Administrative Appointments


  • Senior Associate Dean for Graduate Education and Postdoctoral Affairs, Stanford University School of Medicine (2015 - 2020)
  • Chair, Department of Developmental Biology, Stanford University (2012 - 2015)

Honors & Awards


  • Pew Scholars Award in the Biomedical Sciences, Pew Charitable Trusts (1998-2002)
  • Rita Allen Foundation Scholars Award, Rita Allen Foundation (2002-2004)
  • Fellow, American Association for the Advancement of Science (2014)
  • Catherine R. Kennedy and Daniel L. Grossman Fellow in Human Biology, Stanford University (2014-Present)
  • Award for Excellence in Faculty Advising in Human Biology, Stanford University (2017)
  • Kenneth M. Cuthbertson Award for Contributions to Stanford University, Stanford University (2020)

Professional Education


  • Ph.D., Stanford University, Biochemistry (1993)
  • B.S., University of Florida, Microbiology (1987)

Current Research and Scholarly Interests


Our research focuses on the development and function of glial cells in the vertebrate nervous system. Glia are non-neuronal cells with many essential functions, ranging from forming the myelin sheath to defending the brain against infection.

One of our goals is to use genetic approaches in zebrafish to discover new genes with essential functions in the glial cells that form the myelin sheath, which allows for rapid axonal conduction in vertebrates. Disruption of myelin underlies important human diseases, including Multiple Sclerosis and peripheral neuropathies. The formation of myelin, which involves reciprocal signaling between neurons and glial cells, a dramatic morphological transformation of the glial cells, and organization of the axon into different specialized domains, is fascinating but nonetheless poorly understood.

In genetic screens, we have identified mutations in more than 15 different genes that have specific functions in the development of myelinated axons. Among these are a novel G-protein coupled receptor that instructs Schwann cells to make myelin in peripheral nerves, receptors that control migration of glial cells along growing axons, a kinesin motor protein that is essential for mRNA localization and normal membrane compaction in myelinating oligodendrocytes, and a transcription factor that regulates the migration of the cells that form myelin in the brain and spinal cord.

Another goal of our research is to identify new genes that regulate microglia, which are specialized macrophages that are dedicated to the immune defense of the brain. Microglia also have critical roles in regulating synaptic connectivity and engulfing dead neurons to maintain homeostasis in the brain. Microglial dysfunction has been implicated a wide array of disorders, including autism and Alzheimer disease.

Starting with screens for mutants with abnormal microglia, we have identified novel genes regulate microglial development and function. Examples include a NOD-like receptor that suppresses inappropriate inflammation, a phosphate exporter that functions specifically in microglia and other tissue macrophages, and a regulator of lysosomal action that allows microglia to digest material that they engulf.

These projects provide new insights into glial cell development and function, generate new animal models of human disease, define pathways that may be disrupted in disease, and may provide new avenues toward therapies for diseases of glia.

2023-24 Courses


Stanford Advisees


Graduate and Fellowship Programs


All Publications


  • Unmyelinated sensory neurons use Neuregulin signals to promote myelination of interneurons in the CNS. Cell reports Lysko, D. E., Talbot, W. S. 2022; 41 (7): 111669

    Abstract

    The signaling mechanisms neurons use to modulate myelination of circuits in the central nervous system (CNS) are only partly understood. Through analysis of isoform-specific neuregulin1 (nrg1) mutants in zebrafish, we demonstrate that nrg1 type II is an important regulator of myelination of two classes of spinal cord interneurons. Surprisingly, nrg1 type II expression is prominent in unmyelinated Rohon-Beard sensory neurons, whereas myelination of neighboring interneurons is reduced in nrg1 type II mutants. Cell-type-specific loss-of-function studies indicate that nrg1 type II is required in Rohon-Beard neurons to signal to other neurons, not oligodendrocytes, to modulate spinal cord myelination. Together, our data support a model in which unmyelinated neurons express Nrg1 type II proteins to regulate myelination of neighboring neurons, a mode of action that may coordinate the functions of unmyelinated and myelinated neurons in the CNS.

