes derepression of Smoothened (Smo), resulting in the activation of the Hh pathway. Here, we show that Smo activation is dependent on the levels of the phospholipid phosphatidylinositol-4 phosphate (PI4P). Loss of STT4 kinase, which is required for the generation of PI4P, exhibits hh loss-of-function phenotypes, whereas loss of Sac1 phosphatase, which is required for the degradation of PI4P, results in hh gain-of-function phenotypes in multiple settings during Drosophila development. Furthermore, loss of Ptc function, which results in the activation of Hh pathway, also causes an increase in PI4P levels. Sac1 functions downstream of STT4 and Ptc in the regulation of Smo membrane localization and Hh pathway activation. Taken together, our results suggest a model in which Ptc directly or indirectly functions to suppress the accumulation of PI4P. Binding of Hh to Ptc derepresses the levels of PI4P, which, in turn, promotes Smo activation.

  View details for DOI 10.1016/j.devcel.2010.06.007

  View details for Web of Science ID 000280469100010

  View details for PubMedID 20643350

  View details for PubMedCentralID PMC2945252

• Hedgehog pathway-regulated gene networks in cerebellum development and tumorigenesis PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Lee, E. Y., Ji, H., Ouyang, Z., Zhou, B., Ma, W., Vokes, S. A., McMahon, A. P., Wong, W. H., Scott, M. P. 2010; 107 (21): 9736-9741

  Abstract

  Many genes initially identified for their roles in cell fate determination or signaling during development can have a significant impact on tumorigenesis. In the developing cerebellum, Sonic hedgehog (Shh) stimulates the proliferation of granule neuron precursor cells (GNPs) by activating the Gli transcription factors. Inappropriate activation of Shh target genes results in unrestrained cell division and eventually medulloblastoma, the most common pediatric brain malignancy. We find dramatic differences in the gene networks that are directly driven by the Gli1 transcription factor in GNPs and medulloblastoma. Gli1 binding location analysis revealed hundreds of genomic loci bound by Gli1 in normal and cancer cells. Only one third of the genes bound by Gli1 in GNPs were also bound in tumor cells. Correlation with gene expression levels indicated that 116 genes were preferentially transcribed in tumors, whereas 132 genes were target genes in both GNPs and medulloblastoma. Quantitative PCR and in situ hybridization for some putative target genes support their direct regulation by Gli. The results indicate that transformation of normal GNPs into deadly tumor cells is accompanied by a distinct set of Gli-regulated genes and may provide candidates for targeted therapies.

  View details for DOI 10.1073/pnas.1004602107

  View details for Web of Science ID 000278054700048

  View details for PubMedID 20460306

  View details for PubMedCentralID PMC2906878

• Not Lost in Space: Trafficking in the Hedgehog Signaling Pathway SCIENCE SIGNALING Milenkovic, L., Scott, M. P. 2010; 3 (117)

  Abstract

  Compartmentalization within cells provides spatial organization of signaling pathways and ensures the specificity of signaling. In vertebrates, the primary cilium, a tiny microtubule-based protrusion present on most cells, is essential for organizing events during Hedgehog signal transduction. When cells are stimulated with Hedgehog ligands, proteins in the pathway move in and out of the cilia. Protein kinase A (PKA), which is implicated in diverse cellular processes including protein trafficking, is a component of the Hedgehog signaling pathway. PKA has been localized near primary cilia, at a location suitable for regulating the localization of other proteins in the pathway.

  View details for DOI 10.1126/scisignal.3117pe14

  View details for Web of Science ID 000276644900001

  View details for PubMedID 20388915

• The output of Hedgehog signaling is controlled by the dynamic association between Suppressor of Fused and the Gli proteins GENES & DEVELOPMENT Humke, E. W., Dorn, K. V., Milenkovic, L., Scott, M. P., Rohatgi, R. 2010; 24 (7): 670-682

  Abstract

  The transcriptional program orchestrated by Hedgehog signaling depends on the Gli family of transcription factors. Gli proteins can be converted to either transcriptional activators or truncated transcriptional repressors. We show that the interaction between Gli3 and Suppressor of Fused (Sufu) regulates the formation of either repressor or activator forms of Gli3. In the absence of signaling, Sufu restrains Gli3 in the cytoplasm, promoting its processing into a repressor. Initiation of signaling triggers the dissociation of Sufu from Gli3. This event prevents formation of the repressor and instead allows Gli3 to enter the nucleus, where it is converted into a labile, differentially phosphorylated transcriptional activator. This key dissociation event depends on Kif3a, a kinesin motor required for the function of primary cilia. We propose that the Sufu-Gli3 interaction is a major control point in the Hedgehog pathway, a pathway that plays important roles in both development and cancer.

  View details for DOI 10.1101/gad.1902910

  View details for Web of Science ID 000276235400006

  View details for PubMedID 20360384

  View details for PubMedCentralID PMC2849124

• Planar Cell Polarity Enables Posterior Localization of Nodal Cilia and Left-Right Axis Determination during Mouse and Xenopus Embryogenesis PLOS ONE Antic, D., Stubbs, J. L., Suyama, K., Kintner, C., Scott, M. P., Axelrod, J. D. 2010; 5 (2)

  Abstract

  Left-right asymmetry in vertebrates is initiated in an early embryonic structure called the ventral node in human and mouse, and the gastrocoel roof plate (GRP) in the frog. Within these structures, each epithelial cell bears a single motile cilium, and the concerted beating of these cilia produces a leftward fluid flow that is required to initiate left-right asymmetric gene expression. The leftward fluid flow is thought to result from the posterior tilt of the cilia, which protrude from near the posterior portion of each cell's apical surface. The cells, therefore, display a morphological planar polarization. Planar cell polarity (PCP) is manifested as the coordinated, polarized orientation of cells within epithelial sheets, or as directional cell migration and intercalation during convergent extension. A set of evolutionarily conserved proteins regulates PCP. Here, we provide evidence that vertebrate PCP proteins regulate planar polarity in the mouse ventral node and in the Xenopus gastrocoel roof plate. Asymmetric anterior localization of VANGL1 and PRICKLE2 (PK2) in mouse ventral node cells indicates that these cells are planar polarized by a conserved molecular mechanism. A weakly penetrant Vangl1 mutant phenotype suggests that compromised Vangl1 function may be associated with left-right laterality defects. Stronger functional evidence comes from the Xenopus GRP, where we show that perturbation of VANGL2 protein function disrupts the posterior localization of motile cilia that is required for leftward fluid flow, and causes aberrant expression of the left side-specific gene Nodal. The observation of anterior-posterior PCP in the mouse and in Xenopus embryonic organizers reflects a strong evolutionary conservation of this mechanism that is important for body plan determination.

  View details for DOI 10.1371/journal.pone.0008999

  View details for Web of Science ID 000274207200003

  View details for PubMedID 20126399

  View details for PubMedCentralID PMC2814853

• Lateral transport of Smoothened from the plasma membrane to the membrane of the cilium JOURNAL OF CELL BIOLOGY Milenkovic, L., Scott, M. P., Rohatgi, R. 2009; 187 (3): 365-374

  Abstract

  The function of primary cilia depends critically on the localization of specific proteins in the ciliary membrane. A major challenge in the field is to understand protein trafficking to cilia. The Hedgehog (Hh) pathway protein Smoothened (Smo), a 7-pass transmembrane protein, moves to cilia when a ligand is received. Using microscopy-based pulse-chase analysis, we find that Smo moves through a lateral transport pathway from the plasma membrane to the ciliary membrane. Lateral movement, either via diffusion or active transport, is quite distinct from currently studied pathways of ciliary protein transport in mammals, which emphasize directed trafficking of Golgi-derived vesicles to the base of the cilium. We anticipate that this alternative route will be used by other signaling proteins that function at cilia. The path taken by Smo may allow novel strategies for modulation of Hh signaling in cancer and regeneration.

  View details for DOI 10.1083/jcb.200907126

  View details for Web of Science ID 000271374200008

  View details for PubMedID 19948480

  View details for PubMedCentralID PMC2779247

• Tail Wags Dog: Primary Cilia and Tumorigenesis CANCER CELL Barakat, M. T., Scott, M. P. 2009; 16 (4): 276-277

  Abstract

  Aberrant activation of the Hedgehog (Hh) signaling pathway contributes to many forms of cancer. Primary cilia are Hh signal transduction centers. Two papers in a recent issue of Nature Medicine (Han et al., 2009; Wong et al., 2009) show that mutating cilia can increase or reduce the rates of tumorigenesis depending on how the Hh pathway is disrupted.

  View details for DOI 10.1016/j.ccr.2009.09.019

  View details for Web of Science ID 000270770400005

  View details for PubMedID 19800572

  View details for PubMedCentralID PMC3437380

• Rab35 Controls Actin Bundling by Recruiting Fascin as an Effector Protein SCIENCE Zhang, J., Fonovic, M., Suyama, K., Bogyo, M., Scott, M. P. 2009; 325 (5945): 1250-1254

  Abstract

  Actin filaments are key components of the eukaryotic cytoskeleton that provide mechanical structure and generate forces during cell shape changes, growth, and migration. Actin filaments are dynamically assembled into higher-order structures at specified locations to regulate diverse functions. The Rab family of small guanosine triphosphatases is evolutionarily conserved and mediates intracellular vesicle trafficking. We found that Rab35 regulates the assembly of actin filaments during bristle development in Drosophila and filopodia formation in cultured cells. These effects were mediated by the actin-bundling protein fascin, which directly associated with active Rab35. Targeting Rab35 to the outer mitochondrial membrane triggered actin recruitment, demonstrating a role for an intracellular trafficking protein in localized actin assembly.

  View details for DOI 10.1126/science.1174921

  View details for Web of Science ID 000269523200039

  View details for PubMedID 19729655

• Bortezomib Reverses a Post-Translational Mechanism of Tumorigenesis for Patched1 Haploinsufficiency in Medulloblastoma PEDIATRIC BLOOD & CANCER Taniguchi, E., Cho, M. J., Arenkiel, B. R., Hansen, M. S., Rivera, O. J., McCleish, A. T., Qualman, S. J., Guttridge, D. C., Scott, M. P., Capecchi, M. R., Keller, C. 2009; 53 (2): 136-144

  Abstract

  Tumor initiation has been attributed to haploinsufficiency at a single locus for a large number of cancers. Patched1 (Ptc1) was one of the first such loci, and Ptc1 haploinsufficiency has been asserted to lead to medulloblastoma and rhabdomyosarcoma in mice.To study the role of Ptc1 in cerebellar tumor development and to create a preclinical therapeutic platform, we have generated a conditional Ptc1 haploinsufficiency model of medulloblastoma by inactivating Ptc1 in Pax7-expressing cells of the cerebellum.These mice developed exclusively medulloblastoma. We show that despite the presence of transcription of Ptc1, Ptc1 protein is nearly undetectable or absent in tumors. Our results suggest that Ptc1 loss of function is complete, but achieved at the protein level rather than by the classic genetic two-hit mechanism or a strict half-dosage genetic haploinsufficiency mechanism. Furthermore, we found that bortezomib, a 26S proteasome inhibitor, had a significant anti-tumor activity in vitro and in vivo, which was accompanied by restoration of Ptc1 protein and downregulation of the hedgehog signaling pathway. The same effect was seen for both human and mouse medulloblastoma tumor cell growth.These results suggest that proteasome inhibition is a potential new therapeutic approach in medulloblastoma.

  View details for DOI 10.1002/pbc.21968

  View details for Web of Science ID 000267244500003

  View details for PubMedID 19213072

  View details for PubMedCentralID PMC2850215

• Communicating with Hedgehogs: Signaling in development and disease 16th Annual Conference of the International-Society-of-Development-Biologists Scott, M., Rohatgi, R., Milenkovic, L., Lee, E., Corcoran, R., Barakat, M., Humke, E. ELSEVIER SCIENCE BV. 2009: S26–S27

  View details for DOI 10.1016/j.mod.2009.06.1065

  View details for Web of Science ID 000270034900067

• Distinct developmental roles of planar cell polarity proteins vangl1, prickle1, and prickle2 in cortical crescents and primary cilia 68th Annual Meeting of the Society-for-Developmental-Biology Antic, D., Suyama, K., Axelrod, J. D., Scott, M. P. ACADEMIC PRESS INC ELSEVIER SCIENCE. 2009: 486–86

  View details for DOI 10.1016/j.ydbio.2009.05.375

  View details for Web of Science ID 000267777900366

• Case Studies of Ends-Out Gene Targeting in Drosophila GENESIS Chen, H., Ma, Z., Liu, Z., Tian, Y., Xiang, Y., Wang, C., Scott, M. P., Huang, X. 2009; 47 (5): 305-308

  Abstract

  Ends-in and ends-out gene replacement approaches have been successfully used to disrupt Drosophila genes involved in a variety of biological processes. These methods combine double-strand breaks and homologous recombination to replace a targeted chromosome region with a designed DNA sequence. Unfortunately, these methods require large numbers of single animal crosses, making them both time consuming and labor intensive. Here, we designed a single complete targeting vector for use in a mass crossing ends-out gene targeting study. Importantly, our gene targeting method included a balancer chromosome to block endogenous homologous chromosome pairing and to promote pairing between the foreign targeting DNA fragment and the targeted chromosome. This technique provided successful and efficient gene replacement, greatly facilitating the gene knockout procedure.

  View details for DOI 10.1002/dvg.20501

  View details for Web of Science ID 000266473500003

  View details for PubMedID 19298016

• Manipulation of an Innate Escape Response in Drosophila: Photoexcitation of acj6 Neurons Induces the Escape Response PLOS ONE Zimmermann, G., Wang, L., Vaughan, A. G., Manoli, D. S., Zhang, F., Deisseroth, K., Baker, B. S., Scott, M. P. 2009; 4 (4)

  Abstract

  The genetic analysis of behavior in Drosophila melanogaster has linked genes controlling neuronal connectivity and physiology to specific neuronal circuits underlying a variety of innate behaviors. We investigated the circuitry underlying the adult startle response, using photoexcitation of neurons that produce the abnormal chemosensory jump 6 (acj6) transcription factor. This transcription factor has previously been shown to play a role in neuronal pathfinding and neurotransmitter modality, but the role of acj6 neurons in the adult startle response was largely unknown.We show that the activity of these neurons is necessary for a wild-type startle response and that excitation is sufficient to generate a synthetic escape response. Further, we show that this synthetic response is still sensitive to the dose of acj6 suggesting that that acj6 mutation alters neuronal activity as well as connectivity and neurotransmitter production. RESULTS/SIGNIFICANCE: These results extend the understanding of the role of acj6 and of the adult startle response in general. They also demonstrate the usefulness of activity-dependent characterization of neuronal circuits underlying innate behaviors in Drosophila, and the utility of integrating genetic analysis into modern circuit analysis techniques.

  View details for DOI 10.1371/journal.pone.0005100

  View details for Web of Science ID 000265500700013

  View details for PubMedID 19340304

  View details for PubMedCentralID PMC2660433

• Hedgehog signal transduction by Smoothened: Pharmacologic evidence for a 2-step activation process PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Rohatgi, R., Milenkovic, L., Corcoran, R. B., Scott, M. P. 2009; 106 (9): 3196-3201

  Abstract

  The Hedgehog (Hh) signaling pathway controls growth, cell fate decisions, and morphogenesis during development. Damage to Hh transduction machinery can lead to birth defects and cancer. The transmembrane protein Smoothened (Smo) relays the Hh signal and is an important drug target in cancer. Smo enrichment in primary cilia is thought to drive activation of target genes. Using small-molecule agonists and antagonists to dissect Smo function, we find that Smo enrichment in cilia is not sufficient for signaling and a distinct second step is required for full activation. This 2-step mechanism--localization followed by activation--has direct implications for the design and use of anticancer therapeutics targeted against Smo.

  View details for DOI 10.1073/pnas.0813373106

  View details for Web of Science ID 000263844100041

  View details for PubMedID 19218434

  View details for PubMedCentralID PMC2642660

• A Homozygous Mutation in Human PRICKLE1 Causes an Autosomal-Recessive Progressive Myoclonus Epilepsy-Ataxia Syndrome AMERICAN JOURNAL OF HUMAN GENETICS Bassuk, A. G., Wallace, R. H., Buhr, A., Buller, A. R., Afawi, Z., Shimojo, M., Miyata, S., Chen, S., Gonzalez-Alegre, P., Griesbach, H. L., Wu, S., Nashelsky, M., Vladar, E. K., Antic, D., Ferguson, P. J., Cirak, S., Voit, T., Scott, M. P., Axelrod, J. D., Gurnett, C., Daoud, A. S., Kivity, S., Neufeld, M. Y., Mazarib, A., Straussberg, R., Walid, S., Korczyn, A. D., Slusarski, D. C., Berkovic, S. F., El-Shanti, H. I. 2008; 83 (5): 572-581

  Abstract

  Progressive myoclonus epilepsy (PME) is a syndrome characterized by myoclonic seizures (lightning-like jerks), generalized convulsive seizures, and varying degrees of neurological decline, especially ataxia and dementia. Previously, we characterized three pedigrees of individuals with PME and ataxia, where either clinical features or linkage mapping excluded known PME loci. This report identifies a mutation in PRICKLE1 (also known as RILP for REST/NRSF interacting LIM domain protein) in all three of these pedigrees. The identified PRICKLE1 mutation blocks the PRICKLE1 and REST interaction in vitro and disrupts the normal function of PRICKLE1 in an in vivo zebrafish overexpression system. PRICKLE1 is expressed in brain regions implicated in epilepsy and ataxia in mice and humans, and, to our knowledge, is the first molecule in the noncanonical WNT signaling pathway to be directly implicated in human epilepsy.

  View details for DOI 10.1016/j.ajhg.2008.10.003

  View details for Web of Science ID 000261006900003

  View details for PubMedID 18976727

  View details for PubMedCentralID PMC2668041

• Insulin-like Growth Factor 2 Is Required for Progression to Advanced Medulloblastoma in patched1 Heterozygous Mice CANCER RESEARCH Corcoran, R. B., Raveh, T. B., Barakat, M. T., Lee, E. Y., Scott, M. P. 2008; 68 (21): 8788-8795

  Abstract

  Medulloblastoma (MB) can arise in the cerebellum due to genetic activation of the Sonic Hedgehog (Shh) signaling pathway. During normal cerebellum development, Shh spurs the proliferation of granule neuron precursors (GNP), the precursor cells of MB. Mutations in the Shh receptor gene patched1 (ptc1+/-) lead to increased MB incidence in humans and mice. MB tumorigenesis in mice heterozygous for ptc1+/- shows distinct steps of progression. Most ptc1+/- mice form clusters of preneoplastic cells on the surface of the mature cerebellum that actively transcribe Shh target genes. In approximately 15% of mice, these preneoplastic cells will become fast-growing, lethal tumors. It was previously shown that the loss of function of insulin-like growth factor 2 (igf2) suppresses MB formation in ptc1+/- mice. We found that igf2 is not expressed in preneoplastic lesions but is induced as these lesions progress to more advanced MB tumors. Igf2 is not required for formation of preneoplastic lesions but is necessary for progression to advanced tumors. Exogenous Igf2 protein promoted proliferation of MB precursor cells (GNP) and a MB cell line, PZp53(MED). Blocking igf2 signaling inhibited growth of PZp53(MED) cells, implicating igf2 as a potential clinical target.

  View details for DOI 10.1158/0008-5472.CAN-08-2135

  View details for Web of Science ID 000260698900019

  View details for PubMedID 18974121

  View details for PubMedCentralID PMC2597356

• A nucleostemin family GTPase, NS3, acts in serotonergic neurons to regulate insulin signaling and control body size GENES & DEVELOPMENT Kaplan, D. D., Zimmermann, G., Suyama, K., Meyer, T., Scott, M. P. 2008; 22 (14): 1877-1893

  Abstract

  Growth and body size are regulated by the CNS, integrating the genetic developmental program with assessments of an animal's current energy state and environmental conditions. CNS decisions are transmitted to all cells of the animal by insulin/insulin-like signals. The molecular biology of the CNS growth control system has remained, for the most part, elusive. Here we identify NS3, a Drosophila nucleostemin family GTPase, as a powerful regulator of body size. ns3 mutants reach <60% of normal size and have fewer and smaller cells, but exhibit normal body proportions. NS3 does not act cell-autonomously, but instead acts at a distance to control growth. Rescue experiments were performed by expressing wild-type ns3 in many different cells of ns3 mutants. Restoring NS3 to only 106 serotonergic neurons rescued global growth defects. These neurons are closely apposed with those of insulin-producing neurons, suggesting possible communication between the two neuronal systems. In the brains of ns3 mutants, excess serotonin and insulin accumulate, while peripheral insulin pathway activation is low. Peripheral insulin pathway activation rescues the growth defects of ns3 mutants. The findings suggest that NS3 acts in serotonergic neurons to regulate insulin signaling and thus exert global growth control.

  View details for DOI 10.1101/gad.1670508

  View details for Web of Science ID 000257643400005

  View details for PubMedID 18628395

  View details for PubMedCentralID PMC2492735

• Cell biology - Arrestin' movement in Cilia SCIENCE Rohatgi, R., Scott, M. P. 2008; 320 (5884): 1726-1727

  View details for DOI 10.1126/science.1160448

  View details for Web of Science ID 000257121200023

  View details for PubMedID 18583599

• Disruption of the PACAP gene promotes medulloblastoma in ptc1 mutant mice DEVELOPMENTAL BIOLOGY Lelievre, V., Seksenyan, A., Nobuta, H., Yong, W. H., Chhith, S., Niewiadomski, P., Cohen, J. R., Dong, H., Flores, A., Liau, L. M., Kornblum, H. I., Scott, M. P., Waschek, J. A. 2008; 313 (1): 359-370

  Abstract

  Hedgehog (Hh) proteins and cAMP-dependent protein kinase A (PKA) generally play opposing roles in developmental patterning events. Humans and mice heterozygous for mutations in the sonic hedgehog (Shh) receptor gene patched-1 (ptc1) have an increased incidence of certain types of cancer, including medulloblastoma (MB), a highly aggressive tumor of the cerebellum. Despite the importance of PKA in Hh signaling, little is known about how PKA activity is regulated in the context of Hh signaling, or the consequences of improper regulation. One molecule that can influence PKA activity is pituitary adenylyl cyclase-activating peptide (PACAP), which has been shown to regulate cerebellar granule precursor proliferation in vitro, a cell population thought to give rise to MB. To test for a PACAP/Hh interaction in the initiation or propagation of these tumors, we introduced a PACAP mutation into ptc1 mutant mice. Deletion of a single copy of PACAP increased MB incidence approximate 2.5-fold, to 66%, thereby demonstrating that PACAP exerts a powerful inhibitory action on the induction, growth or survival of these tumors. Tumors from PACAP/ptc1 mutant mice retained PACAP receptor gene expression, and exhibited superinduction of Hh target genes compared to those from ptc1+/- mice. Moreover, PACAP inhibited proliferation of cell lines derived from tumors in a PKA-dependent manner, and inhibited expression of the Hh target gene gli1. The results provide genetic evidence that PACAP acts as a physiological factor that regulates the pathogenesis of Hh pathway-associated MB tumors.

  View details for DOI 10.1016/j.ydbio.2007.10.031

  View details for Web of Science ID 000252300500030

  View details for PubMedID 18036580

  View details for PubMedCentralID PMC2206628

• Drosophila Niemann-Pick Type C-2 genes control sterol homeostasis and steroid biosynthesis: a model of human neurodegenerative disease DEVELOPMENT Huang, X., Warren, J. T., Buchanan, J., Gilbert, L. I., Scott, M. P. 2007; 134 (20): 3733-3742

  Abstract

  Mutations in either of the two human Niemann-Pick type C (NPC) genes, NPC1 and NPC2, cause a fatal neurodegenerative disease associated with abnormal cholesterol accumulation in cells. npc1a, the Drosophila NPC1 ortholog, regulates sterol homeostasis and is essential for molting hormone (20-hydroxyecdysone; 20E) biosynthesis. While only one npc2 gene is present in yeast, worm, mouse and human genomes, a family of eight npc2 genes (npc2a-h) exists in Drosophila. Among the encoded proteins, Npc2a has the broadest expression pattern and is most similar in sequence to vertebrate Npc2. Mutation of npc2a results in abnormal sterol distribution in many cells, as in Drosophila npc1a or mammalian NPC mutant cells. In contrast to the ecdysteroid-deficient, larval-lethal phenotype of npc1a mutants, npc2a mutants are viable and fertile with relatively normal ecdysteroid level. Mutants in npc2b, another npc2 gene, are also viable and fertile, with no significant sterol distribution abnormality. However, npc2a; npc2b double mutants are not viable but can be rescued by feeding the mutants with 20E or cholesterol, the basic precursor of 20E. We conclude that npc2a functions redundantly with npc2b in regulating sterol homeostasis and ecdysteroid biosynthesis, probably by controlling the availability of sterol substrate. Moreover, npc2a; npc2b double mutants undergo apoptotic neurodegeneration, thus constituting a new fly model of human neurodegenerative disease.

  View details for DOI 10.1242/dev.004572

  View details for Web of Science ID 000249812700016

  View details for PubMedID 17804599

• ISWI regulates higher-order chromatin structure and histone H1 assembly in vivo PLOS BIOLOGY Corona, D. F., Siriaco, G., Armstrong, J. A., Snarskaya, N., McClymont, S. A., Scott, M. P., Tamkun, J. W. 2007; 5 (9): 2011-2021

  Abstract

  Imitation SWI (ISWI) and other ATP-dependent chromatin-remodeling factors play key roles in transcription and other processes by altering the structure and positioning of nucleosomes. Recent studies have also implicated ISWI in the regulation of higher-order chromatin structure, but its role in this process remains poorly understood. To clarify the role of ISWI in vivo, we examined defects in chromosome structure and gene expression resulting from the loss of Iswi function in Drosophila. Consistent with a broad role in transcriptional regulation, the expression of a large number of genes is altered in Iswi mutant larvae. The expression of a dominant-negative form of ISWI leads to dramatic alterations in higher-order chromatin structure, including the apparent decondensation of both mitotic and polytene chromosomes. The loss of ISWI function does not cause obvious defects in nucleosome assembly, but results in a significant reduction in the level of histone H1 associated with chromatin in vivo. These findings suggest that ISWI plays a global role in chromatin compaction in vivo by promoting the association of the linker histone H1 with chromatin.