    View details for DOI 10.1016/j.celrep.2022.111669

    View details for PubMedID 36384112

  • A lysosomal regulatory circuit essential for the development and function of microglia. Science advances Iyer, H., Shen, K., Meireles, A. M., Talbot, W. S. 2022; 8 (35): eabp8321

    Abstract

    As the primary phagocytic cells of the central nervous system, microglia exquisitely regulate their lysosomal activity to facilitate brain development and homeostasis. However, mechanisms that coordinate lysosomal activity with microglia development, chemotaxis, and function remain unclear. Here, we show that embryonic macrophages require the lysosomal guanosine triphosphatase (GTPase) RagA and the GTPase-activating protein Folliculin to colonize the brain in zebrafish. We demonstrate that embryonic macrophages in rraga mutants show increased expression of lysosomal genes but display significant down-regulation of immune- and chemotaxis-related genes. Furthermore, we find that RagA and Folliculin repress the key lysosomal transcription factor Tfeb and its homologs Tfe3a and Tfe3b in the macrophage lineage. Using RNA sequencing, we establish that Tfeb and Tfe3 are required for activation of lysosomal target genes under conditions of stress but not for basal expression of lysosomal pathways. Collectively, our data define a lysosomal regulatory circuit essential for macrophage development and function in vivo.

    View details for DOI 10.1126/sciadv.abp8321

    View details for PubMedID 36044568

  • The Lysosomal Transcription Factor TFEB Represses Myelination Downstream of the Rag-Ragulator Complex. Developmental cell Meireles, A. M., Shen, K., Zoupi, L., Iyer, H., Bouchard, E. L., Williams, A., Talbot, W. S. 2018; 47 (3): 319

    Abstract

    Myelin allows for fast and efficient axonal conduction, but much remains to be determined about the mechanisms that regulate myelin formation. To investigate the genetic basis of myelination, we carried out a genetic screen using zebrafish. Here, we show that the lysosomal G protein RagA is essential for CNS myelination. In rraga-/- mutant oligodendrocytes, target genes of the lysosomal transcription factor Tfeb are upregulated, consistent with previous evidence that RagA represses Tfeb activity. Loss of Tfeb function is sufficient to restore myelination in RagA mutants, indicating that hyperactive Tfeb represses myelination. Conversely, tfeb-/- single mutants exhibit ectopic myelin, further indicating that Tfeb represses myelination during development. In a mouse model of de- and remyelination, TFEB expression is increased in oligodendrocytes, but the protein is localized to the cytoplasm, and hence inactive, especially during remyelination. These results define essential regulators of myelination and may advance approaches to therapeutic remyelination.

    View details for PubMedID 30399334

  • Oligodendrocyte development and myelin sheath formation are regulated by the antagonistic interaction between the Rag-Ragulator complex and TFEB. Glia Bouchard, E. L., Meireles, A. M., Talbot, W. S. 2023

    Abstract

    Myelination by oligodendrocytes is critical for fast axonal conduction and for the support and survival of neurons in the central nervous system. Recent studies have emphasized that myelination is plastic and that new myelin is formed throughout life. Nonetheless, the mechanisms that regulate the number, length, and location of myelin sheaths formed by individual oligodendrocytes are incompletely understood. Previous work showed that the lysosomal transcription factor TFEB represses myelination by oligodendrocytes and that the RagA GTPase inhibits TFEB, but the step or steps of myelination in which TFEB plays a role have remained unclear. Here, we show that TFEB regulates oligodendrocyte differentiation and also controls the length of myelin sheaths formed by individual oligodendrocytes. In the dorsal spinal cord of tfeb mutants, individual oligodendrocytes produce myelin sheaths that are longer than those produced by wildtype cells. Transmission electron microscopy shows that there are more myelinated axons in the dorsal spinal cord of tfeb mutants than in wildtype animals, but no significant change in axon diameter. In contrast to tfeb mutants, oligodendrocytes in rraga mutants produce shorter myelin sheaths. The sheath length in rraga; tfeb double mutants is not significantly different from wildtype, consistent with the antagonistic interaction between RagA and TFEB. Finally, we find that the GTPase activating protein Flcn and the RagCa and RagCb GTPases are also necessary for myelination by oligodendrocytes. These findings demonstrate that TFEB coordinates myelin sheath length and number during myelin formation in the central nervous system.