  View details for DOI 10.1371/journal.pbio.0050232

  View details for Web of Science ID 000249552300019

  View details for PubMedID 17760505

  View details for PubMedCentralID PMC1951781

• Patching the gaps in Hedgehog signalling NATURE CELL BIOLOGY Rohatgi, R., Scott, M. P. 2007; 9 (9): 1005-1009

  Abstract

  The Hedgehog (Hh) pathway plays central roles in animal development and stem-cell function. Defects in Hh signalling lead to birth defects and cancer in humans. The first and often genetically damaged step in this pathway is the interaction between two membrane proteins - Patched (Ptc), encoded by a tumour suppressor gene, and Smoothened (Smo), encoded by a proto-oncogene. Recent work linking Hh signalling to sterol metabolites and protein-trafficking events at the primary cilium promises to shed light on the biochemical basis of how Patched inhibits Smoothened, and to provide new avenues for cancer treatment.

  View details for DOI 10.1038/ncb435

  View details for Web of Science ID 000249256500005

  View details for PubMedID 17762891

• Patched1 regulates Hedgehog signaling at the primary cilium SCIENCE Rohatgi, R., Milenkovic, L., Scott, M. P. 2007; 317 (5836): 372-376

  Abstract

  Primary cilia are essential for transduction of the Hedgehog (Hh) signal in mammals. We investigated the role of primary cilia in regulation of Patched1 (Ptc1), the receptor for Sonic Hedgehog (Shh). Ptc1 localized to cilia and inhibited Smoothened (Smo) by preventing its accumulation within cilia. When Shh bound to Ptc1, Ptc1 left the cilia, leading to accumulation of Smo and activation of signaling. Thus, primary cilia sense Shh and transduce signals that play critical roles in development, carcinogenesis, and stem cell function.

  View details for DOI 10.1126/science.1139740

  View details for Web of Science ID 000248150200047

  View details for PubMedID 17641202

• Developmental genomics of the most dangerous animal PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Scott, M. P. 2007; 104 (29): 11865-11866

  View details for DOI 10.1073/pnas.0704795104

  View details for Web of Science ID 000248199200001

  View details for PubMedID 17620616

  View details for PubMedCentralID PMC1924572

• Thirty-one flavors of Drosophila Rab proteins GENETICS Zhang, J., Schulze, K. L., Hiesinger, P. R., Suyama, K., Wang, S., Fish, M., Acar, M., Hoskins, R. A., Bellen, H. J., Scott, M. P. 2007; 176 (2): 1307-1322

  Abstract

  Rab proteins are small GTPases that play important roles in transport of vesicle cargo and recruitment, association of motor and other proteins with vesicles, and docking and fusion of vesicles at defined locations. In vertebrates, >75 Rab genes have been identified, some of which have been intensively studied for their roles in endosome and synaptic vesicle trafficking. Recent studies of the functions of certain Rab proteins have revealed specific roles in mediating developmental signal transduction. We have begun a systematic genetic study of the 33 Rab genes in Drosophila. Most of the fly proteins are clearly related to specific vertebrate proteins. We report here the creation of a set of transgenic fly lines that allow spatially and temporally regulated expression of Drosophila Rab proteins. We generated fluorescent protein-tagged wild-type, dominant-negative, and constitutively active forms of 31 Drosophila Rab proteins. We describe Drosophila Rab expression patterns during embryogenesis, the subcellular localization of some Rab proteins, and comparisons of the localization of wild-type, dominant-negative, and constitutively active forms of selected Rab proteins. The high evolutionary conservation and low redundancy of Drosophila Rab proteins make these transgenic lines a useful tool kit for investigating Rab functions in vivo.

  View details for DOI 10.1534/genetics.106.066761

  View details for Web of Science ID 000247870700046

  View details for PubMedID 17409086

  View details for PubMedCentralID PMC1894592

• Assessing the conservation of mammalian gene expression using high-density exon arrays MOLECULAR BIOLOGY AND EVOLUTION Xing, Y., Ouyang, Z., Kapur, K., Scott, M. P., Wong, W. H. 2007; 24 (6): 1283-1285

  Abstract

  Microarray data from multiple species have been used to study evolutionary constraints on gene expression. Expression measurements from conventional microarray platforms such as the 3' expression arrays are strongly affected by platform-dependent probe effects that may introduce apparent but misleading discrepancies between species. In this manuscript, we assess the conservation of mammalian gene expression in adult tissues using data from a high-density exon array platform. The exon arrays have more than 6 million probes on a single array targeting all exons in a genome. We find that, unlike 3' array data, gene expression measurements from exon arrays reveal patterns of gene expression that are highly conserved between humans and mice in multiple tissues. Our analysis provides strong evidence for widespread stabilizing selection pressure on transcript abundance during mammalian evolution.

  View details for DOI 10.1093/molbev/msm061

  View details for Web of Science ID 000247207700001

  View details for PubMedID 17387099

• Asymmetric distribution of Prickle-like 2 reveals an early underlying polarization of vestibular sensory epithelia in the inner ear JOURNAL OF NEUROSCIENCE Deans, M. R., Antic, D., Suyama, K., Scott, M. P., Axelrod, J. D., Goodrich, L. V. 2007; 27 (12): 3139-3147

  Abstract

  Vestibular hair cells have a distinct planar cell polarity (PCP) manifest in the morphology of their stereocilia bundles and the asymmetric localization of their kinocilia. In the utricle and saccule the hair cells are arranged in an orderly array about an abrupt line of reversal that separates fields of cells with opposite polarity. We report that the putative PCP protein Prickle-like 2 (Pk2) is distributed in crescents on the medial sides of vestibular epithelial cells before the morphological polarization of hair cells. Despite the presence of a line of polarity reversal, crescent position is not altered between hair cells of opposite polarity. Frizzled 6 (Fz6), a second PCP protein, is distributed opposite Pk2 along the lateral side of vestibular support cells. Similar to Pk2, the subcellular localization of Fz6 does not differ between cells located on opposite sides of the line of reversal. In addition, in Looptail/Van Gogh-like2 mutant mice Pk2 is distributed asymmetrically at embryonic day 14.5 (E14.5), but this localization is not coordinated between adjacent cells, and the crescents subsequently are lost by E18.5. Together, these results support the idea that a conserved PCP complex acts before stereocilia bundle development to provide an underlying polarity to all cells in the vestibular epithelia and that cells on either side of the line of reversal are programmed to direct the kinocilium in opposite directions with respect to the polarity axis defined by PCP protein distribution.

  View details for DOI 10.1523/JNEUROSCI.5151-06.2007

  View details for Web of Science ID 000245103700010

  View details for PubMedID 17376975

• Molecular Cell Biology, 6th Edition Lodish, H.F., A.J. Berk, C.A. Kaiser, M. Krieger, M.P. Scott, H. Ploegh, A. Bretscher 2007
• Mes2, a MADF-containing transcription factor essential for Drosophila development DEVELOPMENTAL DYNAMICS Zimmermann, G., Furlong, E. E., Suyama, K., Scott, M. P. 2006; 235 (12): 3387-3395

  Abstract

  The development of the Drosophila mesoderm is initiated by the basic helix-loop-helix transcription factor twist. We identified a gene encoding a putative transcription factor, mes2, in a screen for essential mesoderm-expressed genes that function downstream of twist. Mes2 protein belongs to a family of 48 Drosophila proteins containing MADF domains. MADF domains exist in worms, flies, and fish. Mes2 is a nuclear protein first produced in trunk and head mesoderm during late gastrulation. At later embryonic stages, mes2 is expressed in glia of the central and peripheral nervous systems, and in tissues derived from the head mesoderm. We have identified a null mutation of mes2 that leads to developmental arrest in first instar larvae. Increased production of Mes2 in multiple embryonic and larval tissues almost always causes lethality. The ubiquitous or epidermal misexpression of mes2 in the embryo causes a dramatic loss of epidermal integrity resulting in the failure of dorsal closure. Our data show that the precise regulation of mes2 expression is critical for normal development in Drosophila and implicate Mes2 in the regulation of essential target genes.

  View details for DOI 10.1002/dvdy.20970

  View details for Web of Science ID 000242667600020

  View details for PubMedID 17029287

• Hedgehog modulates cell cycle regulators in stem cells to control hematopoietic regeneration PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Trowbridge, J. J., Scott, M. P., Bhatia, M. 2006; 103 (38): 14134-14139

  Abstract

  The signals that control the regenerative ability of hematopoietic stem cells (HSCs) in response to damage are unknown. Here, we demonstrate that downstream activation of the Hedgehog (Hh) signaling pathway induces cycling and expansion of primitive bone marrow hematopoietic cells under homeostatic conditions and during acute regeneration. However, this effect is at the expense of HSC function, because continued Hh activation during regeneration represses expression of specific cell cycle regulators, leading to HSC exhaustion. In vivo treatment with an inhibitor of the Hh pathway rescues these transcriptional and functional defects in HSCs. Our study establishes Hh signaling as a regulator of the HSC cell cycle machinery that balances hematopoietic homeostasis and regeneration in vivo.

  View details for DOI 10.1073/pnas.0604568103

  View details for Web of Science ID 000240746600042

  View details for PubMedID 16968775

  View details for PubMedCentralID PMC1599924

• Oxysterols stimulate Sonic hedgehog signal transduction and proliferation of medulloblastoma cells PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Corcoran, R. B., Scott, M. P. 2006; 103 (22): 8408-8413

  Abstract

  Sterol synthesis is required for Sonic hedgehog (Shh) signal transduction. Errors in Shh signal transduction play important roles in the formation of human tumors, including medulloblastoma (MB). It is not clear which products of sterol synthesis are necessary for Shh signal transduction or how they act. Here we show that cholesterol or specific oxysterols are the critical products of sterol synthesis required for Shh pathway signal transduction in MB cells. In MB cells, sterol synthesis inhibitors reduce Shh target gene transcription and block Shh pathway-dependent proliferation. These effects of sterol synthesis inhibitors can be reversed by exogenous cholesterol or specific oxysterols. We also show that certain oxysterols can maximally activate Shh target gene transcription through the Smoothened (Smo) protein as effectively as the known Smo full agonist, SAG. Thus, sterols are required and sufficient for Shh pathway activation. These results suggest that oxysterols may be critical regulators of Smo, and thereby Shh signal transduction. Inhibition of Shh signaling by sterol synthesis inhibitors may offer a novel approach to the treatment of MB and other Shh pathway-dependent human tumors.

  View details for DOI 10.1073/pnas.0602852103

  View details for Web of Science ID 000238206800021

  View details for PubMedID 16707575

  View details for PubMedCentralID PMC1462959

• Microoptical characterization of piezoelectric vibratory microinjections in Drosophila embryos for genome-wide RNAi screen JOURNAL OF MICROELECTROMECHANICAL SYSTEMS Zhang, X. J., Scott, M. P., Quate, C. F., Solgaard, O. 2006; 15 (2): 277-286

  View details for DOI 10.1109/JMEMS.2006.872242

  View details for Web of Science ID 000236770200001

• Microoptical characterization and modeling of positioning forces on Drosophila embryos self-assembled in two-dimensional arrays. J. Microelectromechanical Syst. Zhang, X.J., C.-C. Chen, R.W. Bernstein, S. Zappe, M.P. Scott, O. Solgaard 2006; 14: 1187-1197
• Automated MEMS-based Drosophila embryo injection system for high-throughput RNAi screens LAB ON A CHIP Zappe, S., Fish, M., Scott, M. P., Solgaard, O. 2006; 6 (8): 1012-1019

  Abstract

  We have developed an automated system based on microelectromechanical systems (MEMS) injectors for reliable mass-injection of Drosophila embryos. Targeted applications are high-throughput RNA interference (RNAi) screens. Our injection needles are made of silicon nitride. The liquid to be injected is stored in an integrated 500 nl reservoir, and an externally applied air pressure pulse precisely controls the injected volume. A steady-state water flow rate per applied pressure of 1.2 nl s(-1) bar(-1) was measured for a needle with channel width, height and length of 6.1 microm, 2.3 microm and 350 microm, respectively. A typical volume of 60 pl per embryo can be reliably and rapidly delivered within tens of milliseconds. Theoretical predictions of flow rates match measured values within +/-10%. Embryos are attached to a glass slide surface and covered with oil. Packages with the injector chip and the embryo slide are mounted on motorized xyz-stages. Two cameras allow the user to quickly align the needle tip to alignment marks on the glass slide. Our system then automatically screens the glass slide for embryos and reliably detects and injects more than 98% of all embryos. Survival rates after deionized (DI) water injection of 80% and higher were achieved. A first RNAi experiment was successfully performed with double-stranded RNA (dsRNA) corresponding to the segment polarity gene armadillo at a concentration of 0.01 microM. Almost 80% of the injected embryos expressed an expected strong loss-of-function phenotype. Our system can replace current manual injection technologies and will support systematic identification of Drosophila gene functions.

  View details for DOI 10.1039/b600238b

  View details for Web of Science ID 000239313800007

  View details for PubMedID 16874371

• Microoptical characterization of piezoelectric vibratory microinjections in Drosophila embryos for genome-wide RNAi screen J. Microelectromechanical Syst. Zhang, X.J., M.P. Scott, C.F. Quate, O. Solgaard 2006; 15: 277-286
• A Drosophila model of the Niemann-Pick type C lysosome storage disease: dnpc1a is required for molting and sterol homeostasis DEVELOPMENT Huang, X., Suyama, K., Buchanan, J., Zhu, A. J., Scott, M. P. 2005; 132 (22): 5115-5124

  Abstract

  Niemann-Pick type C (NPC) disease is a fatal autosomal-recessive neurodegenerative disorder characterized by the inappropriate accumulation of unesterified cholesterol in aberrant organelles. The disease is due to mutations in either of two genes, NPC1, which encodes a transmembrane protein related to the Hedgehog receptor Patched, and NPC2, which encodes a secreted cholesterol-binding protein. Npc1 mutant mice can be partially rescued by treatment with specific steroids. We have created a Drosophila NPC model by mutating dnpc1a, one of two Drosophila genes related to mammalian NPC1. Cells throughout the bodies of dnpc1a mutants accumulated sterol in a punctate pattern, as in individuals with NPC1 mutations. The mutants developed only to the first larval stage and were unable to molt. Molting after the normal first instar period was restored to various degrees by feeding the mutants the steroid molting hormone 20-hydroxyecdysone, or the precursors of ecdysone biosynthesis, cholesterol and 7-dehydrocholesterol. dnpc1a is normally highly expressed in the ecdysone-producing ring gland. Ring gland-specific expression of dnpc1a in otherwise mutant flies allowed development to adulthood, suggesting that the lack of ecdysone in the mutants is the cause of death. We propose that dnpc1a mutants have sterols trapped in aberrant organelles, leading to a shortage of sterol in the endoplasmic reticulum and/or mitochondria of ring gland cells, and, consequently, inadequate ecdysone synthesis.

  View details for DOI 10.1242/dev.02079

  View details for Web of Science ID 000233831800019

  View details for PubMedID 16221727

• Microoptical characterization and modeling of positioning forces on drosophila embryos self-assembled in two-dimensional arrays JOURNAL OF MICROELECTROMECHANICAL SYSTEMS Zhang, X. J., Chen, C. C., Bernstein, R. W., Zappe, S., Scott, M. P., Solgaard, O. 2005; 14 (5): 1187-1197

  View details for DOI 10.1109/JMEMS.2005.851834

  View details for Web of Science ID 000232571000032

• Cell-autonomous death of cerebellar Purkinje neurons with autophagy in Niemann-Pick type C disease PLOS GENETICS Ko, D. C., Milenkovic, L., Beier, S. M., Manuel, H., Buchanan, J., Scott, M. P. 2005; 1 (1): 81-95

  Abstract

  Niemann-Pick type C is a neurodegenerative lysosomal storage disorder caused by mutations in either of two genes, npc1 and npc2. Cells lacking Npc1, which is a transmembrane protein related to the Hedgehog receptor Patched, or Npc2, which is a secreted cholesterol-binding protein, have aberrant organelle trafficking and accumulate large quantities of cholesterol and other lipids. Though the Npc proteins are produced by all cells, cerebellar Purkinje neurons are especially sensitive to loss of Npc function. Since Niemann-Pick type C disease involves circulating molecules such as sterols and steroids and a robust inflammatory response within the brain parenchyma, it is crucial to determine whether external factors affect the survival of Purkinje cells (PCs). We investigated the basis of neurodegeneration in chimeric mice that have functional npc1 in only some cells. Death of mutant npc1 cells was not prevented by neighboring wild-type cells, and wild-type PCs were not poisoned by surrounding mutant npc1 cells. PCs undergoing cell-autonomous degeneration have features consistent with autophagic cell death. Chimeric mice exhibited a remarkable delay and reduction of wasting and ataxia despite their substantial amount of mutant tissue and dying cells, revealing a robust mechanism that partially compensates for massive PC death.

  View details for DOI 10.1371/journal.pgen.0010007

  View details for Web of Science ID 000234295900010

  View details for PubMedID 16103921

  View details for PubMedCentralID PMC1183526

• Micro-optical characterization of fluidic self-assembly of Drosophila embryos through surface tension: Principle, simulation and experiments International Conference on Optical MEMS and Their Applications Zhang, X. J., Chen, C. C., Scott, M. P., Solgaard, O. OPTICAL SOC JAPAN. 2005: 352–57

  View details for Web of Science ID 000232089300016

• Communicating with Hedgehogs NATURE REVIEWS MOLECULAR CELL BIOLOGY Hooper, J. E., Scott, M. P. 2005; 6 (4): 306-317

  Abstract

  Signalling by secreted Hedgehog (Hh) proteins is important for the development of many tissues and organs. Damage to components of the Hh signal-transduction pathway can lead to birth defects and cancer. The Hh proteins are distributed in tissues in a gradient, and cells respond to different thresholds of Hh with distinct responses. The cellular machinery that is responsible for the unique molecular mechanisms of Hh signal transduction has been largely conserved during metazoan evolution.

  View details for DOI 10.1038/nrm1622

  View details for Web of Science ID 000228060400012

  View details for PubMedID 15803137

• Differential regulation of Hedgehog target gene transcription by Costal2 and Suppressor of Fused DEVELOPMENT Ho, K. S., Suyama, K., Fish, M., Scott, M. P. 2005; 132 (6): 1401-1412

  Abstract

  The mechanism by which the secreted signaling molecule Hedgehog (Hh) elicits concentration-dependent transcriptional responses from cells is not well understood. In the Drosophila wing imaginal disc, Hh signaling differentially regulates the transcription of target genes decapentaplegic (dpp), patched (ptc) and engrailed (en) in a dose-responsive manner. Two key components of the Hh signal transduction machinery are the kinesin-related protein Costal2 (Cos2) and the nuclear protein trafficking regulator Suppressor of Fused [Su(fu)]. Both proteins regulate the activity of the transcription factor Cubitus interruptus (Ci) in response to the Hh signal. We have analyzed the activities of mutant forms of Cos2 in vivo and found effects on differential target gene transcription. A point mutation in the motor domain of Cos2 results in a dominant-negative form of the protein that derepresses dpp but not ptc. Repression of ptc in the presence of the dominant-negative form of Cos2 requires Su(fu), which is phosphorylated in response to Hh in vivo. Overexpression of wild-type or dominant-negative cos2 represses en. Our results indicate that differential Hh target gene regulation can be accomplished by differential sensitivity of Cos2 and Su(Fu) to Hh.

  View details for DOI 10.1242/dev.01689

  View details for Web of Science ID 000228378000021

  View details for PubMedID 15750186

• Microoptical characterization of fluidic self-assembly of Drosophila embryos through surface tension: Principle, simulation, and experiments. Optical Rev. Zhang, X.J., C.C. Chen, M.P. Scott, O. Solgaard 2005; 12: 352-357
• Automated mems based fruit fly embryo injection system for genome-wide high-throughput RNAi screens 8th International Conference on Miniaturized Systems for Chemistry and Life Sciences Zappe, S., Fish, M., Scott, M. P., Solgaard, A. ROYAL SOC CHEMISTRY. 2005: 183–85

  View details for Web of Science ID 000229017400061

• Micromachined silicon diffractive optical force encoders: Principles and applications in biology 8th International Conference on Miniaturized Systems for Chemistry and Life Sciences Zhang, X. J., Zappe, S., Chen, C. C., Scott, M. P., Solgaard, O. ROYAL SOC CHEMISTRY. 2005: 518–20

  View details for Web of Science ID 000229017400172

• Incredible journey: how do developmental signals travel through tissue? GENES & DEVELOPMENT Zhu, A. J., Scott, M. P. 2004; 18 (24): 2985-2997

  Abstract

  How developmental signaling proteins traverse tissue during animal development, through or around tightly packed cells, remains an incompletely resolved mystery. Signaling protein movement is regulated to create gradients, control amounts, impose barriers, or provide direction. Signaling can be controlled by the rate of signal production, modification, active transport, trapping along the path, or by the properties of the receptor apparatus. Signals may move by diffusion outside cells, attached to migrating cells, attached to carrier molecules, through cells by transcytosis, along cell extensions, or in released membrane packets. Recent findings about the movement of Hedgehog, Wingless (Wnt), and TGF-beta signaling proteins have helped to clarify the molecular mechanisms used to ensure that developmental signals carry only good news.

  View details for DOI 10.1101/gad.1233104

  View details for Web of Science ID 000225853200001

  View details for PubMedID 15601817

• Obituary - Edward B. Lewis (1918-2004) NATURE Scott, M. P., Lawrence, P. A. 2004; 431 (7005): 143-143

  View details for DOI 10.1038/431143a

  View details for Web of Science ID 000223746000026

  View details for PubMedID 15356617

• Characterization of fluidic microassembly for immobilization and positioning of Drosophila embryos in 2-D arrays 12th International Conference on Solid-State Sensors, Actuators and Microsystems (TRANSDUCERS 03) Bernstein, R. W., Zhang, X. J., Zappe, S., Fish, M., Scott, M., Solgaard, O. ELSEVIER SCIENCE SA. 2004: 191–96

  View details for DOI 10.1016/j.sna.2003.11.021

  View details for Web of Science ID 000223704800010

• Design and operation of a microfluidic sorter for Drosophila embryos 12th International Conference on Solid-State Sensors, Actuators and Microsystems (TRANSDUCERS 03) Chen, C. C., Zappe, S., Sahin, O., Zhang, X. J., Fish, M., Scott, M., Solgaard, O. ELSEVIER SCIENCE SA. 2004: 59–66

  View details for DOI 10.1016/j.snu.2003.10.015

  View details for Web of Science ID 000223969400009

• Micromachined silicon force sensor based on diffractive optical encoders for characterization of microinjection 12th International Conference on Solid-State Sensors, Actuators and Microsystems (TRANSDUCERS 03) Zhang, X. J., Zappe, S., Bernstein, R. W., Sahin, O., Chen, C. C., Fish, M., Scott, M. P., Solgaard, O. ELSEVIER SCIENCE SA. 2004: 197–203

  View details for DOI 10.1016/j.sna.2003.11.028

  View details for Web of Science ID 000223704800011

• Binding between the Niemann-Pick C1 protein and a photoactivatable cholesterol analog requires a functional sterol-sensing domain PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Ohgami, N., Ko, D. C., Thomas, M., Scott, M. P., Chang, C. C., Chang, T. Y. 2004; 101 (34): 12473-12478

  Abstract

  Niemann-Pick type C (NPC) 1 protein plays important roles in moving cholesterol and other lipids out of late endosomes by means of vesicular trafficking, but it is not known whether NPC1 directly interacts with cholesterol. We performed photoaffinity labeling of intact cells expressing fluorescent protein (FP)-tagged NPC1 by using [(3)H]7,7-azocholestanol ([(3)H]AC). After immunoprecipitation, (3)H-labeled NPC1-GFP appeared as a single band. Including excess unlabeled sterol to the labeling reaction significantly diminished the labeling. Altering the NPC1 sterol-sensing domain (SSD) with loss-of-function mutations (P692S and Y635C) severely reduced the extent of labeling. To further demonstrate the specificity of labeling, we show that NPC2, a late endosomal/lysosomal protein that binds to cholesterol with high affinity, is labeled, whereas mutant NPC2 proteins inactive in binding cholesterol are not. Vamp7, an abundant late endosomal membrane protein without an SSD but with one transmembrane domain, cannot be labeled. Binding between [(3)H]AC and NPC1 does not require NPC2. Treating cells with either U-18666A, a compound that creates an NPC-like phenotype, or with bafilomycin A1, a compound that raises late endosomal pH, has no effect on labeling of NPC1-YFP, suggesting that both drugs affect processes other than NPC1 binding to cholesterol. We also developed a procedure to label the NPC1-YFP by [(3)H]AC in vitro and showed that cholesterol is more effective in protection against labeling than its analogs epicholesterol or 5-alpha-cholestan. Overall, the results demonstrate that there is direct binding between NPC1 and azocholestanol; the binding does not require NPC2 but requires a functional SSD within NPC1.

  View details for DOI 10.1073/pnas.0405255101

  View details for Web of Science ID 000223596200016

  View details for PubMedID 15314240

  View details for PubMedCentralID PMC514655

• Developmental biology and cancer. 63rd Annual Meeting of the Society-for-Developmental-Biology Scott, M. P. ACADEMIC PRESS INC ELSEVIER SCIENCE. 2004: 557–57

  View details for Web of Science ID 000222556200067

• Drosophila Costal1 mutations are alleles of protein kinase A that modulate hedgehog signaling GENETICS Collier, L. S., Suyama, K., Anderson, J. H., Scott, M. P. 2004; 167 (2): 783-796

  Abstract

  Hedgehog (Hh) signaling is crucial for the development of many tissues, and altered Hh signal transduction can result in cancer. The Drosophila Costal1 (Cos1) and costal2 (cos2) genes have been implicated in Hh signaling. cos2 encodes a kinesin-related molecule, one component of a cytoplasmic complex of Hh signal transducers. Mutations in Cos1 enhance loss-of-function cos2 mutations, but the molecular nature of Cos1 has been unknown. We found that previously identified alleles of Cos1 actually map to two separate loci. Four alleles of Cos1 appear to be dominant-negative mutations of a catalytic subunit of protein kinase A (pka-C1) and the fifth allele, Cos1(A1), is a gain-of-function allele of the PKA regulatory subunit pka-RII. PKA-RII protein levels are higher in Cos1(A1) mutants than in wild type. Overexpression of wild-type pka-RII phenocopies Cos1 mutants. PKA activity is aberrant in Cos1(A1) mutants. PKA-RII is uniformly overproduced in the wing imaginal disc in Cos1(A1) mutants, but only certain cells respond by activating the transcription factor Ci and Hh target gene transcription. This work shows that overexpression of a wild-type regulatory subunit of PKA is sufficient to activate Hh target gene transcription.

  View details for Web of Science ID 000222650500019

  View details for PubMedID 15238528

• A genome-wide study of gene activity reveals developmental signaling pathways in the preimplantation mouse embryo DEVELOPMENTAL CELL Wang, Q. T., Piotrowska, K., Ciemerych, M. A., Milenkovic, L., Scott, M. P., Davis, R. W., Zernicka-Goetz, M. 2004; 6 (1): 133-144

  Abstract

  The preimplantation development of the mammalian embryo encompasses a series of critical events: the transition from oocyte to embryo, the first cell divisions, the establishment of cellular contacts, the first lineage differentiation-all the first subtle steps toward a future body plan. Here, we use microarrays to explore gene activity during preimplantation development. We reveal robust and dynamic patterns of stage-specific gene activity that fall into two major phases, one up to the 2-cell stage (oocyte-to-embryo transition) and one after the 4-cell stage (cellular differentiation). The mouse oocyte and early embryo express components of multiple signaling pathways including those downstream of Wnt, BMP, and Notch, indicating that conserved regulators of cell fate and pattern formation are likely to function at the earliest embryonic stages. Overall, these data provide a detailed temporal profile of gene expression that reveals the richness of signaling processes in early mammalian development.

  View details for Web of Science ID 000222442700013

  View details for PubMedID 14723853

• Characterization of fluidic microassembly for immobilization and positioning of Drosophila embryos in 2-D arrays. Sensors and Actuators A: Physical. Bernstein, R.W., X. Zhang, S. Zappe, M. Fish, M.P. Scott, O. Solgaard 2004; 114: 191-196
• Micromachined force sensor based on diffractive optical encoders for characterization of microinjection Sensors and Actuators A: Physical Zhang, X.J., S. Zappe, R.W. Bernstein, O. Sahin, C.-C. Chen, M. Fish, M.P. Scott, O. Solgaard 2004; 114: 197-203
• Design and operation of a microfluidic sorter for Drosophila embryos. Sensors and Actuators B. Chemical. Chen, C.C., S. Zappe, O. Sahin, X.J. Zhang, M. Fish, M.P. Scott, O. Solgaard 2004: 59-66
• BioMEMS for high-throughput handling and microinjection of embryos Conference on MEMS/MOEMS Technologies and Applications II Bernstein, R. W., Scott, M., Solgaard, O. SPIE-INT SOC OPTICAL ENGINEERING. 2004: 67–73

  View details for DOI 10.1117/12.584626

  View details for Web of Science ID 000226862700008

• Characterization of fluidic microassembly for immobilization and positioning of Drosophila embryos in 2-D arrays Sensors and Actuators A: Physical R.W. Bernstein, X. Zhang, S. Zappe, M. Fish, M.P. Scott, O. Solgaard 2004; 114: 191-196
• Notch and Ras signaling pathway effector genes expressed in fusion competent and founder cells during Drosophila myogenesis DEVELOPMENT Artero, R., Furlong, E. E., Beckett, K., Scott, M. P., Baylies, M. 2003; 130 (25): 6257-6272

  Abstract

  Drosophila muscles originate from the fusion of two types of myoblasts, founder cells (FCs) and fusion-competent myoblasts (FCMs). To better understand muscle diversity and morphogenesis, we performed a large-scale gene expression analysis to identify genes differentially expressed in FCs and FCMs. We employed embryos derived from Toll10b mutants to obtain primarily muscle-forming mesoderm, and expressed activated forms of Ras or Notch to induce FC or FCM fate, respectively. The transcripts present in embryos of each genotype were compared by hybridization to cDNA microarrays. Among the 83 genes differentially expressed, we found genes known to be enriched in FCs or FCMs, such as heartless or hibris, previously characterized genes with unknown roles in muscle development, and predicted genes of unknown function. Our studies of newly identified genes revealed new patterns of gene expression restricted to one of the two types of myoblasts, and also striking muscle phenotypes. Whereas genes such as phyllopod play a crucial role during specification of particular muscles, others such as tartan are necessary for normal muscle morphogenesis.

  View details for DOI 10.1242/dev.00843

  View details for Web of Science ID 000187400300010

  View details for PubMedID 14602676

• Tgf beta signaling acts on a Hox response element to confer specificity and diversity to Hox protein function DEVELOPMENT Grienenberger, A., Merabet, S., Manak, J., Iltis, I., Fabre, A., Berenger, H., Scott, M. P., Pradel, J., Graba, Y. 2003; 130 (22): 5445-5455

  Abstract

  Hox proteins play fundamental roles in generating pattern diversity during development and evolution, acting in broad domains but controlling localized cell diversification and pattern. Much remains to be learned about how Hox selector proteins generate cell-type diversity. In this study, regulatory specificity was investigated by dissecting the genetic and molecular requirements that allow the Hox protein Abdominal A to activate wingless in only a few cells of its broad expression domain in the Drosophila visceral mesoderm. We show that the Dpp/Tgfbeta signal controls Abdominal A function, and that Hox protein and signal-activated regulators converge on a wingless enhancer. The signal, acting through Mad and Creb, provides spatial information that subdivides the domain of Abdominal A function through direct combinatorial action, conferring specificity and diversity upon Abdominal A activity.

  View details for DOI 10.1242/dev.00760

  View details for Web of Science ID 000187399800015

  View details for PubMedID 14507783

• Cancer - A twist in a hedgehog's tale NATURE Scott, M. P. 2003; 425 (6960): 780-782

  View details for DOI 10.1038/425780a

  View details for Web of Science ID 000186118500029

  View details for PubMedID 14574396

• Differential requirement for Gli2 and Gli3 in ventral neural cell fate specification DEVELOPMENTAL BIOLOGY Motoyama, J., Milenkovic, L., Iwama, M., Shikata, Y., Scott, M. P., Hui, C. C. 2003; 259 (1): 150-161

  Abstract

  Sonic hedgehog (Shh) directs the development of ventral cell fates, including floor plate and V3 interneurons, in the mouse neural tube. Here, we show that the transcription factors Gli2 and Gli3, mediators of Shh signaling, are required for the development of the ventral cell fates but make distinct contributions to controlling cell fates at different locations along the rostral-caudal axis. Mutants lacking Patched1 (Ptc1), the putative receptor of Shh, were used to analyze Gli functions. Ptc1(-/-) mutants develop floor plate, motor neuron, and V3 interneuron progenitors in lateral and dorsal regions, suggesting that the normal role of Ptc1 is to suppress ventral cell development in dorsal neural tube. The Ptc1(-/-) phenotype is rescued, with restoration of dorsal cell types, by the lack of Gli2, but only in the caudal neural tube. In triple mutants of Gli2, Gli3, and Ptc1, dorsal and lateral cell fates are restored in the entire neural tube. These observations suggest that Gli2 is essential for ventral specification in the caudal neural tube, and that in more rostral regions, only Gli3 can promote development of ventral cells if Gli2 is absent. Thus, Shh signaling is mediated by overlapping but distinct functions of Gli2 and Gli3, and their relative contributions vary along the rostral-caudal axis.

  View details for DOI 10.1016/S0012-1606(3)00159-3

  View details for Web of Science ID 000183791900013

  View details for PubMedID 12812795

• Transcriptional profiling of the Sonic hedgehog response: A critical role for N-myc in proliferation of neuronal precursors PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Oliver, T. G., Grasfeder, L. L., Carroll, A. L., Kaiser, C., Gillingham, C. L., Lin, S. M., Wickramasinghe, R., Scott, M. P., Wechsler-Reya, R. J. 2003; 100 (12): 7331-7336

  Abstract

  Cerebellar granule cells are the most abundant neurons in the brain, and granule cell precursors (GCPs) are a common target of transformation in the pediatric brain tumor medulloblastoma. Proliferation of GCPs is regulated by the secreted signaling molecule Sonic hedgehog (Shh), but the mechanisms by which Shh controls proliferation of GCPs remain inadequately understood. We used DNA microarrays to identify targets of Shh in these cells and found that Shh activates a program of transcription that promotes cell cycle entry and DNA replication. Among the genes most robustly induced by Shh are cyclin D1 and N-myc. N-myc transcription is induced in the presence of the protein synthesis inhibitor cycloheximide, so it appears to be a direct target of Shh. Retroviral transduction of N-myc into GCPs induces expression of cyclin D1, E2F1, and E2F2, and promotes proliferation. Moreover, dominant-negative N-myc substantially reduces Shh-induced proliferation, indicating that N-myc is required for the Shh response. Finally, cyclin D1 and N-myc are overexpressed in murine medulloblastoma. These findings suggest that cyclin D1 and N-myc are important mediators of Shh-induced proliferation and tumorigenesis.

  View details for DOI 10.1073/pnas.0832317100

  View details for Web of Science ID 000183493500078

  View details for PubMedID 12777630

  View details for PubMedCentralID PMC165875

• Altered localization of Drosophila smoothened protein activates hedgehog signal transduction GENES & DEVELOPMENT Zhu, A. J., Zheng, L. M., Suyama, K., Scott, M. P. 2003; 17 (10): 1240-1252

  Abstract

  Hedgehog (Hh) signaling is critical for many developmental events and must be restrained to prevent cancer. A transmembrane protein, Smoothened (Smo), is necessary to transcriptionally activate Hh target genes. Smo activity is blocked by the Hh transmembrane receptor Patched (Ptc). The reception of a Hh signal overcomes Ptc inhibition of Smo, activating transcription of target genes. Using Drosophila salivary gland cells in vivo and in vitro as a new assay for Hh signal transduction, we investigated the regulation of Hh-triggered Smo stabilization and relocalization. Hh causes Smo to move from internal membranes to the cell surface. Relocalization is protein synthesis-independent and occurs within 30 min of Hh treatment. Ptc and the kinesin-related protein Costal2 (Cos2) cause internalization of Smo, a process that is dependent on both actin and microtubules. Disruption of endocytosis by dominant negative dynamin or Rab5 prevents Smo internalization. Fly versions of Smo mutants associated with human tumors are constitutively present at the cell surface. Forced localization of Smo at the plasma membrane activates Hh target gene transcription. Conversely, trapping of activated Smo mutants in the ER prevents Hh target gene activation. Control of Smo localization appears to be a crucial step in Hh signaling in Drosophila.

  View details for DOI 10.1101/gad.1080803

  View details for Web of Science ID 000182941000007

  View details for PubMedID 12730121

  View details for PubMedCentralID PMC196058

• Factors controlling the development of cerebellar granule cells Annual Meeting of the Pediatric-Academic-Societies/Society-for-Pediatric-Research Penn, A. A., Raveh, T., Weissman, I. L., Scott, M. P. NATURE PUBLISHING GROUP. 2003: 548A–548A

  View details for Web of Science ID 000181897903098

• The integrity of a cholesterol-binding pocket in Niemann-Pick C2 protein is necessary to control lysosome cholesterol levels PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Ko, D. C., Binkley, J., Sidow, A., Scott, M. P. 2003; 100 (5): 2518-2525

  Abstract

  The neurodegenerative disease Niemann-Pick Type C2 (NPC2) results from mutations in the NPC2 (HE1) gene that cause abnormally high cholesterol accumulation in cells. We find that purified NPC2, a secreted soluble protein, binds cholesterol specifically with a much higher affinity (K(d) = 30-50 nM) than previously reported. Genetic and biochemical studies identified single amino acid changes that prevent both cholesterol binding and the restoration of normal cholesterol levels in mutant cells. The amino acids that affect cholesterol binding surround a hydrophobic pocket in the NPC2 protein structure, identifying a candidate sterol-binding location. On the basis of evolutionary analysis and mutagenesis, three other regions of the NPC2 protein emerged as important, including one required for efficient secretion.

  View details for DOI 10.1073/pnas.0530027100

  View details for Web of Science ID 000181365000065

  View details for PubMedID 12591949

  View details for PubMedCentralID PMC151373

• Integrated optical diffractive micrograting-based injection force sensor 12th International Conference on Solid-State Sensors, Actuators and Microsystems (TRANSDUCERS 03) Zhang, X. J., Zappe, S., Bernstein, R. W., Sahin, O., Chen, C. C., Fish, M., Scott, M., Solgaard, O. IEEE. 2003: 1051–1054

  View details for Web of Science ID 000184567300264

• Characterization of drosophila embryos immobilized by fluidic microassembly 12th International Conference on Solid-State Sensors, Actuators and Microsystems (TRANSDUCERS 03) Bernstein, R. W., Zhang, X. J., Zappe, S., Fish, M., Scott, M., Solgaard, O. IEEE. 2003: 987–990

  View details for Web of Science ID 000184567300248

• Molecular Cell Biology, 5th Ed. Lodish, H.F., A.J. Berk, C.A. Kaiser, M. Krieger, P. Matsudaira, M.P. Scott, S.L. Zipursky, J.E. Darnell 2003
• Microfluidic switch for embryo and cell sorting 12th International Conference on Solid-State Sensors, Actuators and Microsystems (TRANSDUCERS 03) Chen, C. C., Zappe, S., Sahin, O., Zhang, X. J., Furlong, E. E., Fish, M., Scott, M., Solgaard, O. IEEE. 2003: 659–662

  View details for Web of Science ID 000184567300166

• Zinc-dependent interaction between dishevelled and the Drosophila Wnt antagonist naked cuticle JOURNAL OF BIOLOGICAL CHEMISTRY Rousset, R., Wharton, D. A., Zimmermann, G., Scott, M. P. 2002; 277 (50): 49019-49026

  Abstract

  During Drosophila development, the naked cuticle (nkd) gene attenuates wingless/Wnt signaling through a negative feedback loop mechanism. Fly and vertebrate Nkd proteins contain a putative calcium-binding EF-hand motif, the EFX domain, that interacts with the basic/PDZ region of the Wnt signal transducer, dishevelled (Dsh). Here we show that Dsh binding by Drosophila Nkd in vitro is mediated by the EFX domain as well as an adjacent C-terminal sequence. In vivo data suggest that both of these regions contribute to the ability of Nkd to antagonize Wnt signaling. Mutations in the Nkd EF-hand designed to eliminate potential ion binding affected Nkd-Dsh interactions in the yeast two-hybrid assay but not in the glutathione S-transferase pull-down assay. Addition of the chelating agent EDTA abolished the in vitro Nkd-Dsh interaction. Surprisingly zinc, but not calcium, was able to restore Nkd-Dsh binding, suggesting a zinc-mediated interaction. Calcium 45- and zinc 65-blotting experiments show that Nkd is a zinc-binding metalloprotein. The results further clarify how Nkd may antagonize Wnt signaling via interaction with Dsh, and identify a novel zinc-binding domain in Drosophila Nkd that collaborates with the conserved EFX domain to bind Dsh.

  View details for DOI 10.1074/jbc.M203246200

  View details for Web of Science ID 000179789600135

  View details for PubMedID 12354775

• The Drosophila BRM complex facilitates global transcription by RNA polymerase II EMBO JOURNAL Armstrong, J. A., Papoulas, O., Daubresse, G., Sperling, A. S., Lis, J. T., Scott, M. P., Tamkun, J. W. 2002; 21 (19): 5245-5254

  Abstract

  Drosophila brahma (brm) encodes the ATPase subunit of a 2 MDa complex that is related to yeast SWI/SNF and other chromatin-remodeling complexes. BRM was identified as a transcriptional activator of Hox genes required for the specification of body segment identities. To clarify the role of the BRM complex in the transcription of other genes, we examined its distribution on larval salivary gland polytene chromosomes. The BRM complex is associated with nearly all transcriptionally active chromatin in a pattern that is generally non-overlapping with that of Polycomb, a repressor of Hox gene transcription. Reduction of BRM function dramatically reduces the association of RNA polymerase II with salivary gland chromosomes. A few genes, such as induced heat shock loci, are not associated with the BRM complex; transcription of these genes is not compromised by loss of BRM function. The distribution of the BRM complex thus correlates with a dependence on BRM for gene activity. These data suggest that the chromatin remodeling activity of the BRM complex plays a general role in facilitating transcription by RNA polymerase II.

  View details for Web of Science ID 000178502600024

  View details for PubMedID 12356740

  View details for PubMedCentralID PMC129039

• Gene expression during the life cycle of Drosophila melanogaster SCIENCE Arbeitman, M. N., Furlong, E. E., Imam, F., JOHNSON, E., Null, B. H., Baker, B. S., Krasnow, M. A., Scott, M. P., Davis, R. W., White, K. P. 2002; 297 (5590): 2270-2275

  Abstract

  Molecular genetic studies of Drosophila melanogaster have led to profound advances in understanding the regulation of development. Here we report gene expression patterns for nearly one-third of all Drosophila genes during a complete time course of development. Mutations that eliminate eye or germline tissue were used to further analyze tissue-specific gene expression programs. These studies define major characteristics of the transcriptional programs that underlie the life cycle, compare development in males and females, and show that large-scale gene expression data collected from whole animals can be used to identify genes expressed in particular tissues and organs or genes involved in specific biological and biochemical processes.

  View details for PubMedID 12351791

• Several PATCHED1 missense mutations display activity in patched1-deficient fibroblasts JOURNAL OF BIOLOGICAL CHEMISTRY Bailey, E. C., Milenkovic, L., Scott, M. P., Collawn, J. F., Johnson, R. L. 2002; 277 (37): 33632-33640

  Abstract

  Mutations in mouse and human patched1 (ptc1) genes are associated with birth defects and cancer. Ptc1 is a receptor for Hedgehog (Hh) signaling proteins. Hh proteins activate transcription of target genes, including ptc1, and Ptc1 represses those genes, both by regulating the activity of Gli transcription factors. We have established mammalian cell lines with reduced Ptc1 function and a lacZ reporter to investigate Hh signal transduction. Embryonic fibroblasts were derived from mice, heterozygous or homozygous for a ptc1 mutation that inserts lacZ under the control of the ptc1 promoter (ptc1-lacZ). In heterozygous ptc1 cells, ptc1-lacZ was expressed at low levels but could be induced by Sonic Hedgehog (Shh) and Gli-1. Homozygous ptc1 cells expressed high levels of ptc1-lacZ without Hh stimulation. ptc1-lacZ expression was dependent on cell density in ptc1 homozygotes and Hh-stimulated heterozygotes but was independent of density when Gli1 was used to activate ptc1-lacZ. A wild-type ptc1 transgene introduced into homozygous ptc1 cells greatly reduced ptc1-lacZ expression. Expression of either half of Ptc1 alone resulted in improper maturation of the protein and a failure to complement the ptc1(-/-) cells. When co-expressed, both Ptc1 halves matured and had an activity similar to that of the intact protein. Three missense PTCH1 mutations exhibited significant functions in homozygous ptc1 cells. The missense mutants retained activity when expressed at about 10-fold lower levels and appeared as stable as wild-type Ptc1. These studies suggest that some tumors and disease phenotypes may arise from small reductions in PTCH1 activity.

  View details for DOI 10.1074/jbc.M202203200

  View details for Web of Science ID 000177959100015

  View details for PubMedID 12072433

• Unraveling the genetic hierarchy of muscle development using genetics and genomics. Furlong, E. E., Artero, R. D., Baylies, M. K., Scott, M. P. ACADEMIC PRESS INC ELSEVIER SCIENCE. 2002: 482

  View details for Web of Science ID 000176830700191

• Prickle mediates feedback amplification to generate asymmetric planar cell polarity signaling CELL Tree, D. R., Shulman, J. M., Rousset, R., Scott, M. P., Gubb, D., Axelrod, J. D. 2002; 109 (3): 371-381

  Abstract

  Planar cell polarity signaling in Drosophila requires the receptor Frizzled and the cytoplasmic proteins Dishevelled and Prickle. From initial, symmetric subcellular distributions in pupal wing cells, Frizzled and Dishevelled become highly enriched at the distal portion of the cell cortex. We describe a Prickle-dependent intercellular feedback loop that generates asymmetric Frizzled and Dishevelled localization. In the absence of Prickle, Frizzled and Dishevelled remain symmetrically distributed. Prickle localizes to the proximal side of pupal wing cells and binds the Dishevelled DEP domain, inhibiting Dishevelled membrane localization and antagonizing Frizzled accumulation. This activity is linked to Frizzled activity on the adjacent cell surface. Prickle therefore functions in a feedback loop that amplifies differences between Frizzled levels on adjacent cell surfaces.

  View details for Web of Science ID 000175412100012

  View details for PubMedID 12015986

• Hedgehog signal transduction proteins: contacts of the Fused kinase and Ci transcription factor with the kinesin-related protein Costal2. BMC developmental biology Monnier, V., Ho, K. S., Sanial, M., Scott, M. P., Plessis, A. 2002; 2: 4-?

  Abstract

  Hedgehog signaling proteins play important roles in development by controlling growth and patterning in various animals including Drosophila and mammals. Hedgehog signaling triggers changes in responsive cells through a novel transduction mechanism that ultimately controls the transcription of specific target genes via the activity of zinc finger transcription factors of the Cubitus interruptus/GLI family. In flies, key Hedgehog signal transduction components have been identified including the kinesin-related protein Costal2, the serinethreonine kinase Fused, and the PEST-containing protein Suppressor of Fused. These proteins control Cubitus interruptus cleavage, nucleo-cytoplasmic localization and activation. In fly embryos, Costal2, Fused, Suppressor of Fused and Cubitus interruptus are associated in at least one cytoplasmic complex, which interacts with the microtubules in a Hedgehog-dependent manner.Here we identified and mapped direct interactions between Cos2, Fu, and Ci using an in vitro affinity assay and the yeast two-hybrid system.Our results provide new insights into the possible mechanism of the cytosolic steps of Hedgehog transduction.

  View details for PubMedID 11914126

  View details for PubMedCentralID PMC101406

• Sonic hedgehog in the nervous system: functions, modifications and mechanisms CURRENT OPINION IN NEUROBIOLOGY Ho, K. S., Scott, M. P. 2002; 12 (1): 57-63

  Abstract

  Signaling by Sonic hedgehog (Shh) controls important developmental processes, including dorsoventral neural tube patterning, neural stem cell proliferation, and neuronal and glial cell survival. Shh signaling involves lipid modifications to Shh itself, as well as changes in protein subcellular localization. Recent advances have revealed the importance of palmitoylation and acylation of Shh on its potency and migration capacity. Subsequent trafficking and organelle sorting in the Shh signaling pathway have been observed; these observations offer a new dimension to our understanding of downstream signal transduction events.

  View details for Web of Science ID 000173813000007

  View details for PubMedID 11861165

• Jelly belly: A Drosophila LDL receptor repeat-containing, signal required for mesoderm migration and differentiation CELL Weiss, J. B., Suyama, K. L., Lee, H. H., Scott, M. P. 2001; 107 (3): 387-398

  Abstract

  Inductive interactions subdivide the Drosophila mesoderm into visceral, somatic, and heart muscle precursors. The muscle precursors form organs by executing tissue-specific migrations and cell fusions. We identified a novel gene, jelly belly (jeb), which is required for visceral mesoderm development. jeb encodes a secreted protein that contains an LDL receptor repeat. In jeb mutants, visceral mesoderm precursors form, but they fail to migrate or differentiate normally; no visceral muscles develop. Jeb protein is produced in somatic muscle precursors and taken up by visceral muscle precursors. jeb reveals a signaling process in which somatic muscle precursors support the proper migration and differentiation of visceral muscle cells. Later in embryogenesis, jeb is transcribed in neurons and Jeb protein is found in axons.

  View details for Web of Science ID 000172022800009

  View details for PubMedID 11701128

• Patterns of gene expression during Drosophila mesoderm Development SCIENCE Furlong, E. E., Andersen, E. C., Null, B., White, K. P., Scott, M. P. 2001; 293 (5535): 1629-1633

  Abstract

  The transcription factor Twist initiates Drosophila mesoderm development, resulting in the formation of heart, somatic muscle, and other cell types. Using a Drosophila embryo sorter, we isolated enough homozygous twist mutant embryos to perform DNA microarray experiments. Transcription profiles of twist loss-of-function embryos, embryos with ubiquitous twist expression, and wild-type embryos were compared at different developmental stages. The results implicate hundreds of genes, many with vertebrate homologs, in stage-specific processes in mesoderm development. One such gene, gleeful, related to the vertebrate Gli genes, is essential for somatic muscle development and sufficient to cause neural cells to express a muscle marker.

  View details for Web of Science ID 000170802900054

  View details for PubMedID 11486054

• Signalling and endocytosis: Wnt breaks down on back roads NATURE CELL BIOLOGY Scott, M. 2001; 3 (8): E185-E186

  View details for Web of Science ID 000170393200011

  View details for PubMedID 11483976

• Vertebrate proteins related to Drosophila naked cuticle bind dishevelled and antagonize Wnt signaling DEVELOPMENTAL BIOLOGY Wharton, K. A., Zimmermann, G., Rousset, R., Scott, M. P. 2001; 234 (1): 93-106

  Abstract

  Wnt signals control cell fate decisions and orchestrate cell behavior in metazoan animals. In the fruit fly Drosophila, embryos defective in signaling mediated by the Wnt protein Wingless (Wg) exhibit severe segmentation defects. The Drosophila segment polarity gene naked cuticle (nkd) encodes an EF hand protein that regulates early Wg activity by acting as an inducible antagonist. Nkd antagonizes Wg via a direct interaction with the Wnt signaling component Dishevelled (Dsh). Here we describe two mouse and human proteins, Nkd1 and Nkd2, related to fly Nkd. The most conserved region among the fly and vertebrate proteins, the EFX domain, includes the putative EF hand and flanking sequences. EFX corresponds to a minimal domain required for fly or vertebrate Nkd to interact with the basic/PDZ domains of fly Dsh or vertebrate Dvl proteins in the yeast two-hybrid assay. During mouse development, nkd1 and nkd2 are expressed in multiple tissues in partially overlapping, gradient-like patterns, some of which correlate with known patterns of Wnt activity. Mouse Nkd1 can block Wnt1-mediated, but not beta-catenin-mediated, activation of a Wnt-dependent reporter construct in mammalian cell culture. Misexpression of mouse nkd1 in Drosophila antagonizes Wg function. The data suggest that the vertebrate Nkd-related proteins, similar to their fly counterpart, may act as inducible antagonists of Wnt signals.