    View details for DOI 10.1002/glia.24473

    View details for PubMedID 37767930

  • Promoting validation and cross-phylogenetic integration in model organism research. Disease models & mechanisms Cheng, K. C., Burdine, R. D., Dickinson, M. E., Ekker, S. C., Lin, A. Y., Lloyd, K. C., Lutz, C. M., MacRae, C. A., Morrison, J. H., O'Connor, D. H., Postlethwait, J. H., Rogers, C. D., Sanchez, S., Simpson, J. H., Talbot, W. S., Wallace, D. C., Weimer, J. M., Bellen, H. J. 2022; 15 (9)

    Abstract

    Model organism (MO) research provides a basic understanding of biology and disease due to the evolutionary conservation of the molecular and cellular language of life. MOs have been used to identify and understand the function of orthologous genes, proteins, cells and tissues involved in biological processes, to develop and evaluate techniques and methods, and to perform whole-organism-based chemical screens to test drug efficacy and toxicity. However, a growing richness of datasets and the rising power of computation raise an important question: How do we maximize the value of MOs? In-depth discussions in over 50 virtual presentations organized by the National Institutes of Health across more than 10 weeks yielded important suggestions for improving the rigor, validation, reproducibility and translatability of MO research. The effort clarified challenges and opportunities for developing and integrating tools and resources. Maintenance of critical existing infrastructure and the implementation of suggested improvements will play important roles in maintaining productivity and facilitating the validation of animal models of human biology and disease.

    View details for DOI 10.1242/dmm.049600

    View details for PubMedID 36125045

  • A cAMP Sensor Based on Ligand-Dependent Protein Stabilization. ACS chemical biology Sidoli, M., Chen, L., Lu, A. J., Wandless, T. J., Talbot, W. S. 2022

    Abstract

    cAMP is a ubiquitous second messenger with many functions in diverse organisms. Current cAMP sensors, including Foster resonance energy transfer (FRET)-based and single-wavelength-based sensors, allow for real time visualization of this small molecule in cultured cells and in some cases in vivo. Nonetheless the observation of cAMP in living animals is still difficult, typically requiring specialized microscopes and ex vivo tissue processing. Here we used ligand-dependent protein stabilization to create a new cAMP sensor. This sensor allows specific and sensitive detection of cAMP in living zebrafish embryos, which may enable new understanding of the functions of cAMP in living vertebrates.

    View details for DOI 10.1021/acschembio.2c00333

    View details for PubMedID 35839076

  • Partial loss-of-function variant in neuregulin 1 identified in family with heritable peripheral neuropathy. Human mutation Lysko, D. E., Meireles, A. M., Folland, C., McNamara, E., Laing, N. G., Lamont, P. J., Ravenscroft, G., Talbot, W. S. 2022

    Abstract

    Neuregulin 1 signals are essential for the development and function of Schwann cells, which form the myelin sheath on peripheral axons. Disruption of myelin in the peripheral nervous system (PNS) can lead to peripheral neuropathy, which is characterized by reduced axonal conduction velocity and sensorimotor deficits. Charcot-Marie-Tooth disease is a group of heritable peripheral neuropathies that may be caused by variants in nearly 100 genes. Despite the evidence that Neuregulin 1 is essential for many aspects of Schwann cell development, previous studies have not reported variants in the neuregulin 1 gene (NRG1) in patients with peripheral neuropathy. We have identified a rare missense variant in NRG1 that is homozygous in a patient with sensory and motor deficits consistent with mixed axonal and de-myelinating peripheral neuropathy. Our in vivo functional studies in zebrafish indicate that the patient variant partially reduces NRG1 function. This study tentatively suggests that variants at the NRG1 locus may cause peripheral neuropathy and that NRG1 should be investigated in families with peripheral neuropathy of unknown cause. This article is protected by copyright. All rights reserved.

    View details for DOI 10.1002/humu.24393

    View details for PubMedID 35485770

  • Basic science under threat: Lessons from the Skirball Institute. Cell Sfeir, A., Fishell, G., Schier, A. F., Dustin, M. L., Gan, W. B., Joyner, A., Lehmann, R., Ron, D., Roth, D., Talbot, W. S., Yelon, D., Zychlinsky, A. 2022; 185 (5): 755-758

    Abstract

    Support for basic science has been eclipsed by initiatives aimed at specific medical problems. The latest example is the dismantling of the Skirball Institute at NYU School of Medicine. Here, we reflect on the achievements and mission underlying the Skirball to gain insight into the dividends of maintaining a basic science vision within the academic enterprises.

    View details for DOI 10.1016/j.cell.2022.02.008

    View details for PubMedID 35245477

  • Characterization of mouse Bmp5 regulatory injury element in zebrafish wound models. Bone Heller, I. S., Guenther, C. A., Meireles, A. M., Talbot, W. S., Kingsley, D. M. 2021: 116263

    Abstract

    Many key signaling molecules used to build tissues during embryonic development are re-activated at injury sites to stimulate tissue regeneration and repair. Bone morphogenetic proteins provide a classic example, but the mechanisms that lead to reactivation of BMPs following injury are still unknown. Previous studies have mapped a large "injury response element" (IRE) in the mouse Bmp5 gene that drives gene expression following bone fractures and other types of injury. Here we show that the large mouse IRE region is also activated in both zebrafish tail resection and mechanosensory hair cell injury models. Using the ability to test multiple constructs and image temporal and spatial dynamics following injury responses, we have narrowed the original size of the mouse IRE region by over 100 fold and identified a small 142 bp minimal enhancer that is rapidly induced in both mesenchymal and epithelial tissues after injury. These studies identify a small sequence that responds to evolutionarily conserved local signals in wounded tissues and suggest candidate pathways that contribute to BMP reactivation after injury.

    View details for DOI 10.1016/j.bone.2021.116263

    View details for PubMedID 34826632

  • Myelination induces axonal hotspots of synaptic vesicle fusion that promote sheath growth. Current biology : CB Almeida, R. G., Williamson, J. M., Madden, M. E., Early, J. J., Voas, M. G., Talbot, W. S., Bianco, I. H., Lyons, D. A. 2021

    Abstract

    Myelination of axons by oligodendrocytes enables fast saltatory conduction. Oligodendrocytes are responsive to neuronal activity, which has been shown to induce changes to myelin sheaths, potentially to optimize conduction and neural circuit function. However, the cellular bases of activity-regulated myelination in vivo are unclear, partly due to the difficulty of analyzing individual myelinated axons over time. Activity-regulated myelination occurs in specific neuronal subtypes and can be mediated by synaptic vesicle fusion, but several questions remain: it is unclear whether vesicular fusion occurs stochastically along axons or in discrete hotspots during myelination and whether vesicular fusion regulates myelin targeting, formation, and/or growth. It is also unclear why some neurons, but not others, exhibit activity-regulated myelination. Here, we imaged synaptic vesicle fusion in individual neurons in living zebrafish and documented robust vesicular fusion along axons during myelination. Surprisingly, we found that axonal vesicular fusion increased upon and required myelination. We found that axonal vesicular fusion was enriched in hotspots, namely the heminodal non-myelinated domains into which sheaths grew. Blocking vesicular fusion reduced the stable formation and growth of myelin sheaths, and chemogenetically stimulating neuronal activity promoted sheath growth. Finally, we observed high levels of axonal vesicular fusion only in neuronal subtypes that exhibit activity-regulated myelination. Our results identify a novel "feedforward" mechanism whereby the process of myelination promotes the neuronal activity-regulated signal, vesicular fusion that, in turn, consolidates sheath growth along specific axons selected for myelination.