  View details for Web of Science ID 000169059300008

  View details for PubMedID 11356022

• The HMG-domain protein BAP111 is important for the function of the BRM chromatin-remodeling complex in vivo PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Papoulas, O., Daubresse, G., Armstrong, J. A., Jin, J., Scott, M. P., Tamkun, J. W. 2001; 98 (10): 5728-5733

  Abstract

  The Drosophila trithorax group gene brahma (brm) encodes the ATPase subunit of a SWI/SNF-like chromatin-remodeling complex. A key question about chromatin-remodeling complexes is how they interact with DNA, particularly in the large genomes of higher eukaryotes. Here, we report the characterization of BAP111, a BRM-associated protein that contains a high mobility group (HMG) domain predicted to bind distorted or bent DNA. The presence of an HMG domain in BAP111 suggests that it may modulate interactions between the BRM complex and chromatin. BAP111 is an abundant nuclear protein that is present in all cells throughout development. By using gel filtration chromatography and immunoprecipitation assays, we found that the majority of BAP111 protein in embryos is associated with the BRM complex. Furthermore, heterozygosity for BAP111 enhanced the phenotypes resulting from a partial loss of brm function. These data demonstrate that the BAP111 subunit is important for BRM complex function in vivo.

  View details for Web of Science ID 000168623300065

  View details for PubMedID 11331758

  View details for PubMedCentralID PMC33281

• Information access - Building a "GenBank" of the published literature SCIENCE Roberts, R. J., Varmus, H. E., Ashburner, M., Brown, P. O., Eisen, M. B., Khosla, C., Kirschner, M., Nusse, R., Scott, M., Wold, B. 2001; 291 (5512): 2318-2319

  View details for Web of Science ID 000167618700024

  View details for PubMedID 11269300

• naked cuticle targets dishevelled to antagonize Wnt signal transduction GENES & DEVELOPMENT Rousset, R., Mack, J. A., Wharton, K. A., Axelrod, J. D., Cadigan, K. M., Fish, M. P., Nusse, R., Scott, M. P. 2001; 15 (6): 658-671

  Abstract

  In Drosophila embryos the protein Naked cuticle (Nkd) limits the effects of the Wnt signal Wingless (Wg) during early segmentation. nkd loss of function results in segment polarity defects and embryonic death, but how nkd affects Wnt signaling is unknown. Using ectopic expression, we find that Nkd affects, in a cell-autonomous manner, a transduction step between the Wnt signaling components Dishevelled (Dsh) and Zeste-white 3 kinase (Zw3). Zw3 is essential for repressing Wg target-gene transcription in the absence of a Wg signal, and the role of Wg is to relieve this inhibition. Our double-mutant analysis shows that, in contrast to Zw3, Nkd acts when the Wg pathway is active to restrain signal transduction. Yeast two hybrid and in vitro experiments indicate that Nkd directly binds to the basic-PDZ region of Dsh. Specially timed Nkd overexpression is capable of abolishing Dsh function in a distinct signaling pathway that controls planar-cell polarity. Our results suggest that Nkd acts directly through Dsh to limit Wg activity and thus determines how efficiently Wnt signals stabilize Armadillo (Arm)/beta-catenin and activate downstream genes.

  View details for Web of Science ID 000167821300003

  View details for PubMedID 11274052

• Dynamic movements of organelles containing Niemann-Pick C1 protein: NPC1 involvement in late endocytic events MOLECULAR BIOLOGY OF THE CELL Ko, D. C., Gordon, M. D., Jin, J. Y., Scott, M. P. 2001; 12 (3): 601-614

  Abstract

  People homozygous for mutations in the Niemann-Pick type C1 (NPC1) gene have physiological defects, including excess accumulation of intracellular cholesterol and other lipids, that lead to drastic neural and liver degeneration. The NPC1 multipass transmembrane protein is resident in late endosomes and lysosomes, but its functions are unknown. We find that organelles containing functional NPC1-fluorescent protein fusions undergo dramatic movements, some in association with extending strands of endoplasmic reticulum. In NPC1 mutant cells the NPC1-bearing organelles that normally move at high speed between perinuclear regions and the periphery of the cell are largely absent. Pulse-chase experiments with dialkylindocarbocyanine low-density lipoprotein showed that NPC1 organelles function late in the endocytic pathway; NPC1 protein may aid the partitioning of endocytic and lysosomal compartments. The close connection between NPC1 and the drug U18666A, which causes NPC1-like organelle defects, was established by rescuing drug-treated cells with overproduced NPC1. U18666A inhibits outward movements of NPC1 organelles, trapping membranes and cholesterol in perinuclear organelles similar to those in NPC1 mutant cells, even when cells are grown in lipoprotein-depleted serum. We conclude that NPC1 protein promotes the creation and/or movement of particular late endosomes, which rapidly transport materials to and from the cell periphery.

  View details for Web of Science ID 000170348200009

  View details for PubMedID 11251074

  View details for PubMedCentralID PMC30967

• Automated sorting of live transgenic embryos NATURE BIOTECHNOLOGY Furlong, E. E., Profitt, D., Scott, M. P. 2001; 19 (2): 153-156

  Abstract

  The vast selection of Drosophila mutants is an extraordinary resource for exploring molecular events underlying development and disease. We have designed and constructed an instrument that automatically separates Drosophila embryos of one genotype from a larger population of embryos, based on a fluorescent protein marker. This instrument can also sort embryos from other species, such as Caenorhabditis elegans. The machine sorts 15 living Drosophila embryos per second with more than 99% accuracy. Sorting living embryos will solve longstanding problems, including (1) the need for large quantities of RNA from homozygous mutant embryos to use in DNA microarray or gene-chip experiments, (2) the need for large amounts of protein extract from homozygous mutant embryos for biochemical studies, for example to determine whether a multiprotein complex forms or localizes correctly in vivo when one component is missing, and (3) the need for rapid genetic screening for gene expression changes in living embryos using a fluorescent protein reporter.

  View details for Web of Science ID 000166754700022

  View details for PubMedID 11175730

• A mouse model for medulloblastoma and basal cell nevus syndrome JOURNAL OF NEURO-ONCOLOGY Corcoran, R. B., Scott, M. P. 2001; 53 (3): 307-318

  Abstract

  Medulloblastoma (MB), a tumor of the cerebellum, is the most frequent type of malignant childhood brain tumor. Multiple genes are causally involved in medulloblastoma including PATCHED1 (PTCH). The Patchedl (Ptc1) protein is a receptor for Sonic hedgehog (Shh), a secreted protein ligand. Shh is involved in many signaling processes that control cell fate and growth, among which is its emission from Purkinje cells in the developing cerebellum. Purkinje cell-derived Shh stimulates mitosis of the granule cell precursors that may be the cell type of origin in medulloblastoma. Ptc1 limits the effects of the Shh signal, so mutations in PTCH may lead to persistent granule cell precursors susceptible to further genetic or environmental events that cause medulloblastoma. Mice heterozygous for patched (ptc1) mutations, like heterozygous PTCH humans, have a high rate of medulloblastoma as well as other tumors. We discuss features of the mouse model and how it is contributing to understanding the process of brain tumorigenesis.

  View details for Web of Science ID 000171537400007

  View details for PubMedID 11718263

• The developmental biology of brain tumors ANNUAL REVIEW OF NEUROSCIENCE Wechsler-Reya, R., Scott, M. P. 2001; 24: 385-428

  Abstract

  Tumors of the central nervous system (CNS) can be devastating because they often affect children, are difficult to treat, and frequently cause mental impairment or death. New insights into the causes and potential treatment of CNS tumors have come from discovering connections with genes that control cell growth, differentiation, and death during normal development. Links between tumorigenesis and normal development are illustrated by three common CNS tumors: retinoblastoma, glioblastoma, and medulloblastoma. For example, the retinoblastoma (Rb) tumor suppressor protein is crucial for control of normal neuronal differentiation and apoptosis. Excessive activity of the epidermal growth factor receptor and loss of the phosphatase PTEN are associated with glioblastoma, and both genes are required for normal growth and development. The membrane protein Patched1 (Ptc1), which controls cell fate in many tissues, regulates cell growth in the cerebellum, and reduced Ptc1 function contributes to medulloblastoma. Just as elucidating the mechanisms that control normal development can lead to the identification of new cancer-related genes and signaling pathways, studies of tumor biology can increase our understanding of normal development. Learning that Ptc1 is a medulloblastoma tumor suppressor led directly to the identification of the Ptc1 ligand, Sonic hedgehog, as a powerful mitogen for cerebellar granule cell precursors. Much remains to be learned about the genetic events that lead to brain tumors and how each event regulates cell cycle progression, apoptosis, and differentiation. The prospects for beneficial work at the boundary between oncology and developmental biology are great.

  View details for Web of Science ID 000170109100014

  View details for PubMedID 11283316

• Effects of oncogenic mutations in Smoothened and Patched can be reversed by cyclopamine NATURE Taipale, J., Chen, J. K., Cooper, M. K., Wang, B. L., Mann, R. K., Milenkovic, L., Scott, M. P., Beachy, P. A. 2000; 406 (6799): 1005-1009

  Abstract

  Basal cell carcinoma, medulloblastoma, rhabdomyosarcoma and other human tumours are associated with mutations that activate the proto-oncogene Smoothened (SMO) or that inactivate the tumour suppressor Patched (PTCH). Smoothened and Patched mediate the cellular response to the Hedgehog (Hh) secreted protein signal, and oncogenic mutations affecting these proteins cause excess activity of the Hh response pathway. Here we show that the plant-derived teratogen cyclopamine, which inhibits the Hh response, is a potential 'mechanism-based' therapeutic agent for treatment of these tumours. We show that cyclopamine or synthetic derivatives with improved potency block activation of the Hh response pathway and abnormal cell growth associated with both types of oncogenic mutation. Our results also indicate that cyclopamine may act by influencing the balance between active and inactive forms of Smoothened.

  View details for Web of Science ID 000089020200048

  View details for PubMedID 10984056

• In vivo functions of the patched protein: Requirement of the C terminus for target gene inactivation but not hedgehog sequestration MOLECULAR CELL Johnson, R. L., Milenkovic, L., Scott, M. P. 2000; 6 (2): 467-478

  Abstract

  The membrane protein Patched (Ptc) is a key regulator of Hedgehog (Hh) signaling in development and is mutated in human tumors. Ptc opposes Hh-induced gene transcription and sequesters Hh protein. To dissect these functions, we tested partially deleted forms of Ptc in Drosophila. Deletion of either half of Ptc abolishes all function while coexpression of the halves restores nearly full activity. Deletion of the final 156 residues of Ptc permits Hh sequestration but abolishes inhibition of Hh targets. This deletion has dominant-negative activity, promoting target gene activation in a ligand-independent manner. We observe little or no association of full-length or partially deleted Ptc with the membrane protein Smoothened in Drosophila cultured cells.

  View details for Web of Science ID 000089166100023

  View details for PubMedID 10983992

• Evidence that haploinsufficiency of Ptch leads to medulloblastoma in mice GENES CHROMOSOMES & CANCER Zurawel, R. H., Allen, C., Wechsler-Reya, R., Scott, M. P., Raffel, C. 2000; 28 (1): 77-81

  Abstract

  The PTCH gene encodes a putative tumor suppressor protein; germline alterations in PTCH have been found in patients with the nevoid basal cell carcinoma syndrome (NBCCS). Medulloblastoma, a brain tumor, develops in about 3% of NBCCS patients, and mutations in PTCH have also been described in a subset of sporadic medulloblastomas. The search for the causes of medulloblastoma has been hindered by the lack of an appropriate model system for this tumor type. Recently, a transgenic mouse hemizygous for the Ptch gene was generated by homologous recombination. Medulloblastomas were found in about 19% of these mice within the first 25 weeks after birth. The status of the wild-type PTCH allele in these tumors has not been investigated. For clearer definition of the role of PTCH as a tumor suppressor in medulloblastoma, 13 cerebellar tumors from transgenic Ptch(+/-) mice were examined for alterations in the remaining Ptch allele. A single mutation was found in one tumor, a C-to-A substitution changing a tyrosine to a stop codon; all other tumors exhibited a wild-type sequence. Two tumors with normal Ptch cDNA were examined by in situ hybridization. Ptch cDNA was found in tumor cells but not in associated tumor stroma. We also examined the mRNA expression levels for the remaining Ptch allele, as well as for Gli1, a gene known to be transcriptionally activated by Ptch inactivation. Blot analysis of RNA from the 13 tumors shows that Ptch mRNA of appropriate size is expressed in all tumors at varying levels. Expression of Gli1 was increased in tumors compared to normal cerebellum. These results suggest that deletion of one copy of Ptch may be sufficient to promote medulloblastoma development in mice.

  View details for Web of Science ID 000086120200009

  View details for PubMedID 10738305

• Distinct Hox protein sequences determine specificity in different tissues PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Chauvet, S., Merabet, S., Bilder, D., Scott, M. P., Pradel, J., Graba, Y. 2000; 97 (8): 4064-4069

  Abstract

  Hox genes encode evolutionarily conserved transcription factors that control the morphological diversification along the anteroposterior (A/P) body axis. Expressed in precise locations in the ectoderm, mesoderm, and endoderm, Hox proteins have distinct regulatory activities in different tissues. How Hox proteins achieve tissue-specific functions and why cells lying at equivalent A/P positions but in different germ layers have distinctive responses to the same Hox protein remains to be determined. Here, we examine this question by identifying parts of Hox proteins necessary for Hox function in different tissues. Available genetic markers allow the regulatory effects of two Hox proteins, Abdominal-A (AbdA) and Ultrabithorax (Ubx), to be distinguished in the Drosophila embryonic epidermis and visceral mesoderm (VM). Chimeric Ubx/AbdA proteins were tested in both tissues and used to identify protein sequences that endow AbdA with a different target gene specificity from Ubx. We found that distinct protein sequences define AbdA, as opposed to Ubx, function in the epidermis vs. the VM. These sequences lie mostly outside the homeodomain (HD), emphasizing the importance of non-HD residues for specific Hox activities. Hox tissue specificity is therefore achieved by sensing distinct Hox protein structures in different tissues.

  View details for Web of Science ID 000086461100052

  View details for PubMedID 10737765

  View details for PubMedCentralID PMC18149

• Naked cuticle (nkd) encodes and inducible antagonist of the Wnt signal transduction pathway. Wharton, K. A., Rousset, R., Mack, J., Zeng, W., Klein, P., Axelrod, J., Suyama, K., Scott, M. FEDERATION AMER SOC EXP BIOL. 2000: A724–A724

  View details for Web of Science ID 000085918104192

• Naked cuticle encodes an inducible antagonist of Wnt signalling NATURE Zeng, W. L., Wharton, K. A., Mack, J. A., Wang, K., Gadbaw, M., Suyama, K., Klein, P. S., Scott, M. P. 2000; 403 (6771): 789-795

  Abstract

  During animal development, cells have to respond appropriately to localized secreted signals. Proper responses to Hedgehog, transforming growth factor-beta, epidermal growth factor and fibroblast growth factor/Ras signals require cognate inducible antagonists such as Patched, Dad, Argos and Sprouty. Wnt signals are crucial in development and neoplasia. Here we show that naked cuticle (nkd), a Drosophila segment-polarity gene, encodes an inducible antagonist for the Wnt signal Wingless (Wg). In fly embryos and imaginal discs nkd transcription is induced by Wg. In embryos, decreased nkd function has an effect similar to excess Wg; at later stages such a decrease appears to have no effect. Conversely, overproduction of Nkd in Drosophila and misexpression of Nkd in the vertebrate Xenopus laevis result in phenotypes resembling those of loss of Wg/Wnt function. nkd encodes a protein with a single EF hand (a calcium-binding motif) that is most similar to the recoverin family of myristoyl switch proteins. Nkd may therefore link ion fluxes to the regulation of the potency, duration or distribution of Wnt signals. Signal-inducible feedback antagonists such as nkd may limit the effects of Wnt proteins in development and disease.

  View details for Web of Science ID 000085423100057

  View details for PubMedID 10693810

• Development: The natural history of genes CELL Scott, M. P. 2000; 100 (1): 27-40

  View details for Web of Science ID 000084722600003

  View details for PubMedID 10647929

• Hedgehog signaling in animal development and human disease 29th Ernst-Schering-Research-Foundation Workshop Bailey, E. C., Scott, M. P., Johnson, R. L. SPRINGER-VERLAG BERLIN. 2000: 211–235

  View details for Web of Science ID 000086852400012

  View details for PubMedID 10943312

• Defects in mouse mammary gland development caused by conditional haploinsufficiency of Patched-1 DEVELOPMENT Lewis, M. T., Ross, S., Strickland, P. A., Sugnet, C. W., Jimenez, E., Scott, M. P., Daniel, C. W. 1999; 126 (22): 5181-5193

  Abstract

  In vertebrates, the hedgehog family of cell signaling proteins and associated downstream network components play an essential role in mediating tissue interactions during development and organogenesis. Loss-of-function or misexpression mutation of hedgehog network components can cause birth defects, skin cancer and other tumors. The mammary gland is a specialized skin derivative requiring epithelial-epithelial and epithelial-stromal tissue interactions similar to those required for development of other organs, where these interactions are often controlled by hedgehog signaling. We have investigated the role of the Patched-1 (Ptc1) hedgehog receptor gene in mammary development and neoplasia. Haploinsufficiency at the Ptc1 locus results in severe histological defects in ductal structure, and minor morphological changes in terminal end buds in heterozygous postpubescent virgin animals. Defects are mainly ductal hyperplasias and dysplasias characterized by multilayered ductal walls and dissociated cells impacting ductal lumens. This phenotype is 100% penetrant. Remarkably, defects are reverted during late pregnancy and lactation but return upon involution and gland remodeling. Whole mammary gland transplants into athymic mice demonstrates that the observed dysplasias reflect an intrisic developmental defect within the gland. However, Ptc1-induced epithelial dysplasias are not stable upon transplantation into a wild-type epithelium-free fat pad, suggesting stromal (or epithelial and stromal) function of Ptc1. Mammary expression of Ptc1 mRNA is both epithelial and stromal and is developmentally regulated. Phenotypic reversion correlates with developmentally regulated and enhanced expression of Indian hedgehog (Ihh) during pregnancy and lactation. Data demonstrate a critical mammary role for at least one component of the hedgehog signaling network and suggest that Ihh is the primary hedgehog gene active in the gland.

  View details for Web of Science ID 000084171700024

  View details for PubMedID 10529434

• Ultraviolet and ionizing radiation enhance the growth of BCCs and trichoblastomas in patched heterozygous knockout mice NATURE MEDICINE Aszterbaum, M., Epstein, J., Oro, A., Douglas, V., LeBoit, P. E., Scott, M. P., Epstein, E. H. 1999; 5 (11): 1285-1291

  Abstract

  Basal cell carcinomas, the commonest human skin cancers, consistently have abnormalities of the hedgehog signaling pathway and often have PTCH gene mutations. We report here that Ptch+/- mice develop primordial follicular neoplasms resembling human trichoblastomas, and that exposure to ultraviolet radiation or ionizing radiation results in an increase in the number and size of these tumors and a shift in their histologic features so that they more closely resemble human basal cell carcinoma. The mouse basal cell carcinomas and trichoblastoma-like tumors resemble human basal cell carcinomas in their loss of normal hemidesmosomal components, presence of p53 mutations, frequent loss of the normal remaining Ptch allele, and activation of hedgehog target gene transcription. The Ptch mutant mice provide the first mouse model, to our knowledge, of ultraviolet and ionizing radiation-induced basal cell carcinoma-like tumors, and also demonstrate that Ptch inactivation and hedgehog target gene activation are essential for basal cell carcinoma tumorigenesis.

  View details for Web of Science ID 000086550600039

  View details for PubMedID 10545995

• A putative Drosophila homolog of the Huntington disease gene. Karlovich, C. A., Li, Z., Fish, M. P., Scott, M. P., Myers, R. M. CELL PRESS. 1999: A278–A278

  View details for Web of Science ID 000082879801553

• Mouse patched1 controls body size determination and limb patterning DEVELOPMENT Milenkovic, L., Goodrich, L. V., Higgins, K. M., Scott, M. P. 1999; 126 (20): 4431-4440

  Abstract

  Hedgehog (Hh) proteins control many developmental events by inducing specific cell fates or regulating cell proliferation. The Patched1 (Ptc1) protein, a binding protein for Hh molecules, appears to oppose Hh signals by repressing transcription of genes that can be activated by Hh. Sonic hedgehog (Shh), one of the vertebrate homologs of Hh, controls patterning and growth of the limb but the early embryonic lethality of ptc1(-)(/)(-) mice obscures the roles of ptc1 in later stages of development. We partially rescued ptc1 homozygous mutant embryos using a metallothionein promoter driving ptc1. In a wild-type background, the transgene causes a marked decrease in animal size starting during embryogenesis, and loss of anterior digits. In ptc1 homozygotes, a potent transgenic insert allowed survival to E14 and largely normal morphology except for midbrain overgrowth. A less potent transgene gave rise to partially rescued embryos with massive exencephaly, and polydactyly and branched digits in the limbs. The polydactyly was preceded by unexpected anterior limb bud transcription of Shh, so one function of ptc1 is to repress Shh expression in the anterior limb bud.

  View details for Web of Science ID 000083683300004

  View details for PubMedID 10498679

• A putative Drosophila homolog of the Huntington's disease gene HUMAN MOLECULAR GENETICS Li, Z., Karlovich, C. A., Fish, M. P., Scott, M. P., Myers, R. M. 1999; 8 (9): 1807-1815

  Abstract

  The Huntington's disease (HD) gene encodes a protein, huntingtin, with no known function and no detectable sequence similarity to other proteins in current databases. To gain insight into the normal biological role of huntingtin, we isolated and sequenced a cDNA encoding a protein that is a likely homolog of the HD gene product in Drosophila melanogaster. We also determined the complete sequence of 43 125 contiguous base pairs of genomic DNA that encompass the Drosophila HD gene, allowing the intron-exon structure and 5'- and 3'-flanking regions to be delineated. The predicted Drosophila huntingtin protein has 3583 amino acids, which is several hundred amino acids larger than any other previously characterized member of the HD family. Analysis of the genomic and cDNA sequences indicates that Drosophila HD has 29 exons, compared with the 67 exons present in vertebrate HD genes, and that Drosophila huntingtin lacks the polyglutamine and polyproline stretches present in its mammalian counterparts. The Drosophila HD mRNA is expressed in a broad range of developmental stages and in the adult, a temporal pattern of expression similar to that observed for mammalian HD transcripts. We can discern five regions of high similarity from multiple sequence alignments between Drosophila and vertebrate huntingtins. These regions may define functionally important domains within the protein.

  View details for Web of Science ID 000082600300027

  View details for PubMedID 10441347

• Vertebrate homologs of Drosophila suppressor of fused interact with the Gli family of transcriptional regulators DEVELOPMENTAL BIOLOGY Pearse, R. V., Collier, L. S., Scott, M. P., Tabin, C. J. 1999; 212 (2): 323-336

  Abstract

  The hedgehog (Hh) signaling pathway is crucial for pattern formation during metazoan development. Although originially characterized in Drosophila, vertebrate homologs have been identified for several, but not all, genes in the pathway. Analysis of mutants in Drosophila demonstrates that Suppressor of fused [Su(fu)] interacts genetically with genes encoding proteins in the Hh signal transduction pathway, and its protein product physically interacts with two of the proteins in the Hh pathway. We report here the molecular cloning and characterization of chicken and mouse homologs of Su(fu). The chick and mouse proteins are 27% identical and 53% similar at the amino acid level to the Drosophila melanogaster and Drosophila virilis proteins. Vertebrate Su(fu) is widely expressed in the developing embryo with higher levels in tissues that are known to be patterned by Hh signaling. The chick Su(fu) protein can physically interact with factors known to function in Hh signal transduction including the Drosophila serine/threonine kinase, Fused, and the vertebrate transcriptional regulators Gli1 and Gli3. This interaction may be significant for transcriptional regulation, as recombinant Su(fu) enhances the ability of Gli proteins to bind DNA in electrophoretic mobility shift assays.

  View details for Web of Science ID 000082123900006

  View details for PubMedID 10433824

• Overexpression of ptc1 inhibits induction of Shh target genes and prevents normal patterning in the neural tube DEVELOPMENTAL BIOLOGY Goodrich, L. V., Jung, D., Higgins, K. M., Scott, M. P. 1999; 211 (2): 323-334

  Abstract

  Patched (Ptc) is a human tumor suppressor protein and a candidate receptor for Hedgehog (Hh) proteins, which regulate growth and patterning in embryos. Ptc represses expression of Hh target genes such as Gli1 and ptc1 itself. Localized secretion of Hh appears to induce transcription of target genes in specific patterns by binding to Ptc and preventing it from functioning in recipient cells. People who are heterozygous for PTC1 exhibit a range of developmental defects, suggesting that some genes are inappropriately expressed when there is not enough Ptc protein. To test the idea that a balance between Hh and Ptc activities is essential for normal development, we overexpressed Ptc in the neural tube. We find that excess Ptc is sufficient to inhibit expression of Gli1 and ptc1, suggesting that Sonic hedgehog (Shh) cannot signal effectively. This leads to partial dorsalization of the neural tube and a wide spectrum of neural defects, ranging from embryonic lethality to hydrocephaly.

  View details for Web of Science ID 000081625000012

  View details for PubMedID 10395791

• Analysis of human basal cell carcinoma gene expression at a genomic scale via cDNA microarrays Fan, H., Barry, C. T., Baek, S. C., Lin, Q., Brown, P. O., Khavari, P. A. NATURE PUBLISHING GROUP. 1999: 528–28

  View details for Web of Science ID 000079495800056

• Regulation of Shh target gene expression plays a critical role in hair follicle and BCC formation Oro, A. E., Higgins, K., Scott, M. P. NATURE PUBLISHING GROUP. 1999: 525–25

  View details for Web of Science ID 000079495800037

• The Drosophila kismet gene is related to chromatin-remodeling factors and is required for both segmentation and segment identity DEVELOPMENT Daubresse, G., Deuring, R., Moore, L., Papoulas, O., Zakrajsek, I., Waldrip, W. R., Scott, M. P., Kennison, J. A., Tamkun, J. W. 1999; 126 (6): 1175-1187

  Abstract

  The Drosophila kismet gene was identified in a screen for dominant suppressors of Polycomb, a repressor of homeotic genes. Here we show that kismet mutations suppress the Polycomb mutant phenotype by blocking the ectopic transcription of homeotic genes. Loss of zygotic kismet function causes homeotic transformations similar to those associated with loss-of-function mutations in the homeotic genes Sex combs reduced and Abdominal-B. kismet is also required for proper larval body segmentation. Loss of maternal kismet function causes segmentation defects similar to those caused by mutations in the pair-rule gene even-skipped. The kismet gene encodes several large nuclear proteins that are ubiquitously expressed along the anterior-posterior axis. The Kismet proteins contain a domain conserved in the trithorax group protein Brahma and related chromatin-remodeling factors, providing further evidence that alterations in chromatin structure are required to maintain the spatially restricted patterns of homeotic gene transcription.