    View details for DOI 10.1016/j.cub.2021.06.036

    View details for PubMedID 34270947

  • The lysosomal GPCR-like protein GPR137B regulates Rag and mTORC1 localization and activity NATURE CELL BIOLOGY Gan, L., Seki, A., Shen, K., Iyer, H., Han, K., Hayer, A., Wollman, R., Ge, X., Lin, J. R., Dey, G., Talbot, W. S., Meyer, T. 2019; 21 (5): 614-+
  • The lysosomal GPCR-like protein GPR137B regulates Rag and mTORC1 localization and activity. Nature cell biology Gan, L. n., Seki, A. n., Shen, K. n., Iyer, H. n., Han, K. n., Hayer, A. n., Wollman, R. n., Ge, X. n., Lin, J. R., Dey, G. n., Talbot, W. S., Meyer, T. n. 2019

    Abstract

    Cell growth is controlled by a lysosomal signalling complex containing Rag small GTPases and mammalian target of rapamycin complex 1 (mTORC1) kinase. Here, we carried out a microscopy-based genome-wide human short interfering RNA screen and discovered a lysosome-localized G protein-coupled receptor (GPCR)-like protein, GPR137B, that interacts with Rag GTPases, increases Rag localization and activity, and thereby regulates mTORC1 translocation and activity. High GPR137B expression can recruit and activate mTORC1 in the absence of amino acids. Furthermore, GPR137B also regulates the dissociation of activated Rag from lysosomes, suggesting that GPR137B controls a cycle of Rag activation and dissociation from lysosomes. GPR137B-knockout cells exhibited defective autophagy and an expanded lysosome compartment, similar to Rag-knockout cells. Like zebrafish RagA mutants, GPR137B-mutant zebrafish had upregulated TFEB target gene expression and an expanded lysosome compartment in microglia. Thus, GPR137B is a GPCR-like lysosomal regulatory protein that controls dynamic Rag and mTORC1 localization and activity as well as lysosome morphology.

    View details for PubMedID 31036939

  • The Lysosomal Transcription Factor TFEB Represses Myelination Downstream of the Rag-Ragulator Complex DEVELOPMENTAL CELL Meireles, A. M., Shen, K., Zoupi, L., Lyer, H., Bouchard, E. L., Williams, A., Talbot, W. S. 2018; 47 (3): 319-+
  • Non-nuclear Pool of Splicing Factor SFPQ Regulates Axonal Transcripts Required for Normal Motor Development NEURON Thomas-Jinu, S., Gordon, P. M., Fielding, T., Taylor, R., Smith, B. N., Snowden, V., Blanc, E., Vance, C., Topp, S., Wong, C., Bielen, H., Williams, K. L., McCann, E. P., Nicholson, G. A., Pan-Vazquez, A., Fox, A. H., Bond, C. S., Talbot, W. S., Blair, I. P., Shaw, C. E., Houart, C. 2017; 94 (2): 322-?

    Abstract

    Recent progress revealed the complexity of RNA processing and its association to human disorders. Here, we unveil a new facet of this complexity. Complete loss of function of the ubiquitous splicing factor SFPQ affects zebrafish motoneuron differentiation cell autonomously. In addition to its nuclear localization, the protein unexpectedly localizes to motor axons. The cytosolic version of SFPQ abolishes motor axonal defects, rescuing key transcripts, and restores motility in the paralyzed sfpq null mutants, indicating a non-nuclear processing role in motor axons. Novel variants affecting the conserved coiled-coil domain, so far exclusively found in fALS exomes, specifically affect the ability of SFPQ to localize in axons. They broadly rescue morphology and motility in the zebrafish mutant, but alter motor axon morphology, demonstrating functional requirement for axonal SFPQ. Altogether, we uncover the axonal function of the splicing factor SFPQ in motor development and highlight the importance of the coiled-coil domain in this process. VIDEO ABSTRACT.