  View details for Web of Science ID 000079622400009

  View details for PubMedID 10021337

• Development - Hox proteins reach out round DNA NATURE Scott, M. P. 1999; 397 (6721): 649-?

  View details for Web of Science ID 000078840100022

• Recruitment of a hedgehog regulatory circuit in butterfly eyespot evolution SCIENCE Keys, D. N., Lewis, D. L., Selegue, J. E., Pearson, B. J., Goodrich, L. V., Johnson, R. L., Gates, J., Scott, M. P., Carroll, S. B. 1999; 283 (5401): 532-534

  Abstract

  The origin of new morphological characters is a long-standing problem in evolutionary biology. Novelties arise through changes in development, but the nature of these changes is largely unknown. In butterflies, eyespots have evolved as new pattern elements that develop from special organizers called foci. Formation of these foci is associated with novel expression patterns of the Hedgehog signaling protein, its receptor Patched, the transcription factor Cubitus interruptus, and the engrailed target gene that break the conserved compartmental restrictions on this regulatory circuit in insect wings. Redeployment of preexisting regulatory circuits may be a general mechanism underlying the evolution of novelties.

  View details for Web of Science ID 000078203300035

  View details for PubMedID 9915699

• Control of neuronal precursor proliferation in the cerebellum by sonic hedgehog NEURON Wechsler-Reya, R. J., Scott, M. P. 1999; 22 (1): 103-114

  Abstract

  Cerebellar granule cells are the most abundant type of neuron in the brain, but the molecular mechanisms that control their generation are incompletely understood. We show that Sonic hedgehog (Shh), which is made by Purkinje cells, regulates the division of granule cell precursors (GCPs). Treatment of GCPs with Shh prevents differentiation and induces a potent, long-lasting proliferative response. This response can be inhibited by basic fibroblast growth factor or by activation of protein kinase A. Blocking Shh function in vivo dramatically reduces GCP proliferation. These findings provide insight into the mechanisms of normal growth and tumorigenesis in the cerebellum.

  View details for Web of Science ID 000078410300016

  View details for PubMedID 10027293

• Hedgehog and Patched in neural development and disease NEURON Goodrich, L. V., Scott, M. P. 1998; 21 (6): 1243-1257

  View details for Web of Science ID 000077910600008

  View details for PubMedID 9883719

• Splitting hairs: Dissecting roles of signaling systems in epidermal development CELL Oro, A. E., Scott, M. P. 1998; 95 (5): 575-578

  View details for Web of Science ID 000077253700001

  View details for PubMedID 9845357

• Dorsoventral patterning in the Drosophila central nervous system: the intermediate neuroblasts defective homeobox gene specifies intermediate column identity GENES & DEVELOPMENT Weiss, J. B., Von Ohlen, T., Mellerick, D. M., Dressler, G., Doe, C. Q., Scott, M. P. 1998; 12 (22): 3591-3602

  Abstract

  One of the first steps in neurogenesis is the diversification of cells along the dorsoventral axis. In Drosophila the central nervous system develops from three longitudinal columns of cells: ventral cells that express the vnd/nk2 homeobox gene, intermediate cells, and dorsal cells that express the msh homeobox gene. Here we describe a new Drosophila homeobox gene, intermediate neuroblasts defective (ind), which is expressed specifically in the intermediate column cells. ind is essential for intermediate column development: Null mutants have a transformation of intermediate to dorsal column neuroectoderm fate, and only 10% of the intermediate column neuroblasts develop. The establishment of dorsoventral column identity involves negative regulation: Vnd represses ind in the ventral column, whereas ind represses msh in the intermediate column. Vertebrate genes closely related to vnd (Nkx2.1 and Nkx2.2), ind (Gsh1 and Gsh2), and msh (Msx1 and Msx3) are expressed in corresponding ventral, intermediate, and dorsal domains during vertebrate neurogenesis, raising the possibility that dorsoventral patterning within the central nervous system is evolutionarily conserved.

  View details for Web of Science ID 000077326700011

  View details for PubMedID 9832510

  View details for PubMedCentralID PMC317240

• Progressive ataxia, myoclonic epilepsy and cerebellar apoptosis in cystatin B-deficient mice NATURE GENETICS Pennacchio, L. A., Bouley, D. M., Higgins, K. M., Scott, M. P., Noebels, J. L., Myers, R. M. 1998; 20 (3): 251-258

  Abstract

  Loss-of-function mutations in the gene (CSTB) encoding human cystatin B, a widely expressed cysteine protease inhibitor, are responsible for a severe neurological disorder known as Unverricht-Lundborg disease (EPM1). The primary cellular events and mechanisms underlying the disease are unknown. We found that mice lacking cystatin B develop myoclonic seizures and ataxia, similar to symptoms seen in the human disease. The principal cytopathology appears to be a loss of cerebellar granule cells, which frequently display condensed nuclei, fragmented DNA and other cellular changes characteristic of apoptosis. This mouse model of EPM1 provides evidence that cystatin B, a non-caspase cysteine protease inhibitor, has a role in preventing cerebellar apoptosis.

  View details for Web of Science ID 000076698100018

  View details for PubMedID 9806543

• Hedgehog and Wingless induce metameric pattern in the Drosophila visceral mesoderm DEVELOPMENTAL BIOLOGY Bilder, D., Scott, M. P. 1998; 201 (1): 43-56

  Abstract

  The Drosophila visceral mesoderm (VM) is a favorite system for studying the regulation of target genes by Hox proteins. The VM is formed by cells from only the anterior subdivision of each mesodermal parasegment (PS). We show here that the VM itself acquires modular anterior-posterior subdivisions similar to those found in the ectoderm. As VM progenitors merge to form a continuous band running anterior to posterior along the embryo, expression of connectin (con) in 11 metameric patches within the VM reveals VM subdivisions analagous to ectodermal compartments. The VM subdivisions form in response to ectodermal production of secreted signals encoded by the segment polarity genes hedgehog (hh) and wingless (wg) and are independent of Hox gene activity. A cascade of induction from ectoderm to mesoderm to endoderm thus subdivides the gut tissues along the A-P axis. Induction of VM subdivisions may converge with Hox-mediated information to refine spatial patterning in the VM. Con patches align with ectodermal engrailed stripes, so the VM subdivisions correspond to PS 2-12 boundaries in the VM. The PS boundaries demarcated by Con in the VM can be used to map expression domains of Hox genes and their targets with high resolution. The resultant map suggests a model for the origins of VM-specific Hox expression in which Hox domains clonally inherited from blastoderm ancestors are modified by diffusible signals acting on VM-specific enhancers.

  View details for Web of Science ID 000075918500004

  View details for PubMedID 9733572

• New players and puzzles in the Hedgehog signaling pathway CURRENT OPINION IN GENETICS & DEVELOPMENT Johnson, R. L., Scott, M. P. 1998; 8 (4): 450-456

  Abstract

  Members of the Hedgehog (Hh) family of signaling proteins control cell fates and proliferation during animal development in part by regulating the transcription of specific genes. Depending on the tissue, Hh can act over long or short distances, to signal directly or by inducing secondary signals. Recent discoveries include new components of the pathway as well as novel regulatory mechanisms involving cholesterol, proteolysis, and the cytoskeleton. The role of Hh in carcinogenesis is underscored by the identification of mutations in several pathway components in skin and brain tumors.

  View details for Web of Science ID 000075603800012

  View details for PubMedID 9729722

• The function and subcellular localization of Costal2, a novel kinesin-related protein that interacts with members of the hedgehog signal transduction pathway Ho, K. S., Scott, M. P. ACADEMIC PRESS INC ELSEVIER SCIENCE. 1998: 216–16

  View details for Web of Science ID 000074281900340

• Identification of mutations in the human PATCHED gene in sporadic basal cell carcinomas and in patients with the basal cell nevus syndrome JOURNAL OF INVESTIGATIVE DERMATOLOGY Aszterbaum, M., Rothman, A., Johnson, R. L., Fisher, M., Xie, J. W., Bonifas, J. M., Zhang, X. L., Scott, M. P., Epstein, E. H. 1998; 110 (6): 885-888

  Abstract

  Mutations in PATCHED (PTC), the human homolog of the Drosophila patched gene, have been identified in most exons of the gene in patients with the basal cell nevus syndrome and in sporadic basal cell carcinomas. We have screened the 23 PTC exons for mutations using single strand conformation polymorphism analysis of DNA from 86 basal cell nevus syndrome probands, 26 sporadic basal cell carcinomas, and seven basal cell nevus syndrome-associated basal cell carcinomas. This screen identified mutations located in eight exons in 13 of the basal cell nevus syndrome patients and in three of the tumors. The most common mutations were frameshifts resulting in premature chain termination. These results provide further evidence for the crucial role of PTC as a tumor suppressor in human keratinocytes.

  View details for Web of Science ID 000073697600007

  View details for PubMedID 9620294

• The formation and evolution of butterfly wing color patterns Carroll, S. B., Galant, R., Keys, D., Lewis, D., Skeath, J. B., Goodrich, L., Scott, M. P., Paddock, S. ACADEMIC PRESS INC ELSEVIER SCIENCE. 1998: 164–64

  View details for Web of Science ID 000074281900037

• Balance of hedgehog and patched required for proper limb patterning. Milenkovic, L., Goodrich, L. V., Scott, M. P., Hughes, H. ACADEMIC PRESS INC ELSEVIER SCIENCE. 1998: 185–85

  View details for Web of Science ID 000074281900161

• Sonic hedgehog regulates proliferation of granule cell precursors in the developing cerebellum Wechsler-Reya, R. J., Roosa, J., Scott, M. P. ACADEMIC PRESS INC ELSEVIER SCIENCE. 1998: 194–94

  View details for Web of Science ID 000074281900216

• Regulation of growth by the patched tumor suppressor gene. Wechsler-Reya, R., Milenkovic, L., Goodrich, L. V., Oro, A. E., Higgins, K. M., Scott, M. P. ACADEMIC PRESS INC. 1998: 166

  View details for Web of Science ID 000074281900049

• Regulation of EGF receptor dimerization monitored in live cells Rossi, F. M., Blakely, B. T., Blau, H. M. ACADEMIC PRESS INC ELSEVIER SCIENCE. 1998: 166–66

  View details for Web of Science ID 000074281900048

• A screen for new genes involved in heart development Furlong, E. E., Beitel, G., Scott, M. P. ACADEMIC PRESS INC ELSEVIER SCIENCE. 1998: 206–

  View details for Web of Science ID 000074281900282

• The Drosophila naked gene (nkd) encodes a novel Wg-induced EF-hand protein which acts in the Wg receiving cells to inhibit Wg signaling. Mack, J., Wharton, K., Zeng, W. L., Wang, K., Scott, M. P. ACADEMIC PRESS INC ELSEVIER SCIENCE. 1998: 221–21

  View details for Web of Science ID 000074281900370

• Towards development of an in vitro model of human BCC formation. Oro, A. E., Fan, H., Khavari, P. A., Scott, M. P. ACADEMIC PRESS INC ELSEVIER SCIENCE. 1998: 196–96

  View details for Web of Science ID 000074281900230

• Wnt and TGF beta signals subdivide the AbdA Hox domain during Drosophila mesoderm patterning DEVELOPMENT Bilder, D., Graba, Y., Scott, M. P. 1998; 125 (9): 1781-1790

  Abstract

  Hox genes have large expression domains yet control the formation of fine pattern elements at specific locations. We have examined the mechanism underlying subdivision of the abdominal-A (abdA) Hox domain in the visceral mesoderm. AbdA directs formation of an embryonic midgut constriction at a precise location within the broad and uniform abdA expression domain. The constriction divides the abdA domain of the midgut into two chambers, the anterior one producing the Pointed (Pnt) ETS transcription factors and the posterior one the Odd-paired (Opa) zinc finger protein. Transcription of both pnt and opa is activated by abdA but the adjacent non-overlapping patterns are not due to mutual opa-pnt regulation. Near the anterior limit of the abdA domain, two signals, Dpp (a TGFbeta) and Wg (a Wnt), are produced, in adjacent non-overlapping patterns, under Hox control in mesoderm cells. The two signals are known to regulate local mesodermal cell fates and to signal to the endoderm. We find that, in addition, they precisely subdivide the abdA domain: Wg acts upon anterior abdA domain cells to activate pnt transcription, while Dpp is essential in the same region to prevent abdA from activating opa transcription. pnt activation is required to determine the appropriate numbers of mesodermal cells in the third midgut chamber.

  View details for Web of Science ID 000073880000021

  View details for PubMedID 9521915

• Towards development of an in vitro model of human BC formation. Oro, A. E., Fan, H., Khavari, P. A., Scott, M. P. NATURE PUBLISHING GROUP. 1998: 500–500

  View details for Web of Science ID 000072738200196

• Identification of Tinman regulated genes Weiss, J., Mellerick, D., Scott, M. P. FEDERATION AMER SOC EXP BIOL. 1998: A45–A45

  View details for Web of Science ID 000076006400263

• Hedgehog/patched signaling in animal development and disease Accomplishments in Cancer Research 1997, pp. 120-124. Eds. S.A. Wells and P.A. Sharp. Lippincott, Williams & Williams. Scott, M.P. 1998
• Transcriptional regulation of the Drosophila homeotic gene teashirt by the homeodomain protein Fushi tarazu MECHANISMS OF DEVELOPMENT Core, N., Charroux, B., McCormick, A., Vola, C., Fasano, L., Scott, M. P., Kerridge, S. 1997; 68 (1-2): 157-172

  Abstract

  The Drosophila melanogaster gene teashirt (tsh) is essential for segment identity of the embryonic thorax and abdomen. A deletion 3' to the tsh transcription unit causes the loss of tsh early expression in the even-numbered parasegments, and the corresponding larval cuticular patterns are disrupted. tsh function in the odd-numbered parasegments in these mutants is normal by both criteria. The in vivo activities of genomic fragments from the deleted region were tested in transgenic embryos. A 2.0 kb enhancer from the 3' region acts mainly in the even-numbered parasegments and is dependent on fushi tarazu (ftz) activity, which encodes a homeodomain protein required for the development of even-numbered parasegments. Ftz protein binds in vitro to four distinct sequences in a 220 bp sub-fragment; these and neighboring sequences are conserved in the equivalent enhancer isolated from Drosophila virilis. Tsh protein produced under the control of the 220 bp enhancer partially rescues a null tsh mutation, with its strongest effect in the even-numbered parasegments. Mutation of the Ftz binding sites partially abrogates the capacity for rescue. These results suggest a composite mechanism for regulation of tsh, with different activators such as ftz contributing to the overall pattern of expression of this key regulator.

  View details for Web of Science ID 000071085500014

  View details for PubMedID 9431813

• Targeted disruption of the mouse cystatin B gene. Pennacchio, L. A., Higgins, K. M., Scott, M. P., Myers, R. M. CELL PRESS. 1997: A343–A343

  View details for Web of Science ID 000071446002009

• Altered neural cell fates and medulloblastoma in mouse patched mutants SCIENCE Goodrich, L. V., Milenkovic, L., Higgins, K. M., Scott, M. P. 1997; 277 (5329): 1109-1113

  Abstract

  The PATCHED (PTC) gene encodes a Sonic hedgehog (Shh) receptor and a tumor suppressor protein that is defective in basal cell nevus syndrome (BCNS). Functions of PTC were investigated by inactivating the mouse gene. Mice homozygous for the ptc mutation died during embryogenesis and were found to have open and overgrown neural tubes. Two Shh target genes, ptc itself and Gli, were derepressed in the ectoderm and mesoderm but not in the endoderm. Shh targets that are, under normal conditions, transcribed ventrally were aberrantly expressed in dorsal and lateral neural tube cells. Thus Ptc appears to be essential for repression of genes that are locally activated by Shh. Mice heterozygous for the ptc mutation were larger than normal, and a subset of them developed hindlimb defects or cerebellar medulloblastomas, abnormalities also seen in BCNS patients.

  View details for Web of Science ID A1997XT03300039

  View details for PubMedID 9262482

• Hedgehog/patched signaling in animal development and disease Scott, M. P., Goodrich, L. V., Higgins, K. M., Johnson, R. L., Milenkovic, L., Oro, A. E. FEDERATION AMER SOC EXP BIOL. 1997: A1302–A1302

  View details for Web of Science ID 000073305603086

• A genetic and immunochemical analysis of the segment polarity gene naked (nkd). Wang, K., Mack, J., Scott, M. P. FEDERATION AMER SOC EXP BIOL. 1997: A1114–A1114

  View details for Web of Science ID 000073305601991

• Costal2, a novel kinesin-related protein in the Hedgehog signaling pathway CELL Sisson, J. C., Ho, K. S., Suyama, K., Scott, M. P. 1997; 90 (2): 235-245

  Abstract

  The Hedgehog (HH) signaling proteins control cell fates and patterning during animal development. In Drosophila, HH protein induces the transcription of target genes encoding secondary signals such as DPP and WG proteins by opposing a repressor system. The repressors include Costal2, protein kinase A, and the HH receptor Patched. Like HH, the kinase Fused and the transcription factor Cubitus interruptus (CI) act positively upon targets. Here we show that costal2 encodes a kinesin-related protein that accumulates preferentially in cells capable of responding to HH. COS2 is cytoplasmic and binds microtubules. We find that CI associates with COS2 in a large protein complex, suggesting that COS2 directly controls the activity of CI.

  View details for Web of Science ID A1997XN24900008

  View details for PubMedID 9244298

• Induction of basal cell carcinoma features in transgenic human skin expressing Sonic Hedgehog NATURE MEDICINE Fan, H. R., Oro, A. E., Scott, M. P., Khavari, P. A. 1997; 3 (7): 788-792

  Abstract

  Hedgehog (HH) signaling proteins mediate inductive events during animal development. Mutation of the only known HH receptor gene, Patched (PTC), has recently been implicated in inherited and sporadic forms of the most common human cancer, basal cell carcinoma (BCC). In Drosophila, HH acts by inactivating PTC function, raising the possibility that overexpression of Sonic Hedgehog (SHH) in human epidermis might have a tumorigenic effect equivalent to loss of PTC function. We used retroviral transduction of normal human keratinocytes to constitutively express SHH. SHH-expressing cells demonstrated increased expression of both the known HH target, BMP-2B, as well as bcl-2, a protein prominently expressed by keratinocytes in BCCs. These keratinocytes were then used to regenerate human skin transgenic for long terminal repeat-driven SHH (LTR-SHH) on immune-deficient mice. LTR-SHH human skin consistently displays the abnormal specific histologic features seen in BCCs, including downgrowth of epithelial buds into the dermis, basal cell palisading and separation of epidermis from the underlying dermis. In addition, LTR-SHH skin displays the gene expression abnormalities previously described for human BCCs, including decreased BP180/BPAG2 and laminin 5 adhesion proteins and expression of basal epidermal keratins. These data indicate that expression of SHH in human skin recapitulates features of human BCC in vivo, suggest that activation of this conserved signaling pathway contributes to the development of epithelial neoplasia and describe a new transgenic human tissue model of neoplasia.

  View details for Web of Science ID A1997XG76700042

  View details for PubMedID 9212109

• Mutations of the PATCHED gene in several types of sporadic extracutaneous tumors CANCER RESEARCH Xie, J. W., Johnson, R. L., Zhang, X. L., BARE, J. W., Waldman, F. M., Cogen, P. H., Menon, A. G., Warren, R. S., Chen, L. C., Scott, M. P., Epstein, E. H. 1997; 57 (12): 2369-2372

  Abstract

  Patients with basal cell nevus syndrome have a high incidence of multiple basal cell carcinomas, medulloblastomas, and meningiomas. Because somatic PATCHED (PTCH) mutations have been found in sporadic basal cell carcinomas, we have screened for PTCH mutations in several types of sporadic extracutaneous tumors. We found that 2 of 14 sporadic medulloblastomas bear somatic nonsense mutations in one copy of the gene and also deletion of the other copy. In addition, we identified missense mutations in PTCH in two of seven breast carcinomas, one of nine meningiomas, and one colon cancer cell line. No PTCH gene mutations were detected in 10 primary colon carcinomas and eighteen bladder carcinomas.

  View details for Web of Science ID A1997XD80000013

  View details for PubMedID 9192811

• NK-2/vnd patterns the developing CNS in Drosophila Mellerick, D. M., Weiss, J., Scott, M. P. ACADEMIC PRESS INC ELSEVIER SCIENCE. 1997: A200–A200

  View details for Web of Science ID A1997XH77400304

• Cell fate specification in the vertebrate neural tube: Shh receptors and the proto-oncogene GLI. Hynes, M., Stone, D., Armanini, M., Phillips, H., Johnson, R. L., Scott, M. P., Swanson, T., Pennica, D., Gu, Q., Goddard, A., Noll, M., Hooper, J., DeSauvage, F., Dowd, M., PITTSMEEK, S., Rosenthal, A. ACADEMIC PRESS INC ELSEVIER SCIENCE. 1997: B207–B207

  View details for Web of Science ID A1997XH77400590

• Costal2, a novel kinesin-related protein, genetically and biochemically interacts with other members of the hedgehog signalling pathway Ho, K. S., Sisson, J. C., Scott, M. P. ACADEMIC PRESS INC ELSEVIER SCIENCE. 1997: A91–A91

  View details for Web of Science ID A1997XH77400196

• Butterfly eyespots and the co-option of A-P patterning genes Keys, D. N., Lewis, D. L., Goodrich, L. V., Selegue, J., Gates, J., Scott, M. P., Carroll, S. B. ACADEMIC PRESS INC ELSEVIER SCIENCE. 1997: A32–A32

  View details for Web of Science ID A1997XH77400137

• The role of Patched in the developing mammalian nervous system Goodrich, L. V., Milenkovic, L., Higgins, K. M., Scott, M. P. ACADEMIC PRESS INC ELSEVIER SCIENCE. 1997: A89–A89

  View details for Web of Science ID A1997XH77400194

• Basal cell carcinomas in mice overexpressing sonic hedgehog SCIENCE Oro, A. E., Higgins, K. M., Hu, Z. L., Bonifas, J. M., Epstein, E. H., Scott, M. P. 1997; 276 (5313): 817-821

  Abstract

  Mutations in the tumor suppressor gene PATCHED (PTC) are found in human patients with the basal cell nevus syndrome, a disease causing developmental defects and tumors, including basal cell carcinomas. Gene regulatory relationships defined in the fruit fly Drosophila suggest that overproduction of Sonic hedgehog (SHH), the ligand for PTC, will mimic loss of ptc function. It is shown here that transgenic mice overexpressing SHH in the skin develop many features of basal cell nevus syndrome, demonstrating that SHH is sufficient to induce basal cell carcinomas in mice. These data suggest that SHH may have a role in human tumorigenesis.

  View details for Web of Science ID A1997WW90000064

  View details for PubMedID 9115210

• The role of hedgehog signaling in the pathogenesis of the basal cell news syndrome. Oro, A. E., Higgins, K. M., Scott, M. P. NATURE PUBLISHING GROUP. 1997: 44–44

  View details for Web of Science ID A1997WP04000069

• Hox genes in evolution: Protein surfaces and paralog groups TRENDS IN GENETICS Sharkey, M., Graba, Y., Scott, M. P. 1997; 13 (4): 145-151

  Abstract

  The clustered Hox genes, which encode homeodomain transcription factors, control cell fates along the anterior-posterior axis. Differences between Hox proteins cause differences between body parts. Vertebrates have 13 Hox subgroups, called paralog groups, which can be correlated with some of the insect and Amphioxus genes, and have remained distinctive for hundreds of millions of years. We identify characteristic residues that define the different paralog groups. Some paralog groups can be recognized by the homeodomain sequence alone; others only by using characteristic residues outside the homeodomain. Mapping characteristic residues onto the known homeodomain crystal structure reveals that most of the homeodomain amino acids that distinguish paralog groups are oriented away from the DNA, in positions where they might engage in protein-protein interactions.

  View details for Web of Science ID A1997WQ96900008

  View details for PubMedID 9097725

• Induction of features of basal cell epithelioma in transgenic human skin expressing sonic Hedgehog in vivo. Fan, H., Oro, A. E., Scott, M. P., Khavari, P. A. NATURE PUBLISHING GROUP. 1997: 38–38

  View details for Web of Science ID A1997WP04000063

• Hox genes, arms and the man NATURE GENETICS Scott, M. P. 1997; 15 (2): 117-118

  View details for Web of Science ID A1997WF08700005

  View details for PubMedID 9020831

• Summary: A common language COLD SPRING HARBOR SYMPOSIA ON QUANTITATIVE BIOLOGY Scott, M. P. 1997; 62: 555-562

  View details for Web of Science ID 000073570200061

  View details for PubMedID 9598389

• Control of cell growth and fate by patched genes Cold Spring Harbor Symposium on Quantitative Biology - Pattern Formation During Development Johnson, R. L., Scott, M. P. COLD SPRING HARBOR LAB PRESS, PUBLICATIONS DEPT. 1997: 205–215

  View details for Web of Science ID 000073570200025

  View details for PubMedID 9598353

• Regulation of Drosophila development by transcription factors and cell-cell signaling Exploring Genetic Mechanisms. P. Berg and M. Singer, Eds. Kornberg, T., M.P. Scott 1997
• The tumour-suppressor gene patched encodes a candidate receptor for Sonic hedgehog NATURE Stone, D. M., Hynes, M., Armanini, M., Swanson, T. A., Gu, Q. M., Johnson, R. L., Scott, M. P., Pennica, D., Goddard, A., Phillips, H., Noll, M., Hooper, J. E., DeSauvage, F., Rosenthal, A. 1996; 384 (6605): 129-134

  Abstract

  The protein Sonic hedgehog (Shh) controls patterning and growth during vertebrate development. Here we demonstrate that it binds Patched (vPtc), which has been identified as a tumour-suppressor protein in basal cell carcinoma, with high affinity. We show that Ptc can form a physical complex with a newly cloned vertebrate homologue of the Drosophila protein Smoothened (vSmo), and that vSmo is coexpressed with vPtc in many tissues but does not bind Shh directly. These findings, combined with available genetic evidence from Drosophila, support the hypothesis that Ptc is a receptor for Shh, and that vSmo could be a signalling component that is linked to Ptc.

  View details for Web of Science ID A1996VT33600055

  View details for PubMedID 8906787

• Spatial regulation of a zebrafish patched homologue reflects the roles of sonic hedgehog and protein kinase A in neural tube and somite patterning DEVELOPMENT Concordet, J. P., Lewis, K. E., Moore, J. W., Goodrich, L. V., Johnson, R. L., Scott, M. P., Ingham, P. W. 1996; 122 (9): 2835-2846

  Abstract

  Signalling by members of the Hedgehog family of secreted proteins plays a central role in the development of vertebrate and invertebrate embryos. In Drosophila, transduction of the Hedgehog signal is intimately associated with the activity of protein kinase A and the product of the segment polarity gene patched. We have cloned a homologue of patched from the zebrafish Danio rerio and analysed the spatiotemporal regulation of its transcription during embryonic development in both wild-type and mutant animals. We find a striking correlation between the accumulation of patched1 transcripts and cells responding to sonic hedgehog activity both in the neurectoderm and mesoderm, suggesting that like its Drosophila counterpart, patched1 is regulated by sonic hedgehog activity. Consistent with this interpretation, mis-expression of sonic hedgehog results in ectopic activation of patched1 transcription. Using dominant negative and constitutively active forms of the protein kinase A subunits, we also show that expression of patched1 as well as of other sonic hedgehog targets, is regulated by protein kinase A activity. Taken together, our findings suggest that the mechanism of signalling by Hedgehog family proteins has been highly conserved during evolution.

  View details for Web of Science ID A1996VJ07500026

  View details for PubMedID 8787757

• Antagonizing cAMP-dependent protein kinase A in the dorsal CNS activates a conserved Sonic hedgehog signaling pathway DEVELOPMENT Epstein, D. J., Marti, E., Scott, M. P., MCMAHON, A. P. 1996; 122 (9): 2885-2894

  Abstract

  Hedgehog (Hh) signaling plays a significant role in defining the polarity of a variety of tissue types along the anterior/posterior and dorsal/ventral axes in both vertebrate and invertebrate organisms. The pathway through which Hh transduces its signal is still obscure, however, recent data have implicated the cyclic AMP-dependent protein kinase A as a negative regulator of the Hh signal transduction pathway. One of the vertebrate Hh family members, Sonic hedgehog (Shh), can induce ventral neural cell types both in vivo and in vitro; high concentrations induce floor plate and lower concentrations motor neurons. To investigate whether PKA plays an active role in the suppression of ventral neural differentiation, we generated transgenic embryos expressing a dominant negative form of PKA (dnPKA) in primarily dorsal aspects of the mouse CNS. Similar to our earlier results with Shh, we observed the induction of floor plate and motor neuron markers in embryos expressing the dominant negative PKA transgene and the loss of dorsal gene expression at rostral levels. Thus suppression of PKA activity is sufficient to activate targets of the Shh signaling pathway in the vertebrate CNS suggesting that induction of ventral cell types occurs via the antagonistic action of Shh on PKA activity. Two mammalian target genes that are strongly expressed in ectopic dorsal locations in response to dnPKA are Ptc and Gli. As both of these are targets of Drosophila Hh signaling, our data point to an evolutionary conservation in both the mechanisms of signaling and the effectors of the signaling pathway.

  View details for Web of Science ID A1996VJ07500030

  View details for PubMedID 8787761

• Human homolog of patched, a candidate gene for the basal cell nevus syndrome SCIENCE Johnson, R. L., Rothman, A. L., Xie, J. W., Goodrich, L. V., BARE, J. W., Bonifas, J. M., Quinn, A. G., Myers, R. M., Cox, D. R., Epstein, E. H., Scott, M. P. 1996; 272 (5268): 1668-1671

  Abstract

  The basal cell nevus syndrome (BCNS) is characterized by developmental abnormalities and by the postnatal occurrence of cancers, especially basal cell carcinomas (BCCs), the most common human cancer. Heritable mutations in BCNS patients and a somatic mutation in a sporadic BCC were identified in a human homolog of the Drosophila patched (ptc) gene. The ptc gene encodes a transmembrane protein that in Drosophila acts in opposition to the Hedgehog signaling protein, controlling cell fates, patterning, and growth in numerous tissues. The human PTC gene appears to be crucial for proper embryonic development and for tumor suppression.

  View details for Web of Science ID A1996UR09300052

  View details for PubMedID 8658145

• Conservation in hedgehog signaling: Induction of a chicken patched homolog by Sonic hedgehog in the developing limb DEVELOPMENT Marigo, V., Scott, M. P., Johnson, R. L., Goodrich, L. V., Tabin, C. J. 1996; 122 (4): 1225-1233

  Abstract

  Hedgehog genes have been implicated in inductive signaling during development in a variety of organisms. A key element of the hedgehog signaling system is encoded by the gene patched. In Drosophila hedgehog regulates gene expression by antagonizing the action of patched. In addition, patched is itself a transcriptional target of hedgehog signaling. We have isolated a chicken patched homolog and find it to be strongly expressed adjacent to all tissues where members of the hedgehog family are expressed. As in Drosophila, ectopic expression of Sonic hedgehog leads to ectopic induction of chicken Patched. Based on this regulatory conservation, vertebrate Patched is likely to be directly downstream of Sonic hedgehog signaling. An important role of Sonic hedgehog is the regulation of anterior/posterior pattern in the developing limb bud. Since Patched is directly downstream of the hedgehog signal, the extent of high level Patched expression provides a measure of the distance that Sonic hedgehog diffuses and directly acts. On this basis, we find that Sonic hedgehog directly acts as a signal over only the posterior third of the limb bud. During limb patterning, secondary signals are secreted in both the mesoderm (e.g. Bone Morphogenetic Protein-2) and apical ectodermal ridge (e.g. Fibroblast Growth Factor-4) in response to Sonic hedgehog. Thus knowing which is the direct target tissue is essential for unraveling the molecular patterning of the limb. The expression of Patched provides a strong indication that the mesoderm and not the ectoderm is the direct target of Sonic hedgehog signaling in the limb bud. Finally we demonstrate that induction of Patched requires Sonic hedgehog but, unlike Bone Morphogenetic Protein-2 and Hox genes, does not require Fibroblast Growth Factor as a co-inducer. It is therefore a more direct target of Sonic hedgehog than previously reported patterning genes.

  View details for Web of Science ID A1996UF83100018

  View details for PubMedID 8620849

• Conservation of the hedgehog/patched signaling pathway from flies to mice: Induction of a mouse patched gene by Hedgehog GENES & DEVELOPMENT Goodrich, L. V., Johnson, R. L., Milenkovic, L., McMahon, J. A., Scott, M. P. 1996; 10 (3): 301-312

  Abstract

  The signaling protein Hedgehog (Hh) controls cell fate and polarizes tissues in both flies and vertebrates. In flies, Hh exerts its effects by opposing the function of a novel transmembrane protein, Patched, while also locally inducing patched (ptc) transcription. We have identified a mouse homolog of ptc which in many tissues is transcribed near cells making either Sonic or Indian hedgehog. In addition, ectopic Sonic hedgehog expression in the mouse central nervous system induces ptc transcription. As in flies, mouse ptc transcription appears to be indicative of hedgehog signal reception. The results support the existence of a conserved signaling pathway used for pattern formation in insects and mammals.

  View details for Web of Science ID A1996TV71100006

  View details for PubMedID 8595881

• patched overexpression alters wing disc size and pattern: Transcriptional and post-transcriptional effects on hedgehog targets DEVELOPMENT Johnson, R. L., Grenier, J. K., Scott, M. P. 1995; 121 (12): 4161-4170

  Abstract

  The membrane protein, Patched, plays a critical role in patterning embryonic and imaginal tissues in Drosophila. patched constitutively inactivates the transcription of target genes such as wingless, decapentaplegic, and patched itself. The secreted protein, Hedgehog, induces transcription of target genes by opposing the Patched signaling pathway. Using the Gal4 UAS system we have overexpressed patched in wing imaginal discs and found that high Patched levels, expressed in either normal or ectopic patterns, result in loss of wing vein patterning in both compartments centering at the anterior/posterior border. In addition, patched inhibits the formation of the mechanosensory neurons, the campaniform sensilla, in the wing blade. The patched wing vein phenotype is modulated by mutations in hedgehog and cubitus interruptus (ci). Patched overexpression inhibits transcription of patched and decapentaplegic and post-transcriptionally decreases the amount of Ci protein at the anterior/posterior boundary. In hedgehogMrt wing discs, which express ectopic hedgehog, Ci levels are correspondingly elevated, suggesting that hedgehog relieves patched repression of Ci accumulation. Protein kinase A also regulates Ci; protein kinase A mutant clones in the anterior compartment have increased levels of Ci protein. Thus patched influences wing disc patterning by decreasing Ci protein levels and inactivating hedgehog target genes in the anterior compartment.

  View details for Web of Science ID A1995TM47800024

  View details for PubMedID 8575316

• GENOMIC REGIONS REQUIRED FOR MORPHOGENESIS OF THE DROSOPHILA EMBRYONIC MIDGUT GENETICS Bilder, D., Scott, M. P. 1995; 141 (3): 1087-1100

  Abstract

  The Drosophila midgut is an excellent system for studying the cell migration, cell-cell communication, and morphogenetic events that occur in organ formation. Genes representative of regulatory gene families common to all animals, including homeotic, TGF beta, and Wnt genes, play roles in midgut development. To find additional regulators of midgut morphogenesis, we screened a set of genomic deficiencies for midgut phenotypes. Fifteen genomic intervals necessary for proper midgut morphogenesis were identified, three contain genes already known to act in the midgut. Three other genomic regions are required for formation of the endoderm or visceral mesoderm components of the midgut. Nine regions are required for proper formation of the midgut constrictions. The E75 ecdysone-induced gene, which encodes a nuclear receptor superfamily member, is the relevant gene in one region and is essential for proper formation of midgut constrictions. E75 acts downstream of the previously known constriction regulators or in parallel. Temporal hormonal control may therefore work in conjunction with spatial regulation by the homeotic genes in midgut development. Another genomic region is required to activate transcription of the homeotic genes Antp and Scr specifically in visceral mesoderm. The genomic regions identified by this screen provide a map to novel midgut development regulators.

  View details for Web of Science ID A1995TB62900026

  View details for PubMedID 8582615

  View details for PubMedCentralID PMC1206832

• SELECTION AND CHARACTERIZATION OF SEQUENCES WITH HIGH-AFFINITY FOR THE ENGRAILED PROTEIN OF DROSOPHILA MECHANISMS OF DEVELOPMENT SAENZROBLES, M. T., Maschat, F., Tabata, T., Scott, M. P., Kornberg, T. B. 1995; 53 (2): 185-195

  Abstract

  The engrailed gene helps to direct Drosophila melanogaster development by encoding a homeodomain-containing DNA binding protein. To identify genes whose transcription engrailed regulates, we developed a method to isolate genomic sequences to which engrailed protein binds with high affinity. Fragments of genomic DNA were fractionated on an engrailed protein affinity column, and fragments that were retained in the presence of 0.4-1.0 M KCl were isolated and cloned. The isolated fragments include regions of the engrailed and cubitus interruptus gene promoters, both of which are candidate targets of engrailed, and most fragments contain regions that engrailed protein protects from DNaseI digestion. Chromosomal deletions that remove some of the engrailed binding sites (located either at 64D, 96B or 99D) interact genetically with engrailed. Characterization of a transcript encoded in region 64D revealed its dependence on engrailed protein.

  View details for Web of Science ID A1995TA28400003

  View details for PubMedID 8562421

• HOMEOTIC RESPONSE ELEMENTS ARE TIGHTLY LINKED TO TISSUE-SPECIFIC ELEMENTS IN A TRANSCRIPTIONAL ENHANCER OF THE TEASHIRT GENE DEVELOPMENT MCCORMICK, A. M., Core, N., Kerridge, S., Scott, M. P. 1995; 121 (9): 2799-2812

  Abstract

  Along the anterior-posterior axis of animal embryos, the choice of cell fates, and the organization of morphogenesis, is regulated by transcription factors encoded by clustered homeotic or 'Hox' genes. Hox genes function in both epidermis and internal tissues by regulating the transcription of target genes in a position- and tissue-specific manner. Hox proteins can have distinct targets in different tissues; the mechanisms underlying tissue and homeotic protein specificity are unknown. Light may be shed by studying the organization of target gene enhancers. In flies, one of the target genes is teashirt (tsh), which encodes a zinc finger protein. tsh itself is a homeotic gene that controls trunk versus head development. We identified a tsh gene enhancer that is differentially activated by Hox proteins in epidermis and mesoderm. Sites where Antennapedia (Antp) and Ultrabithorax (Ubx) proteins bind in vitro were mapped within evolutionarily conserved sequences. Although Antp and Ubx bind to identical sites in vitro, Antp activates the tsh enhancer only in epidermis while Ubx activates the tsh enhancer in both epidermis and in somatic mesoderm. We show that the DNA elements driving tissue-specific transcriptional activation by Antp and Ubx are separable. Next to the homeotic protein-binding sites are extensive conserved sequences likely to control tissue activation by different homeodomain proteins. We propose that local interactions between homeotic proteins and other factors effect activation of targets in proper cell types.

  View details for Web of Science ID A1995RV11900009

  View details for PubMedID 7555708

• COORDINATION OF LIMB PATTERNING BY SONIC HEDGEHOG AND FGF-4 Laufer, E., Marigo, V., Nelson, C., Johnson, R. L., Scott, M. P., Tabin, C. ACADEMIC PRESS INC ELSEVIER SCIENCE. 1995: 739–39

  View details for Web of Science ID A1995RQ08400058

• THE DROSOPHILA SNR1 AND BRM PROTEINS ARE RELATED TO YEAST SWI/SNF PROTEINS AND ARE COMPONENTS OF A LARGE PROTEIN COMPLEX MOLECULAR BIOLOGY OF THE CELL Dingwall, A. K., Beek, S. J., McCallum, C. M., Tamkun, J. W., Kalpana, G. V., Goff, S. P., Scott, M. P. 1995; 6 (7): 777-791

  Abstract

  During most of Drosophila development the regulation of homeotic gene transcription is controlled by two groups of regulatory genes, the trithorax group of activators and the Polycomb group of repressors. brahma (brm), a member of the trithorax group, encodes a protein related to the yeast SWI2/SNF2 protein, a subunit of a protein complex that assists sequence-specific activator proteins by alleviating the repressive effects of chromatin. To learn more about the molecular mechanisms underlying the regulation of homeotic gene transcription, we have investigated whether a similar complex exists in flies. We identified the Drosophila snr1 gene, a potential homologue of the yeast SNF5 gene that encodes a subunit of the yeast SWI/SNF complex. The snr1 gene is essential and genetically interacts with brm and trithorax (trx), suggesting cooperation in regulating homeotic gene transcription. The spatial and temporal patterns of expression of snr1 are similar to those of brm. The snr1 and brm proteins are present in a large (> 2 x 10(6) Da) complex, and they co-immunoprecipitate from Drosophila extracts. These findings provide direct evidence for conservation of the SWI/SNF complex in higher eucaryotes and suggest that the Drosophila brm/snr1 complex plays an important role in maintaining homeotic gene transcription during development by counteracting the repressive effects of chromatin.

  View details for Web of Science ID A1995RH68700003

  View details for PubMedID 7579694

  View details for PubMedCentralID PMC301240

• INTIMATIONS OF A CREATURE CELL Scott, M. P. 1994; 79 (7): 1121-1124

  View details for Web of Science ID A1994PZ86000001

  View details for PubMedID 8001147

• REGULATION OF A DECAPENTAPLEGIC MIDGUT ENHANCER BY HOMEOTIC PROTEINS DEVELOPMENT Manak, J. R., Mathies, L. D., Scott, M. P. 1994; 120 (12): 3605-3619

  Abstract

  The clustered homeotic genes encode transcription factors that regulate pattern formation in all animals, conferring cell fates by coordinating the activities of downstream 'target' genes. In the Drosophila midgut, the Ultrabithorax (Ubx) protein activates and the abdominalA (abd-A) protein represses transcription of the decapentaplegic (dpp) gene, which encodes a secreted signalling protein of the TGF beta class. We have identified an 813 bp dpp enhancer which is capable of driving expression of a lacZ gene in a correct pattern in the embryonic midgut. The enhancer is activated ectopically in the visceral mesoderm by ubiquitous expression of Ubx or Antennapedia but not by Sex combs reduced protein. Ectopic expression of abd-A represses the enhancer. Deletion analysis reveals regions required for repression and activation. A 419 bp subfragment of the 813 bp fragment also drives reporter gene expression in an appropriate pattern, albeit more weakly. Evolutionary sequence conservation suggests other factors work with homeotic proteins to regulate dpp. A candidate cofactor, the extradenticle protein, binds to the dpp enhancer in close proximity to homeotic protein binding sites. Mutation of either this site or another conserved motif compromises enhancer function. A 45 bp fragment of DNA from within the enhancer correctly responds to both UBX and ABD-A in a largely tissue-specific manner, thus representing the smallest in vivo homeotic response element (HOMRE) identified to date.

  View details for Web of Science ID A1994QB20000024

  View details for PubMedID 7821226

• ROLE OF THE TEASHIRT GENE IN DROSOPHILA MIDGUT MORPHOGENESIS - SECRETED PROTEINS MEDIATE THE ACTION OF HOMEOTIC GENES DEVELOPMENT Mathies, L. D., Kerridge, S., Scott, M. P. 1994; 120 (10): 2799-2809

  Abstract

  Homeotic genes control the development of embryonic structure by coordinating the activities of downstream 'target' genes. The identities and functions of target genes must be understood in order to learn how homeotic genes control morphogenesis. Drosophila midgut development is regulated by homeotic genes expressed in the visceral mesoderm, where two of their target genes have been identified. Both encode secreted proteins. The Ultrabithorax (Ubx) homeotic gene activates transcription of the decapentaplegic (dpp) gene, which encodes a TGF beta class protein, while in adjacent mesoderm cells the abdominal-A (abd-A) homeotic gene activates transcription of the wingless (wg) gene, which encodes a Wnt class protein. The homeotic genes Antennapedia (Antp) and Sex combs reduced (Scr) act in more anterior midgut regions. Here we report the identification of another homeotic gene target in the midgut mesoderm, the teashirt (tsh) gene, which encodes a protein with zinc finger motifs. tsh is necessary for proper formation of anterior and central midgut structures. Antp activates tsh in anterior midgut mesoderm. In the central midgut mesoderm Ubx, abd-A, dpp, and wg are required for proper tsh expression. The control of tsh by Ubx and abd-A, and probably also by Antp, is mediated by secreted signaling molecules. By responding to signals as well as localized transcription regulators, the tsh transcription factor is produced in a spatial pattern distinct from any of the homeotic genes.

  View details for Web of Science ID A1994PN07600006

  View details for PubMedID 7607071

• PATCHED OVEREXPRESSION CAUSES LOSS OF WINGLESS EXPRESSION IN DROSOPHILA EMBRYOS DEVELOPMENTAL BIOLOGY Schuske, K., Hooper, J. E., Scott, M. P. 1994; 164 (1): 300-311

  Abstract

  The patched (ptc) segment polarity gene of Drosophila encodes a transmembrane protein involved in cell signaling that establishes pattern within the segment. In the posterior half of the parasegment Patched protein represses transcription of the wingless (wg) gene by an unknown mechanism. In the most posterior row of cells in each parasegment this repression is neutralized by a signal possibly carried by the product of the hedgehog gene, allowing wg expression. High levels of Patched expression might therefore overcome the repression and repress wg in all cells. Here we use a heat shock-inducible promoter to transiently express high levels of Patched in all cells. A single pulse of Patched transgene expression has little or no effect on the segmental pattern, as has been previously reported. Repeated pulses of Patched production drastically alter the segment pattern to mimic embryos lacking one of the wg class of segment polarity genes. We observe repression of wg and gooseberry (a wg class gene) transcription in the germband ectoderm but not in the head. Expression of two other segment polarity genes, engrailed and cubitus interruptus, is unaffected. Thus excess Patched is capable of overcoming the neutralizing signal.

  View details for Web of Science ID A1994NW44400026

  View details for PubMedID 8026632

• 5 SWI/SNF GENE-PRODUCTS ARE COMPONENTS OF A LARGE MULTISUBUNIT COMPLEX REQUIRED FOR TRANSCRIPTIONAL ENHANCEMENT PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Peterson, C. L., Dingwall, A., Scott, M. P. 1994; 91 (8): 2905-2908

  Abstract

  The Saccharomyces cerevisiae SWI1, SWI2 (SNF2), SWI3, SNF5, and SNF6 gene products play a crucial role in the regulation of transcription. We provide here direct biochemical evidence that all five SWI/SNF polypeptides are components of a large multisubunit complex. These five polypeptides coelute from a gel-filtration column with an apparent molecular mass of approximately 2 MDa. The five SWI/SNF polypeptides do not copurify when extracts are prepared from swi- or snf- mutants. We show that SWI/SNF polypeptides also remain associated during an affinity-chromatography step followed by gel filtration. Assembly of the SWI/SNF complex is not disrupted by a mutation in the putative APT-binding site of SWI2, although this mutation eliminates SWI2 function.

  View details for Web of Science ID A1994NF96800009

  View details for PubMedID 8159677

  View details for PubMedCentralID PMC43482

• SETTING LIMITS ON HOMEOTIC GENE-FUNCTION - RESTRAINT OF SEX-COMBS-REDUCED ACTIVITY BY TEASHIRT AND OTHER HOMEOTIC GENES EMBO JOURNAL Andrew, D. J., Horner, M. A., PETITT, M. G., Smolik, S. M., Scott, M. P. 1994; 13 (5): 1132-1144

  Abstract

  Each of the homeotic genes of the HOM or HOX complexes is expressed in a limited domain along the anterior-posterior axis. Each homeotic protein directs the formation of characteristic structures, such as wings or ribs. In flies, when a heat shock-inducible homeotic gene is used to produce a homeotic protein in all cells of the embryo, only some cells respond by altering their fates. We have identified genes that limit where the homeotic gene Sex combs reduced (Scr) can affect cell fates in the Drosophila embryo. In the abdominal cuticle Scr is prevented from inducing prothoracic structures by the three bithorax complex (BX-C) homeotic genes. However, two of the BX-C homeotic genes, Ultrabithorax (Ubx) and abdominal-A (abd-A), have no effect on the ability of Scr to direct the formation of salivary glands. Instead, salivary gland induction by Scr is limited in the trunk by the homeotic gene teashirt (tsh) and in the last abdominal segment by the third BX-C gene, Abdominal-B (AbdB). Therefore, spatial restrictions on homeotic gene activity differ between tissues and result both from the regulation of homeotic gene transcription and from restraints on where homeotic proteins can function.

  View details for PubMedID 7907545

• IMMUNOLOGICAL METHODS FOR MAPPING PROTEIN DISTRIBUTIONS ON POLYTENE CHROMOSOMES METHODS IN CELL BIOLOGY, VOL 44 Andrew, D. J., Scott, M. P. 1994; 44: 353-370

  View details for Web of Science ID A1994BC27U00021

  View details for PubMedID 7707963

• A CLASS ACT - CONSERVATION OF HOMEODOMAIN PROTEIN FUNCTIONS DEVELOPMENT Manak, J. R., Scott, M. P. 1994: 61-77

  Abstract

  Dramatic successes in identifying vertebrate homeobox genes closely related to their insect relatives have led to the recognition of classes within the homeodomain superfamily. To what extent are the homeodomain protein classes dedicated to specific functions during development? Although information on vertebrate gene functions is limited, existing evidence from mice and nematodes clearly supports conservation of function for the Hox genes. Less compelling, but still remarkable, is the conservation of other homeobox gene classes and of regulators of homeotic gene expression and function. It is too soon to say whether the cases of conservation are unique and exceptional, or the beginning of a profoundly unified view of gene regulation in animal development. In any case, new questions are raised by the data: how can the differences between mammals and insects be compatible with conservation of homeobox gene function? Did the evolution of animal form involve a proliferation of new homeodomain proteins, new modes of regulation of existing gene types, or new relationships with target genes, or is evolutionary change largely the province of other classes of genes? In this review, we summarize what is known about conservation of homeobox gene function.

  View details for Web of Science ID A1994QK89700008

  View details for PubMedID 7579525

• ECTOPIC EXPRESSION AND FUNCTION OF THE ANTP AND SCR HOMEOTIC GENES - THE N-TERMINUS OF THE HOMEODOMAIN IS CRITICAL TO FUNCTIONAL SPECIFICITY DEVELOPMENT Zeng, W. L., Andrew, D. J., Mathies, L. D., Horner, M. A., Scott, M. P. 1993; 118 (2): 339-352

  Abstract

  The transcription factors encoded by homeotic genes determine cell fates during development. Each homeotic protein causes cells to follow a distinct pathway, presumably by differentially regulating downstream 'target' genes. The homeodomain, the DNA-binding part of homeotic proteins, is necessary for conferring the specificity of each homeotic protein's action. The two Drosophila homeotic proteins encoded by Antennapedia and Sex combs reduced determine cell fates in the epidermis and internal tissues of the posterior head and thorax. Genes encoding chimeric Antp/Scr proteins were introduced into flies and their effects on morphology and target gene regulation observed. We find that the N terminus of the homeodomain is critical for determining the specific effects of these homeotic proteins in vivo, but other parts of the proteins have some influence as well. The N-terminal part of the homeodomain has been observed, in crystal structures and in NMR studies in solution, to contact the minor groove of the DNA. The different effects of Antennapedia and Sex combs reduced proteins in vivo may depend on differences in DNA binding, protein-protein interactions, or both.

  View details for PubMedID 7900989

• GENE-REGULATION - ABLE ASSISTANTS FOR HOMEODOMAIN PROTEINS CURRENT BIOLOGY Manak, J. R., Scott, M. P. 1993; 3 (5): 318-320

  View details for Web of Science ID A1993LL95700018

  View details for PubMedID 15335761

• A RATIONAL NOMENCLATURE FOR VERTEBRATE HOMEOBOX (HOX) GENES NUCLEIC ACIDS RESEARCH Scott, M. P. 1993; 21 (8): 1687-1688

  Abstract

  Developmental fates along the anterior-posterior axes of animals are controlled by clustered homeotic genes which in vertebrates are called Hox genes. The gene clusters are similar and probably functionally homologous in animals as different as nematodes, flies, and mammals. A new set of names for Hox genes was recently agreed upon by many workers in the field. Remarkably, the order of the Hox genes along the chromosome reflects where they are expressed along the body axis. This simple principle is reflected in the new nomenclature system.