    View details for DOI 10.1016/j.neuron.2017.03.026

    View details for Web of Science ID 000399451400016

    View details for PubMedID 28392072

  • Interactions between mural cells and endothelial cells stabilize the developing zebrafish dorsal aorta DEVELOPMENT Stratman, A. N., Pezoa, S. A., Farrelly, O. M., Castranova, D., Dye, L. E., Butler, M. G., Sidik, H., Talbot, W. S., Weinstein, B. M. 2017; 144 (1): 115-127

    Abstract

    Mural cells (vascular smooth muscle cells and pericytes) play an essential role in the development of the vasculature, promoting vascular quiescence and long-term vessel stabilization through their interactions with endothelial cells. However, the mechanistic details of how mural cells stabilize vessels are not fully understood. We have examined the emergence and functional role of mural cells investing the dorsal aorta during early development using the zebrafish. Consistent with previous literature, our data suggest that cells ensheathing the dorsal aorta emerge from a sub-population of cells in the adjacent sclerotome. Inhibition of mural cell recruitment to the dorsal aorta through disruption of pdgfr signaling leads to a reduced vascular basement membrane, which in turn results in enhanced dorsal aorta vessel elasticity and failure to restrict aortic diameter. Our results provide direct in vivo evidence for a functional role for mural cells in patterning and stabilization of the early vasculature through production and maintenance of the vascular basement membrane to prevent abnormal aortic expansion and elasticity.

    View details for DOI 10.1242/dev.143131

    View details for Web of Science ID 000393454900015

    View details for PubMedID 27913637

    View details for PubMedCentralID PMC5278630

  • Mapping the Pairwise Choices Leading from Pluripotency to Human Bone, Heart, and Other Mesoderm Cell Types CELL Loh, K. M., Chen, A., Koh, P. W., Deng, T. Z., Sinha, R., Tsai, J. M., Barkal, A. A., Shen, K. Y., Jain, R., Morganti, R. M., Shyh-Chang, N., Fernhoff, N. B., George, B. M., Wernig, G., Salomon, R. E., Chen, Z., Vogel, H., Epstein, J. A., Kundaje, A., Talbot, W. S., Beachy, P. A., Ang, L. T., Weissman, I. L. 2016; 166 (2): 451-467

    Abstract

    Stem-cell differentiation to desired lineages requires navigating alternating developmental paths that often lead to unwanted cell types. Hence, comprehensive developmental roadmaps are crucial to channel stem-cell differentiation toward desired fates. To this end, here, we map bifurcating lineage choices leading from pluripotency to 12 human mesodermal lineages, including bone, muscle, and heart. We defined the extrinsic signals controlling each binary lineage decision, enabling us to logically block differentiation toward unwanted fates and rapidly steer pluripotent stem cells toward 80%-99% pure human mesodermal lineages at most branchpoints. This strategy enabled the generation of human bone and heart progenitors that could engraft in respective in vivo models. Mapping stepwise chromatin and single-cell gene expression changes in mesoderm development uncovered somite segmentation, a previously unobservable human embryonic event transiently marked by HOPX expression. Collectively, this roadmap enables navigation of mesodermal development to produce transplantable human tissue progenitors and uncover developmental processes. VIDEO ABSTRACT.

    View details for DOI 10.1016/j.cell.2016.06.011

    View details for PubMedID 27419872

  • Individual Neuronal Subtypes Exhibit Diversity in CNS Myelination Mediated by Synaptic Vesicle Release CURRENT BIOLOGY Koudelka, S., Voas, M. G., Almeida, R. G., Baraban, M., Soetaert, J., Meyer, M. P., Talbot, W. S., Lyons, D. A. 2016; 26 (11): 1447-1455