  View details for Web of Science ID A1993LC21000001

  View details for PubMedID 8098522

  View details for PubMedCentralID PMC309401

• VERTEBRATE HOMEOBOX GENE NOMENCLATURE CELL Scott, M. P. 1992; 71 (4): 551-553

  View details for Web of Science ID A1992JY67600003

  View details for PubMedID 1358459

• COMPARATIVE-STUDIES OF DROSOPHILA ANTENNAPEDIA GENES GENETICS Hooper, J. E., PEREZALONSO, M., Bermingham, J. R., Prout, M., Rocklein, B. A., Wagenbach, M., Edstrom, J. E., DEFRUTOS, R., Scott, M. P. 1992; 132 (2): 453-469

  Abstract

  The Antennapedia (Antp) homeotic gene of Drosophila melanogaster controls cell fates and pattern formation in the epidermis, nervous system and mesoderm of thoracic segments. Its expression is controlled at the levels of transcription, alternative RNA splicing, polyadenylation and translation. Two nested Antp transcription units extend over 103 kb and produce sixteen different transcripts. We have compared the Antp genes of Drosophila virilis, Drosophila subobscura and D. melanogaster to determine which structural features are conserved and therefore may be important to the gene's function. The overall gene structures are similar. There are many conserved sequence blocks throughout the large introns, at least 15 kb upstream of the first promoter, and at least 3 kb downstream of the last polyadenylation site. Intron and exon sequence conservation around alternative splice sites indicates that alternative protein coding forms may also be conserved. Protein coding potential is perfectly conserved around the C-terminal homeodomain, well conserved in the N-terminal region, and more variable in the middle. The large size of the Antp gene may reflect a large number of control elements necessary for appropriate Antp protein expression. The conservation of transcript complexity suggests functional requirements for the different protein forms.

  View details for Web of Science ID A1992JQ14600015

  View details for PubMedID 1358753

• HOMEOTIC GENE ANTENNAPEDIA MESSENGER-RNA CONTAINS 5'-NONCODING SEQUENCES THAT CONFER TRANSLATIONAL INITIATION BY INTERNAL RIBOSOME BINDING GENES & DEVELOPMENT Oh, S. K., Scott, M. P., Sarnow, P. 1992; 6 (9): 1643-1653

  Abstract

  The Antennapedia (Antp) homeotic gene of Drosophila melanogaster has two promoters, P1 and P2. The resulting Antp mRNAs contain 1512-nucleotide (P1) and 1727-nucleotide (P2) 5'-noncoding regions, composed of exons A, B, D, and E (P1) or exons C, D, and E (P2), respectively. Multiple AUG codons are present in exons A, B, and C. We have found that 252-nucleotide exon D, common to mRNAs from both transcription units and devoid of AUG codons, can mediate initiation of translation by internal ribosome binding in cultured cells. Many mRNAs in Drosophila contain long 5'-noncoding regions with apparently unused AUG codons, suggesting that internal ribosome binding may be a common mechanism of translational initiation, and possibly its regulation, in Drosophila.

  View details for Web of Science ID A1992JM56900005

  View details for PubMedID 1355457

• BRAHMA - A REGULATOR OF DROSOPHILA HOMEOTIC GENES STRUCTURALLY RELATED TO THE YEAST TRANSCRIPTIONAL ACTIVATOR SNF2 SW12 CELL Tamkun, J. W., Deuring, R., Scott, M. P., Kissinger, M., Pattatucci, A. M., Kaufman, T. C., Kennison, J. A. 1992; 68 (3): 561-572

  Abstract

  The brahma (brm) gene is required for the activation of multiple homeotic genes in Drosophila. Loss-of-function brm mutations suppress mutations in Polycomb, a repressor of homeotic genes, and cause developmental defects similar to those arising from insufficient expression of the homeotic genes of the Antennapedia and Bithorax complexes. The brm gene encodes a 1638 residue protein that is similar to SNF2/SWI2, a protein involved in transcriptional activation in yeast, suggesting possible models for the role of brm in the transcriptional activation of homeotic genes. In addition, both brm and SNF2 contain a 77 amino acid motif that is found in other Drosophila, yeast, and human regulatory proteins and may be characteristic of a new family of regulatory proteins.

  View details for Web of Science ID A1992HD39800015

  View details for PubMedID 1346755

• Gene control systems affecting insect development. (Eds. J.M. Crampton and P. Eggleston). in Insect Molecular Science, 16th Symp. Roy. Entomol. Soc. Petitt, M.G., M.P. Scott 1992
• DOWNSTREAM OF THE HOMEOTIC GENES NEW BIOLOGIST Andrew, D. J., Scott, M. P. 1992; 4 (1): 5-15

  Abstract

  The homeotic genes of Drosophila melanogaster determine which structures form in each of the body segments. Disrupting the function of the homeotic genes causes body parts found in one domain of the animal to be replaced by body parts normally found elsewhere. Each of the homeotic genes encodes a protein, or a closely related family of proteins, which is capable of binding DNA and controlling the transcriptional activities of downstream genes. The homeotic genes are in the middle of a complex regulatory network, and many of the genes that control homeotic expression have been well characterized. However, very little is known about what comes after the homeotic genes, the downstream genes whose activities are regulated by the homeotic genes. Here, we review the known relationships between the homeotic proteins and the few identified target genes. The details of these interactions may be characteristic and may thus guide the search for additional targets.

  View details for PubMedID 1346973

• The molecular genetic basis of positional information in insect segments. Results and problems in cell differentiation Hooper, J. E., Scott, M. P. 1992; 18: 1-48

  View details for PubMedID 1475532

• Editorial CURRENT OPINION IN GENETICS & DEVELOPMENT Laskey, R., Scott, M. P. 1991; 1 (1): 1

  View details for DOI 10.1016/0959-437X(91)80030-P

  View details for Web of Science ID 000450881200001

• THE PROSPERO GENE SPECIFIES CELL FATES IN THE DROSOPHILA CENTRAL-NERVOUS-SYSTEM CELL Doe, C. Q., CHULAGRAFF, Q., Wright, D. M., Scott, M. P. 1991; 65 (3): 451-464

  Abstract

  The molecular mechanisms used to generate neuronal diversity are largely unknown. To identify genes controlling cell fate in the Drosophila central nervous system, we screened for mutations that alter expression of homeobox genes in the developing central nervous system (indicating changes in cell fates). We also screened "enhancer trap" lines to identify genes expressed in neuronal stem cells (neuroblasts). The prospero gene was discovered in both screens. prospero is expressed in a subset of neuroblasts, sensory neuron precursors, and identified glial precursors. It is not expressed in neurons. Neuroblasts lacking prospero function generate abnormal cell lineages, producing incorrectly specified progeny that differentiate into neurons showing axon pathfinding defects. prospero is therefore a novel type of gene expressed in neuroblasts and known to specify neuronal fate.

  View details for Web of Science ID A1991FK18200012

  View details for PubMedID 1673362

• THE ARFLIKE GENE ENCODES AN ESSENTIAL GTP-BINDING PROTEIN IN DROSOPHILA PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Tamkun, J. W., Kahn, R. A., Kissinger, M., Brizuela, B. J., RULKA, C., Scott, M. P., Kennison, J. A. 1991; 88 (8): 3120-3124

  Abstract

  We have identified a Drosophila gene (arflike, arl) encoding a protein that is structurally related (approximately 55% identity) to the ADP-ribosylation factors (ARFs) of yeast and mammals. Biochemical analyses of purified recombinant arl-encoded protein revealed properties similar to the ARF proteins, including the ability to bind and hydrolyze GTP. Clear functional differences between arl and ARF proteins, including a complete lack of ARF activity, suggest that arl is not a functional homolog of ARF. A recessive lethal arl mutation was recovered, demonstrating that the arl locus is an essential gene. We conclude that the arl locus encodes an essential member of the ARF subfamily of small GTP-binding proteins in Drosophila.

  View details for Web of Science ID A1991FG91300033

  View details for PubMedID 1901655

• REPRESSION OF THE DROSOPHILA-FUSHI-TARAZU (FTZ) SEGMENTATION GENE EMBO JOURNAL Brown, J. L., Sonoda, S., Ueda, H., Scott, M. P., Wu, C. 1991; 10 (3): 665-674

  Abstract

  The striped expression of the Drosophila segmentation gene fushi tarazu in alternate parasegments of the early embryo is controlled by the 740 bp zebra element. Among multiple protein factors that bind to the zebra element, FTZ-F2 behaves as a transcriptional repressor of ftz. Point mutations in the zebra element which disrupt FTZ-F2 binding to DNA cause ectopic expression of zebra-lacZ activity in transformed embryos. The mutant constructs are expressed from the zygotic genome in preblastoderm embryos as early as the third nuclear division cycle. This unprecedented early transcription suggests that ftz requires active repression during initial nuclear division cycles, a novel type of embryonic gene regulation. A putative FTZ-F2 cDNA clone isolated by recognition site screening of an expression library was found to be identical in sequence with the zinc finger protein tramtrack (Harrison and Travers, 1990).

  View details for Web of Science ID A1991FB41300021

  View details for PubMedID 2001679

• A DROSOPHILA GROWTH-FACTOR HOMOLOG, DECAPENTAPLEGIC, REGULATES HOMEOTIC GENE-EXPRESSION WITHIN AND ACROSS GERM LAYERS DURING MIDGUT MORPHOGENESIS DEVELOPMENT PANGANIBAN, G. E., Reuter, R., Scott, M. P., HOFFMANN, F. M. 1990; 110 (4): 1041-1050

  Abstract

  The decapentaplegic (dpp) gene product, a member of the transforming growth factor-beta family, is required in Drosophila embryos for normal gastrulation and the establishment of dorsal-ventral polarity in the embryo. dpp is also expressed at specific positions in the visceral mesoderm along the developing midgut. We find that mutations that eliminate the visceral mesoderm expression of dpp lead to defects in midgut morphogenesis and alter the spatially localized expression of the homeotic genes Sex combs reduced (Scr), Ultrabithorax (Ubx), and Antennapedia (Antp) in the visceral mesoderm. The extracellular dpp protein migrates from the visceral mesoderm across the apposing endodermal cell layer in a region of the endoderm that expresses the homeotic gene labial (lab). Mesodermal expression of dpp is required for the expression of lab in these endodermal cells indicating that dpp mediates an inductive interaction between the two germ layers. We propose that extracellular dpp protein regulates gut morphogenesis, in part, by regulating homeotic gene expression in the visceral mesoderm and endoderm of the developing midgut.

  View details for Web of Science ID A1990EQ22300004

  View details for PubMedID 1983114

• HOMEOTIC GENES REGULATE THE SPATIAL EXPRESSION OF PUTATIVE GROWTH-FACTORS IN THE VISCERAL MESODERM OF DROSOPHILA EMBRYOS DEVELOPMENT Reuter, R., PANGANIBAN, G. E., HOFFMANN, F. M., Scott, M. P. 1990; 110 (4): 1031-1040

  Abstract

  During Drosophila embryogenesis homeotic genes control the developmental diversification of body structures. The genes probably coordinate the expression of as yet unidentified target genes that carry out cell differentiation processes. At least four homeotic genes expressed in the visceral mesoderm are required for midgut morphogenesis. In addition, two growth factor homologs are expressed in specific regions of the visceral mesoderm surrounding the midgut epithelium. One of these, decapentaplegic (dpp), is a member of the transforming growth factor beta (TGF-beta) family; the other, wingless (wg), is a relative of the mammalian proto-oncogene int-1. Here we show that the spatially restricted expression of dpp in the visceral mesoderm is regulated by the homeotic genes Ubx and abd-A. Ubx is required for the expression of dpp while abd-A represses dpp. One consequence of dpp expression is the induction of labial (lab) in the underlying endoderm cells. In addition, abd-A function is required for the expression of wg in the visceral mesoderm posterior to the dpp-expressing cells. The two growth factor genes therefore are excellent candidates for target genes that are directly regulated by the homeotic genes.

  View details for Web of Science ID A1990EQ22300003

  View details for PubMedID 1983113

• WHAT DETERMINES THE SPECIFICITY OF ACTION OF DROSOPHILA HOMEODOMAIN PROTEINS CELL Hayashi, S., Scott, M. P. 1990; 63 (5): 883-894

  View details for Web of Science ID A1990EL75900004

  View details for PubMedID 1979524

• THE PUTATIVE DROSOPHILA GROWTH-FACTOR DPP IS REGULATED BY HOMEOTIC GENES DURING EMBRYOGENESIS Reuter, R., PANGANIBAN, G. E., HOFFMANN, F. M., Scott, M. P. WALTER DE GRUYTER & CO. 1990: 812–12

  View details for Web of Science ID A1990EC97200188

• DIFFERENT PATTERNS OF TRANSCRIPTION FROM THE 2 ANTENNAPEDIA PROMOTERS DURING DROSOPHILA EMBRYOGENESIS DEVELOPMENT Bermingham, J. R., MARTINEZARIAS, A., PETITT, M. G., Scott, M. P. 1990; 109 (3): 553-566

  Abstract

  The homeotic genes of Drosophila control the differentiation of segments during development. Mutations in these genes cause one or more segments to develop structures normally found elsewhere in the organism. Several studies have shown that the spatial patterns of homeotic gene transcription are highly complex, and that these precise patterns of transcription are critical to normal development. The homeotic gene Antennapedia (Antp), a member of the Antennapedia Complex, is required for the correct differentiation of thoracic segments in both embryos and adults. The patterns of total Antp transcript and protein accumulation have been described in detail, but the contribution of each promoter to the overall pattern in embryos has not been reported. We have examined in detail the spatial distribution of transcripts from each of the Antp promoters in both embryo sections and whole embryos by in situ hybridization using promoter-specific probes. We show that the transcripts from each of the two promoters accumulate in distinct, but overlapping patterns during embryogenesis. The results demonstrate that the two Antp promoters are differentially regulated in embryos and provide a basis for examining the regulation of the two promoters and characterizing more fully the function of Antp during embryogenesis. In addition, we have examined the regulation of each of the Antp promoters by genes of the bithorax complex (BX-C). We show that in BX-C- embryos both promoters are derepressed in the abdomen.

  View details for Web of Science ID A1990DQ06500004

  View details for PubMedID 1976090

• EXPRESSION AND FUNCTION OF THE HOMEOTIC GENES ANTENNAPEDIA AND SEX COMBS REDUCED IN THE EMBRYONIC MIDGUT OF DROSOPHILA DEVELOPMENT Reuter, R., Scott, M. P. 1990; 109 (2): 289-?

  Abstract

  Drosophila homoeotic genes control the formation of external morphological features of the embryo and adult, and in addition affect differentiation of the nervous system. Here we describe the morphogenetic events in the midgut that are controlled by the homoeotic genes Sex combs reduced (Scr) and Antennapedia (Antp). The midgut is composed of two cell layers, an inner endoderm and an outer visceral mesoderm that surround the yolk. Scr and Antp are expressed in the visceral mesoderm but not in the endoderm. The two genes are required for different aspects of the midgut morphogenesis. In Scr null mutant embryos the gastric caeca fail to form. Scr is expressed in the visceral mesoderm cells posterior to the primordia of the gastric caeca and appears to be indirectly required for the formation of the caeca. Antp is expressed in visceral mesoderm cells that overlie a part of the midgut where a constriction will form, and Antp null mutant embryos fail to form this constriction. An ultrastructural analysis of the midgut reveals that the visceral mesoderm imposes the constriction on the endoderm and the yolk. The mesodermal tissue contracts within the constriction and thereby penetrates the layer of the midgut endoderm. Microtubules participate in the morphological changes of the visceral mesoderm cells. The analysis of the expression of Scr in Antp mutant embryos revealed a case of tissue-specific regulation of Scr expression by Antp. In the epidermis, Antp has been shown to negatively regulate Scr, but it positively regulates Scr in the visceral mesoderm.

  View details for Web of Science ID A1990DK60900004

  View details for PubMedID 1976087

• A SEQUENCE-SPECIFIC DNA-BINDING PROTEIN THAT ACTIVATES FUSHI-TARAZU SEGMENTATION GENE-EXPRESSION GENES & DEVELOPMENT Ueda, H., Sonoda, S., Brown, J. L., Scott, M. P., Wu, C. 1990; 4 (4): 624-635

  Abstract

  The Drosophila segmentation gene fushi tarazu (ftz) is expressed at the cellular blastoderm stage in a pattern of seven transverse stripes; the stripes lie out of register with the segmental primordia, spanning alternate segmental boundaries. The zebra element, a 740-bp DNA sequence upstream of the ftz translational start, directs striped expression of lacZ when introduced into the fly genome. We have purified to homogeneity a sequence-specific DNA-binding factor, FTZ-F1, that binds to two sites located within the zebra element and to two sites within the ftz protein-coding sequence. FTZ-F1 DNA-binding activity is first detected in extracts of 1.5- to 4-hr embryos, coincident with the time of ftz expression in stripes; the activity then diminishes before reappearing during late embryo, larval, and adult stages. When one of the FTZ-F1-binding sequences in the zebra element is mutated by 2- or 4-base substitutions, the binding to FTZ-F1 is disrupted in vitro, and the intensity of lacZ expression is reduced in transformed embryos, especially in stripes 1, 2, 3, and 6. The results suggest that FTZ-F1 is a transcriptional activator necessary for the proper expression of the ftz gene.

  View details for Web of Science ID A1990DD09700013

  View details for PubMedID 2113881

• THE DROSOPHILA PATCHED GENE ENCODES A PUTATIVE MEMBRANE-PROTEIN REQUIRED FOR SEGMENTAL PATTERNING CELL Hooper, J. E., Scott, M. P. 1989; 59 (4): 751-765

  Abstract

  The patched (ptc) gene is one of several segment polarity genes required for correct patterning within every segment of Drosophila. The absence of ptc gene function causes a transformation of the fate of cells in the middle part of each segment so that they form pattern elements characteristic of cells positioned around the segment border. Analysis of the mutant phenotype demonstrates that both segment and parasegment borders are included in the duplicated pattern of ptc mutants. We have cloned the ptc gene and deduced that the product is a 1286 amino acid protein with at least seven putative transmembrane alpha helices. ptc RNA is expressed in embryos in broad stripes of segmental periodicity that later split into two stripes per segment primordium. The pattern of expression does not directly predict the transformation seen in ptc mutant embryos, suggesting that ptc participates in cell interactions that establish pattern within the segment.

  View details for Web of Science ID A1989CA40200019

  View details for PubMedID 2582494

• THE STRUCTURE AND FUNCTION OF THE HOMEODOMAIN BIOCHIMICA ET BIOPHYSICA ACTA Scott, M. P., Tamkun, J. W., HARTZELL, G. W. 1989; 989 (1): 25-48

  View details for Web of Science ID A1989AH83600003

  View details for PubMedID 2568852

• TRANSCRIPTIONAL ACTIVATION BY THE ANTENNAPEDIA AND FUSHI-TARAZU PROTEINS IN CULTURED DROSOPHILA CELLS CELL Winslow, G. M., Hayashi, S., Krasnow, M., Hogness, D. S., Scott, M. P. 1989; 57 (6): 1017-1030

  Abstract

  Drosophila homeodomain proteins bind to specific DNA sequences in vitro and are hypothesized to regulate the transcription of other genes during development. Using a cotransfection assay, we have shown that homeodomain proteins encoded by the homeotic gene Antennapedia (Antp) and the segmentation gene fushi tarazu, as well as a hybrid homeodomain protein, are activators of transcription from specific promoters in cultured Drosophila cells. Sequences downstream of the Antp P1 and Ultrabithorax transcription start sites mediate the observed activation. A TAA-rich DNA sequence to which the Antp protein binds in vitro is sufficient to confer regulation on a heterologous promoter. The results demonstrate that homeodomain proteins are transcriptional regulators in vivo and that in cultured cells, different homeodomain-containing proteins can act upon a common sequence to modulate gene transcription.

  View details for Web of Science ID A1989AC40300015

  View details for PubMedID 2567631

• CONTROL ELEMENTS OF THE P2 PROMOTER OF THE ANTENNAPEDIA GENE GENES & DEVELOPMENT Boulet, A. M., Scott, M. P. 1988; 2 (12A): 1600-1614

  Abstract

  Antennapedia (Antp), a homeotic gene of Drosophila required for proper differentiation of the thorax of the fly, is expressed in complex spatial patterns during development. The gene is greater than 100 kb long and has two independently regulated promoters. To characterize cis-acting regulatory elements responsible for the expression pattern, fusions of the Antp promoter 2 cap site and upstream sequences to an Adh-lacZ gene were introduced into flies. A 10-kb sequence directs beta-galactosidase production in a pattern that closely resembles the endogenous P2 pattern. Transcription from the 10-kb fusions is regulated by three genes that regulate Antp transcription. Control elements, including a target of action of homeo-domain-containing proteins, were mapped by deleting parts of the 10-kb sequence.

  View details for Web of Science ID A1988R431000008

  View details for PubMedID 2905685

• DEVELOPMENTALLY REGULATED ALTERNATIVE SPLICING OF TRANSCRIPTS FROM THE DROSOPHILA HOMEOTIC GENE ANTENNAPEDIA CAN PRODUCE 4 DIFFERENT PROTEINS EMBO JOURNAL Bermingham, J. R., Scott, M. P. 1988; 7 (10): 3211-3222

  Abstract

  Antennapedia (Antp) is a Drosophila homeotic gene that controls differentiation of the thoracic segments. Antp transcripts are produced from either of two promoters that are independently regulated in temporally and spatially distinct patterns. In addition, Antp transcripts utilize either of two major polyadenylation sites. Antp primary transcripts contain the same protein coding sequences. Alternative RNA splicing at two positions within the primary transcripts produces mRNAs that can encode four slightly different Antp proteins. Different classes of alternatively spliced transcript predominate early and late in Drosophila development, indicating that the Antp gene is regulated by the processing of its transcripts as well as by controlling their transcription. Alternative splicing appears to be independent of which promoter and which polyadenylation site is used.

  View details for Web of Science ID A1988Q285200033

  View details for PubMedID 2903048

• MATERNAL-EFFECT GENES THAT ALTER THE FATE MAP OF THE DROSOPHILA BLASTODERM EMBRYO DEVELOPMENTAL BIOLOGY Winslow, G. M., Carroll, S. B., Scott, M. P. 1988; 129 (1): 72-83

  Abstract

  The pattern of segmentation in the Drosophila embryo is controlled by at least 25 zygotically active genes and at least 20 maternally active genes. We have examined the pattern of expression of the protein product of the zygotically active segmentation gene fushi tarazu (ftz) at the cellular blastoderm stage in progeny of mutant females homozygous for each of six maternal-effect segmentation genes to observe the early effects of the maternal-effect genes on zygotic gene expression. The genes included exuperantia (a member of the anterior class of maternal-effect segmentation genes); staufen and vasa (members of the posterior class); and torso, trunk, and fs(1)N (members of the terminal class). Mutations in the genes caused a disruption of the normal pattern of ftz stripes in regions of the embryo where gene activity is known to be required. The ftz stripes provide a marker for segmental determination at the cellular blastoderm stage, making it possible to correlate aberrant patterns of ftz protein with defects in cuticle morphology at the end of embryogenesis. ftz protein expression in progeny of females mutant for combinations of the above genes was also examined. The changes in the ftz pattern in progeny of females doubly mutant for genes of the anterior and terminal classes or of the posterior and terminal classes can largely be understood as the result of the additive effects of the single mutations. In contrast, clearly nonadditive effects on the ftz pattern were seen when a mutation in a gene of the anterior class (exuperantia) was combined with mutations in posterior class genes.

  View details for Web of Science ID A1988P803800007

  View details for PubMedID 3410162

• SEGMENTATION AND HOMEOTIC GENE-FUNCTION IN THE DEVELOPING NERVOUS-SYSTEM OF DROSOPHILA TRENDS IN NEUROSCIENCES Doe, C. Q., Scott, M. P. 1988; 11 (3): 101-106

  View details for Web of Science ID A1988M242900008

  View details for PubMedID 2465606

• TEMPORAL AND SPATIAL RELATIONSHIPS BETWEEN SEGMENTATION AND HOMEOTIC GENE-EXPRESSION IN DROSOPHILA EMBRYOS - DISTRIBUTIONS OF THE FUSHI-TARAZU, ENGRAILED, SEX COMBS REDUCED, ANTENNAPEDIA, AND ULTRABITHORAX PROTEINS GENES & DEVELOPMENT Carroll, S. B., DiNardo, S., OFARRELL, P. H., White, R. A., Scott, M. P. 1988; 2 (3): 350-360

  Abstract

  The specification of segment number and identity in the Drosophila embryo requires the activity of several classes of genes that may be grouped according to the array of pattern elements that they control. Double-label immunofluorescence was used to simultaneously localize the products of genes representative of the pair-rule segmentation class (fushi tarazu), the segment polarity class (engrailed), and the homeotic class (Sex combs reduced, Antennapedia, and Ultrabithorax) of pattern-regulating genes. The temporal order of appearance of each class of proteins and the precise spatial relationships between the products of the different genes are described with single-cell resolution. Boundaries of gene expression, particularly the parasegmental boundaries, are established by early-acting genes such as fushi tarazu and subsequently respected by the expression patterns of later appearing gene products such as engrailed and Ultrabithorax, suggesting regulatory relationships between certain pairs of genes. In addition, the dynamic transitions observed in spatial relationship among the Sex combs reduced, Antennapedia, and Ultrabithorax homeotic protein patterns during the early period of embryogenesis may reflect cross-regulatory interactions among these genes. Finally, some cells contain a single homeotic product, whereas other cells simultaneously contain several, suggesting that certain cells may be determined by the combinatorial action of homeotic genes.

  View details for Web of Science ID A1988P138300008

  View details for PubMedID 2897943

• THE INTERACTION OF PROTEINS ENCODED BY DROSOPHILA HOMEOTIC AND SEGMENTATION GENES WITH SPECIFIC DNA-SEQUENCES DEVELOPMENT Laughon, A., Howell, W., Scott, M. P. 1988; 104: 75-83

  Abstract

  The ANT-C gene cluster is part of a network of genes that govern pattern formation in the development of Drosophila. The ANT-C genes encode proteins that contain a conserved 60 amino acid sequence, the homeodomain. Here we show that the homeodomains encoded by two of the ANT-C loci confer sequence-specific DNA-binding activity. The DNA sequence specificities of the Dfd and ftz homeodomains appear to overlap completely in vitro, indicating that differences in regulatory specificity among ANT-C and BX-C proteins (assuming that differences exist) must be a consequence of the nonconserved protein sequences found outside of the homeodomains. Deletions that remove sequences from either end of the ftz homeodomain abolish DNA-binding activity, consistent with the commonly held assumption that the homeodomain is a structural domain. The relevance of in vitro DNA-binding experiments to the regulatory function of ftz is supported by our finding that a temperature-sensitive ftz mutation that causes a pairwise fusion of embryonic segments also reduces the affinity of the ftz homeodomain for DNA. Restriction fragments containing ftz homeodomain binding sites were identified within a 90 kb stretch of DNA extending the Antp P1 and P2 promoters. Binding sites appear to be clustered near the P1 promoter but also occur near P2 and in the region between the two. The task remains of determining which of these sequences mediate regulation of Antp by ftz or by other genes that encode closely related homeodomains.

  View details for Web of Science ID A1988R717900009

  View details for PubMedID 2908297

• The molecular biology of pattern formation in the early embryonic development of Drosophila. Developmental biology (New York, N.Y. : 1985) Scott, M. P. 1988; 5: 151-185

  View details for PubMedID 2908379

• THE SEGMENTATION AND HOMEOTIC GENE NETWORK IN EARLY DROSOPHILA DEVELOPMENT CELL Scott, M. P., Carroll, S. B. 1987; 51 (5): 689-698

  View details for Web of Science ID A1987L208200001

  View details for PubMedID 2890437

• THE EXPRESSION AND REGULATION OF SEX COMBS REDUCED PROTEIN IN DROSOPHILA EMBRYOS GENES & DEVELOPMENT Riley, P. D., Carroll, S. B., Scott, M. P. 1987; 1 (7): 716-730

  Abstract

  Homeotic genes are expressed in spatially precise patterns during Drosophila development to control segmental differentiation. The Sex combs reduced (Scr) gene of the Antennapedia gene complex is involved in the determination of the labial and prothoracic segments of the embryo. To study both the wild-type pattern of Scr expression and the regulatory relationships of Scr to other regulatory genes, an antibody probe that detects the Scr protein was prepared. We find that the Scr gene product is expressed in a dynamic pattern over the course of embryogenesis, beginning in the ectoderm in parasegment 2 while the germ band is elongated and extending to parasegment 3 during the completion of germ band shortening. The locations of Scr protein correlate well with the part of the embryo that are altered in Scr- mutants. After head involution occurs, Scr protein is also expressed in the ganglion corresponding to parasegment 2 of the ventral nervous system. The precise spatial expression of Scr is attained through regulation by both homeotic genes and segmentation genes. The lack of proper Antennapedia or Polycomb gene function causes ectopic Scr protein expression. Mutations in the segmentation genes fushi tarazu, hunchback, Krüppel, and giant alter the spatial pattern of Scr expression.

  View details for Web of Science ID A1987K161900009

  View details for PubMedID 2892760

• THE SPATIAL AND TEMPORAL DEPLOYMENT OF DFD AND SCR TRANSCRIPTS THROUGHOUT DEVELOPMENT OF DROSOPHILA DEVELOPMENT MARTINEZARIAS, A., Ingham, P. W., Scott, M. P., Akam, M. E. 1987; 100 (4): 673-683

  Abstract

  In Drosophila, the Deformed (Dfd) and Sex combs reduced (Scr) genes determined the developmental pathways followed by the most anterior metameric units. Using in situ hybridization, we have monitored the spatial distributions of transcripts from these two genes. Dfd RNA accumulates in parasegments 0 and 1; Scr RNA accumulation shows a dynamic pattern spanning parasegments 2 and 3. The expression of Dfd and Scr seems to change from parasegmental to segmental during formation of the gnathal appendages. Both genes are transcribed during imaginal development: Dfd in a portion of the eye-antennal disc; Scr in the labial and prothoracic discs. In addition, we find Scr RNA in the adepithelial cells of all mesothoracic discs.

  View details for Web of Science ID A1987J842400012

  View details for PubMedID 2450726

• GENES THAT CONTROL DORSOVENTRAL POLARITY AFFECT GENE-EXPRESSION ALONG THE ANTEROPOSTERIOR AXIS OF THE DROSOPHILA EMBRYO DEVELOPMENT Carroll, S. B., Winslow, G. M., TWOMBLY, V. J., Scott, M. P. 1987; 99 (3): 327-332

  Abstract

  At least 13 genes control the establishment of dorsoventral polarity in the Drosophila embryo and more than 30 genes control the anteroposterior pattern of body segments. Each group of genes is thought to control pattern formation along one body axis, independently of the other group. We have used the expression of the fushi tarazu (ftz) segmentation gene as a positional marker to investigate the relationship between the dorsoventral and anteroposterior axes. The ftz gene is normally expressed in seven transverse stripes. Changes in the striped pattern in embryos mutant for other genes (or progeny of females homozygous for maternal-effect mutations) can reveal alterations of cell fate resulting from such mutations. We show that in the absence of any of ten maternal-effect dorsoventral polarity gene functions, the characteristic stripes of ftz protein are altered. Normally there is a difference between ftz stripe spacing on the dorsal and ventral sides of the embryo; in dorsalized mutant embryos the ftz stripes appear to be altered so that dorsal-type spacing occurs on all sides of the embryo. These results indicate that cells respond to dorsoventral positional information in establishing early patterns of gene expression along the anteroposterior axis and that there may be more significant interactions between the different axes of positional information than previously determined.

  View details for Web of Science ID A1987G384800004

  View details for PubMedID 3653004

• COMPLEX LOCI OF DROSOPHILA ANNUAL REVIEW OF BIOCHEMISTRY Scott, M. P. 1987; 56: 195-227

  View details for Web of Science ID A1987H963200008

  View details for PubMedID 2887142

• STRUCTURE OF TRANSCRIPTS FROM THE HOMEOTIC ANTENNAPEDIA GENE OF DROSOPHILA-MELANOGASTER - 2 PROMOTERS CONTROL THE MAJOR PROTEIN-CODING REGION MOLECULAR AND CELLULAR BIOLOGY Laughon, A., Boulet, A. M., Bermingham, J. R., Laymon, R. A., Scott, M. P. 1986; 6 (12): 4676-4689

  Abstract

  The Antennapedia (Antp) homeotic gene of Drosophila melanogaster regulates segmental identity in the thorax. Loss of Antp function results in altered development of the embryonic thoracic segments or can cause legs to be transformed into antennae. Certain combinations of Antp recessive lethal alleles complement to permit normal development. The structure of the Antp gene, analyzed by sequencing cDNA clones and exons and by transcript mapping, revealed some of the basis for its genetic complexity. It has two promoters governing two nested transcription units, one unit 36 and one 103 kilobase pairs (kb) long. Both units incorporated the same protein-coding exons, all of which are located in the 3'-most 13 kb of the gene. The two promoters resulted in the attachment of either of two long noncoding leader sequences (1.5 and 1.7 kb) to a 1.1-kb open reading frame. Both transcription units used the same pair of alternative polyadenylation sites 1.4 kb apart; the choice of sites was developmentally regulated. Some of the mutations that disrupt the larger transcription unit complemented a mutation affecting the smaller one. Dominant mutations that transform antennae into legs split the gene but left the coding exons intact. The encoded protein has unusually long runs of glutamine and a homeodomain near the C terminus.

  View details for Web of Science ID A1986E956800062

  View details for PubMedID 2879223

• THE LOCALIZATION AND REGULATION OF ANTENNAPEDIA PROTEIN EXPRESSION IN DROSOPHILA EMBRYOS CELL Carroll, S. B., Laymon, R. A., MCCUTCHEON, M. A., Riley, P. D., Scott, M. P. 1986; 47 (1): 113-122

  Abstract

  The homeotic Antennapedia (Antp) gene of Drosophila is required for the normal differentiation of the thoracic segments during embryonic development and metamorphosis. Antibodies to a recombinant Antp protein were used to localize the protein in whole mount embryos. Antp is expressed in the nuclei of cells of the thoracic embryonic epidermis and several segments of the ventral and peripheral nervous systems. Analysis of Antp expression in mutant embryos revealed three levels of Antp regulation by genes of the bithorax complex, pleiotropic homeotic loci, and Antp itself. The distributions of the Antp and the Ultrabithorax (Ubx) proteins in doubly-labeled embryos suggest that the Ubx protein may be one direct negative regulator of Antp gene expression.

  View details for Web of Science ID A1986E471700014

  View details for PubMedID 3093083

• MATERNAL CONTROL OF DROSOPHILA SEGMENTATION GENE-EXPRESSION NATURE Carroll, S. B., Winslow, G. M., Schupbach, T., Scott, M. P. 1986; 323 (6085): 278-280

  Abstract

  Several genes have been identified that are involved in establishing the segmented body pattern during development of the fruit-fly Drosophila melanogaster. These fall into several classes on the basis of the kind of alteration to the wild-type segmentation pattern observed in mutant embryos. For example, mutations of the pair-rule class, such as fushi tarazu (ftz), cause the deletion of pattern elements with a two-segment periodicity; those of the gap class, such as knirps, cause the deletion of contiguous groups of segments. The availability of antibodies against the ftz protein has allowed its spatial pattern of expression to be studied during the development of wild-type and mutant embryos. The aim of the latter kind of experiment is to investigate possible interactions between these important genes. We have recently reported that knirps mutations cause a striking alteration to the pattern of transverse stripes of ftz expression usually seen during embryogenesis. Knirps is a zygotically-expressed gene, but recently a class of maternally-active genes has been identified that causes similar defects in pattern formation. We have now investigated the pattern of ftz expression in mutants of this class and have found that while they do have features seen in knirps mutants, they also exhibit significant differences between the different mutations reflecting the distinct but overlapping domains of gene activity. These observations demonstrate that maternally-active segmentation genes regulate zygotic gene expression, and that some of their effects on ftz may be directed through the knirps gene.

  View details for Web of Science ID A1986E040800071

  View details for PubMedID 3093889

• MORE ON THE HOMEOBOX BIOESSAYS Scott, M. P. 1986; 5 (2): 88-89

  View details for Web of Science ID A1986D564300009

  View details for PubMedID 2878664

• ZYGOTICALLY ACTIVE GENES THAT AFFECT THE SPATIAL EXPRESSION OF THE FUSHI-TARAZU SEGMENTATION GENE DURING EARLY DROSOPHILA EMBRYOGENESIS CELL Carroll, S. B., Scott, M. P. 1986; 45 (1): 113-126

  Abstract

  The establishment of the segmental body pattern of Drosophila requires the coordinated functions of three classes of zygotically active genes early in development. We have examined the effects of mutations in these genes on the spatial expression of the fushi tarazu (ftz) pair-rule segmentation gene. Mutations in four gap loci and in three pair-rule loci dramatically affect the initial pattern of transverse stripes of ftz-containing nuclei. Five other pair-rule genes and several other loci that affect the larval cuticular pattern do not detectably affect ftz expression. No simple regulatory relationships can be deduced. Rather, expression of the ftz gene depends upon the interactions among the different segmentation genes active at each position along the anterior-posterior axis of the early embryo.

  View details for Web of Science ID A1986A896100012

  View details for PubMedID 3082519

• SPATIAL PROGRAMMING OF GENE-EXPRESSION IN EARLY DROSOPHILA EMBRYOGENESIS ANNUAL REVIEW OF CELL BIOLOGY Scott, M. P., OFARRELL, P. H. 1986; 2: 49-80

  View details for Web of Science ID A1986E986500003

  View details for PubMedID 2881561

• Localization of homoeodomain-containing proteins using antibodies against a synthetic oligopeptide Symp. Soc. Devel. Biol. Carroll, S.B., P.D. Riley, M.W. Klymkowsky, J. van Blerkom, J. Stewart, M.P. Scott 1985; 43: 257-262
• LOCALIZATION OF THE FUSHI-TARAZU PROTEIN DURING DROSOPHILA EMBRYOGENESIS CELL Carroll, S. B., Scott, M. P. 1985; 43 (1): 47-57

  Abstract

  The fushi tarazu (ftz) gene of Drosophila acts early in embryogenesis to regulate body segmentation. The localization of the ftz protein product in embryos was examined using indirect immunofluorescence microscopy. Antibodies were prepared against a beta-galactosidase-ftz hybrid protein made in E. coli. The ftz protein was first detectable in blastoderm-stage embryos as seven stripes of nuclei encircling the embryos transversely. The stripes persist through the early events of gastrulation, but disappear before overt segmentation is visible. The ftz protein is expressed a second time in some nuclei of the developing nervous system. In contrast to the early pattern, at the later stage, ftz is expressed in each of fifteen metameric subunits of the embryo.

  View details for Web of Science ID A1985AVE3900008

  View details for PubMedID 3000605

• Molecules and puzzles from the antennapedia homeotic gene complex of Drosophila Trends in Genetics Scott MP 1985; 1: 74-80
• COMMON PROPERTIES OF PROTEINS ENCODED BY THE ANTENNAPEDIA COMPLEX GENES OF DROSOPHILA-MELANOGASTER COLD SPRING HARBOR SYMPOSIA ON QUANTITATIVE BIOLOGY Laughon, A., Carroll, S. B., STORFER, F. A., Riley, P. D., Scott, M. P. 1985; 50: 253-262

  View details for Web of Science ID A1985C628800031

  View details for PubMedID 3938363

• STRUCTURAL RELATIONSHIPS AMONG GENES THAT CONTROL DEVELOPMENT - SEQUENCE HOMOLOGY BETWEEN THE ANTENNAPEDIA, ULTRABITHORAX, AND FUSHI TARAZU LOCI OF DROSOPHILA PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA-BIOLOGICAL SCIENCES Scott, M. P., Weiner, A. J. 1984; 81 (13): 4115-4119

  Abstract

  Genes that regulate the development of the fruit fly Drosophila melanogaster exist as tightly linked clusters in at least two cases. These clusters, the bithorax complex (BX-C) and the Antennapedia complex (ANT-C), both contain multiple homoeotic loci: mutations in each locus cause a transformation of one part of the fly into another. Several repetitive DNA sequences, including at least one transposon, were mapped in the ANT-C. DNA from the 3' exon of Antennapedia (Antp), a homoeotic locus in the ANT-C, hybridized weakly to DNA from the 3' exon of Ultrabithorax (Ubx), a homoeotic locus in the BX-C. DNA from each of these 3' exons also hybridized weakly to DNA from the fushi tarazu locus of the ANT-C. The fushi tarazu (ftz) locus controls the number and differentiation of segments in the developing embryo. Sequence analysis of the cross-hybridizing DNA from the three loci revealed the conservation of predicted amino acid sequences derived from coding parts of the genes. This suggests that two homoeotic loci and a "segment-deficient" locus encode protein products with partially shared structures and that the three loci may be evolutionarily and functionally related.

  View details for Web of Science ID A1984TA46100039

  View details for PubMedID 6330741

• Homoeotic gene transcripts in the neural tissue of insects Trends in Neurosciences Scott MP 1984; 7 (7): 221-223
• A MOLECULAR ANALYSIS OF FUSHI-TARAZU, A GENE IN DROSOPHILA-MELANOGASTER THAT ENCODES A PRODUCT AFFECTING EMBRYONIC SEGMENT NUMBER AND CELL FATE CELL Weiner, A. J., Scott, M. P., Kaufman, T. C. 1984; 37 (3): 843-851

  Abstract

  Mutations at the fushi tarazu locus in Drosophila melanogaster affect both segment number and the pattern of cuticular structures on alternating segments of embryos. The ftz gene has been cloned and characterized. Two mutations, ftzw20 and ftzRpl are associated with lesions in a 3.2 kb fragment of DNA cloned in the Antennepedia Complex (ANT-C) chromosome "walk." The structure of DNA isolated from the ftzw20 and ftzRpl chromosomes indicates that the mutations are associated with a 4.9 kb insertion of DNA and a chromosomal rearrangement breakpoint, respectively. The 3.2 kb genomic DNA fragment hybridizes to a 1.8 kb polyadenylated transcript which accumulates maximally at 2-4 hr of embryonic development. The ftzw20 and ftzRpl mutations have different phenotypic consequences for the developing embryo, although both mutations interrupt the 1.8 kb transcription unit. The genetic and molecular data indicate that the 1.8 kb transcript derives from the ftz locus. The gene products are synthesized and utilized several hours prior to the visibly detectable morphogenetic events which the gene apparently regulates.

  View details for Web of Science ID A1984TB80600016

  View details for PubMedID 6430569

• SEQUENCE OF A DROSOPHILA SEGMENTATION GENE - PROTEIN-STRUCTURE HOMOLOGY WITH DNA-BINDING PROTEINS NATURE Laughon, A., Scott, M. P. 1984; 310 (5972): 25-31

  Abstract

  Mutations in the fushi tarazu (ftz) locus of Drosophila result in embryos with half the usual number of body segments. The sequences of the wild-type gene, a temperature-sensitive allele and a dominant mutant allele are presented. A portion of the conserved protein domain present in ftz and several homoeotic genes resembles the DNA-binding region of prokaryotic DNA-binding proteins, and is also similar to products of the yeast mating-type locus.

  View details for Web of Science ID A1984SY85100036

  View details for PubMedID 6330566

• THE MOLECULAR-ORGANIZATION OF THE ANTENNAPEDIA LOCUS OF DROSOPHILA CELL Scott, M. P., Weiner, A. J., Hazelrigg, T. I., Polisky, B. A., Pirrotta, V., SCALENGHE, F., Kaufman, T. C. 1983; 35 (3): 763-776

  Abstract

  The Antennapedia complex (ANT-C) of Drosophila is a cluster of genes involved in the regulation of morphogenesis, including at least three homoeotic loci (Antp, Scr, and pb), in which mutations cause switches of cell fates from one developmental pathway to another. We have isolated DNA clones containing most of the ANT-C, a region of about 300 kb. Antp mutations are distributed across more than 100 kb of the ANT-C. Dominant Antp mutations are associated with certain chromosome rearrangements and insertions that interrupt the 100 kb region; other chromosome breaks within the region cause recessive lethality and have no dominant effects. Two prominent transcripts (3.5 and 5.0 kb) are derived from exons within and at the two ends of the 100 kb region.

  View details for Web of Science ID A1983RX39200021

  View details for PubMedID 6418389

• TRANSLATIONAL CONTROL IN LYSATES OF DROSOPHILA-MELANOGASTER CELLS PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA-BIOLOGICAL SCIENCES Scott, M. P., Pardue, M. L. 1981; 78 (6): 3353-3357

  Abstract

  Cell-free protein-synthesizing systems made from Drosophila melanogaster cells were used to study the translational control induced in these cells by heat shock. Lysates of normally growing cells, termed 25 degrees C cells, translate both normal and heat shock mRNAs. Lysates of cells heat shocked at 36 degrees C for 1 hr, termed 36 degrees C cells, translate preferentially heat-shock mRNAs and a few 25 degrees C cell mRNAs. Thus, both lysates appear to reproduce the control displayed in vivo. Both lysates are optimally active at 28 degrees C, and all translations are done at that temperature, demonstrating that, once established, the discrimination system does not require heat-shock temperature for its activity. Addition of crude ribosome fractions from 25 degrees C cell lysates to lysates from heat-shocked cells "rescues" translation of 25 degrees C cell mRNA, which suggests that the discriminating elements are associated with ribosomes. Neither the heat-shock crude ribosome supplements nor the soluble fractions have any effect on either lysate. The experiments also show that RNA selection is determined by some feature of the RNA structure that is insensitive to protease digestion and phenol/chloroform extraction. The essential structural feature may not be unique to Drosophila mRNAs because the Drosophila lysate is capable of discriminating among mRNAs from other organisms.

  View details for Web of Science ID A1981LW77700009

  View details for PubMedID 6267586

• Expression of the heat shock genes in Drosophila melanogaster. Developmental Biology Using Purified Genes (D.D. Brown and D.F. Fox, eds.), ICN-UCLA Symp. Pardue, M.L., D.G. Ballinger, M.P. Scott 1981; 23: 415-427
• TRANSLATIONAL CONTROL OF PROTEIN-SYNTHESIS IN RESPONSE TO HEAT-SHOCK IN DROSOPHILA-MELANOGASTER CELLS CELL Storti, R. V., Scott, M. P., Rich, A., Pardue, M. L. 1980; 22 (3): 825-834

  Abstract

  In response to elevated temperature, Drosophila cells synthesize a small set of proteins known as the heat-shock proteins, while synthesis of most other proteins ceases. In vitro translation has been used to demonstrate that the messenger RNAs encoding the normal (25 degrees) spectrum of proteins are not broken down or irreversibly inactivated in response to the temperature change. During the heat shock only the heat-shock mRNAs plus a small number of preexisting mRNAs are translated, while most other messages are stored and can be reactivated upon return of the cells to their normal temperature. After recovery from heat shock, cells translate both the normal mRNA and the remaining heat-shock mRNA. The translational control operating in intact cells has been reproduced in cell-free translation systems directed by purified mRNA from normal and heat-shocked cells. Lysates prepared from heat-shocked Drosophila cells preferentially translated the heat-shock messages, while the lysate made from normally growing Drosophila cells indiscriminately translated both normal and heat-shock messages. Therefore there must be stable alterations in the translational components of heat-shocked cells which are capable of causing selective translation of the heat-shock messages. In addition there must be information encoded in the heat-shock messages that allows their selection.

  View details for Web of Science ID A1980KW22300024

  View details for PubMedID 6780199

• 2 CLUSTERS OF GENES FOR MAJOR CHORION PROTEINS OF DROSOPHILA-MELANOGASTER CELL Spradling, A. C., Digan, M. E., Mahowald, A. P., Scott, M., Craig, E. A. 1980; 19 (4): 905-914

  View details for Web of Science ID A1980JQ05600011

  View details for PubMedID 6769591

• A NEW TYPE OF VIRUS FROM CULTURED DROSOPHILA CELLS - CHARACTERIZATION AND USE IN STUDIES OF THE HEAT-SHOCK RESPONSE CELL Scott, M. P., Fostel, J. M., Pardue, M. L. 1980; 22 (3): 929-941

  Abstract

  An infectious virus isolated from the cultured Drosophila melanogster cell line, Schneider 2-L, appears to be a member of a new group of animal viruses. The virus, HPS-1 has a genome composed of a single segment of double-stranded RNA approximately 6 kb in length. Virions are particles approximately 36 nm in diameter. They contain only two proteins and no lipid coat. The major protein, presumably the viral coat protein, is 120,000 daltons. A 200,000 dalton protein is present in much lower quantities. Two other proteins, synthesized in virus-producing cells, are encoded in the viral genome but not included in the mature virion. Synthesis of viral proteins is not affected by the heat-shock-induced translational control that inhibits translation of most normal mRNAs but allows protein synthesis on heat-shock mRNAs. The viral mRNAs thus appear to share the structural features of heat-shock mRNAs which permit heir translation in heat-shocked cells. Viral RNA serves as a probe to study heat-shock translational control. Cells heat-shocked in the presence of actinomycin D cannot transcribe heat-shock mRNA and therefore cannot make heat-shock proteins. Although these cells are making neither heat-shock mRNA nor heat-shock protein, the translational control appears fully induced. The normal cell proteins are not made in these cells but viral proteins are synthesized. These results indicate that the heat shock-induced proteins are not components of the translational control mechanism.

  View details for Web of Science ID A1980KW22300035

  View details for PubMedID 6780201

• The heat shock response: a model system for the study of gene regulation in Drosophila. Basic life sciences Pardue, M. L., Scott, M. P., Storti, R. V., Lengyel, J. A. 1980; 16: 41-55

  View details for PubMedID 6161601

• CELL-FREE PROTEIN-SYNTHESIS IN LYSATES OF DROSOPHILA-MELANOGASTER CELLS BIOCHEMISTRY Scott, M. P., Storti, R. V., Pardue, M. L., Rich, A. 1979; 18 (8): 1588-1594

  Abstract

  A procedure is described for preparing cell-free protein synthesizing lysates from Drosophila melanogaster tissue culture cells and embryos. Preparation of translationally active lysates from tissue culture cells is dependent on the presence of rat liver supernatant during cell lysis to inhibit ribonuclease activity. After micrococcal nuclease treatment of the lysate, protein synthesis is dependent on the addition of exogenous messenger RNA. The fidelity of translation is very high. The conditions for optimal translation have been determined. In addition, the effects on translation of a variety of supplements, including rat liver supernatant, have been analyzed. The products of translation by the Drosophila lysate have been compared with those of wheat germ extracts and of micrococcal nuclease treated rabbit reticulocyte lysates. Translation in vitro of bovine parathyroid hormone messenger RNA yielded two products tentatively identified as preproparathyroid hormone and proparathyroid hormone, as well as an unidentified third product. This result suggests that insect enzymes can accurately process mammalian precursor proteins.

  View details for Web of Science ID A1979GR87100032

  View details for PubMedID 106888

• The structure and function of mitochondrial DNA of Drosophila melanogaster. Alfred Benzon Symp. Berninger, M., T. Cech, J. Fostel, K. Potter, M.P. Scott, M.L. Pardue 1979; 13: 229-241
• MYOGENESIS IN PRIMARY-CELL CULTURES FROM DROSOPHILA-MELANOGASTER - PROTEIN-SYNTHESIS AND ACTIN HETEROGENEITY DURING DEVELOPMENT CELL Storti, R. V., HOROVITCH, S. J., Scott, M. P., Rich, A., Pardue, M. L. 1978; 13 (4): 589-598

  Abstract

  Muscle cell cultures from Drosophila melanogaster were obtained by plating dissociated gastrula stage embryo cells on protamine-treated culture dishes. They myogenic cells in these cultures fuse to form multinucleated pulsating cells by 15 hr after plating. An analysis of protein synthesis during myogenesis in these cultures, as measured by the incorporation of 35S-methionine and analyzed by two-dimensional polyacrylamide gel electrophoresis, showed profound changes in the pattern of protein synthesis. This analysis enabled us to identify three distinct classes of proteins. Class A proteins, the most abundant, are synthesized continuously throughout myogenesis, class B proteins are those proteins whose synthesis is initiated during myogenesis and continued throughout development; class C proteins are those synthesized at specific times during development. In addition, three forms of actin have been identified in these cultures. Actin I, which shows increased synthesis concomitant with the myogenic development in these cultures, is apparently a muscle-specific form of actin. Actin II, the predominant "cytoplasmic" form of actin in the nonmuscle Schneider cell line 2, is also the major form in the gastrula cultures before differentiation begins. Synthesis of this actin continues in the myogenic cultures. Actin III is a rapidly turning over form of actin which does not accumulate in either the Schneider cells or the myogenic cultures.

  View details for Web of Science ID A1978EW03300001

  View details for PubMedID 418880