    Abstract

    Regulation of myelination by oligodendrocytes in the CNS has important consequences for higher-order nervous system function (e.g., [1-4]), and there is growing consensus that neuronal activity regulates CNS myelination (e.g., [5-9]) through local axon-oligodendrocyte synaptic-vesicle-release-mediated signaling [10-12]. Recent analyses have indicated that myelination along axons of distinct neuronal subtypes can differ [13, 14], but it is not known whether regulation of myelination by activity is common to all neuronal subtypes or only some. This limits insight into how specific neurons regulate their own conduction. Here, we use a novel fluorescent fusion protein reporter to study myelination along the axons of distinct neuronal subtypes over time in zebrafish. We find that the axons of reticulospinal and commissural primary ascending (CoPA) neurons are among the first myelinated in the zebrafish CNS. To investigate how activity regulates myelination by different neuronal subtypes, we express tetanus toxin (TeNT) in individual reticulospinal or CoPA neurons to prevent synaptic vesicle release. We find that the axons of individual tetanus toxin expressing reticulospinal neurons have fewer myelin sheaths than controls and that their myelin sheaths are 50% shorter than controls. In stark contrast, myelination along tetanus-toxin-expressing CoPA neuron axons is entirely normal. These results indicate that while some neuronal subtypes modulate myelination by synaptic vesicle release to a striking degree in vivo, others do not. These data have implications for our understanding of how different neurons regulate myelination and thus their own function within specific neuronal circuits.

    View details for DOI 10.1016/j.cub.2016.03.070

    View details for Web of Science ID 000377390700023

    View details for PubMedID 27161502

    View details for PubMedCentralID PMC4906267

  • The Rag-Ragulator Complex Regulates Lysosome Function and Phagocytic Flux in Microglia. Cell reports Shen, K., Sidik, H., Talbot, W. S. 2016; 14 (3): 547-559

    Abstract

    Microglia are resident macrophages of the CNS that are essential for phagocytosis of apoptotic neurons and weak synapses during development. We show that RagA and Lamtor4, two components of the Rag-Ragulator complex, are essential regulators of lysosomes in microglia. In zebrafish lacking RagA function, microglia exhibit an expanded lysosomal compartment, but they are unable to properly digest apoptotic neuronal debris. Previous biochemical studies have placed the Rag-Ragulator complex upstream of mTORC1 activation in response to cellular nutrient availability. Nonetheless, RagA and mTOR mutant zebrafish have distinct phenotypes, indicating that the Rag-Ragulator complex has functions independent of mTOR signaling. Our analysis reveals an essential role of the Rag-Ragulator complex in proper lysosome function and phagocytic flux in microglia.

    View details for DOI 10.1016/j.celrep.2015.12.055

    View details for PubMedID 26774477

    View details for PubMedCentralID PMC4731305

  • A zinc finger protein that regulates oligodendrocyte specification, migration and myelination in zebrafish DEVELOPMENT Sidik, H., Talbot, W. S. 2015; 142 (23): 4119-4128

    Abstract

    Precise control of oligodendrocyte migration and development is crucial for myelination of axons in the central nervous system (CNS), but important questions remain unanswered about the mechanisms controlling these processes. In a zebrafish screen for myelination mutants, we identified a mutation in zinc finger protein 16-like (znf16l). znf16l mutant larvae have reduced myelin basic protein (mbp) expression and reduced CNS myelin. Marker, time-lapse and ultrastructural studies indicated that oligodendrocyte specification, migration and myelination are disrupted in znf16l mutants. Transgenic studies indicated that znf16l acts autonomously in oligodendrocytes. Expression of Zfp488 from mouse rescued mbp expression in znf16l mutants, indicating that these homologs have overlapping functions. Our results defined the function of a new zinc finger protein with specific function in oligodendrocyte specification, migration and myelination in the developing CNS.

    View details for DOI 10.1242/dev.128215

    View details for Web of Science ID 000366363800012

    View details for PubMedID 26459222

    View details for PubMedCentralID PMC4712842

  • Autosomal recessive mutations of GPR126 are responsible for severe Arthrogryposis multiplex congenita Ravenscroft, G., Nolen, F., Rajagopalan, S., Meireles, A., Paavola, K., Gaillard, D., Alanio, E., Buckland, M., Arbuckle, S., Krivanek, M., Maluenda, J., Pannell, S., Gooding, R., Ong, R., Allcock, R., Kok, F., Talbot, W., Melki, J., Laing, N. PERGAMON-ELSEVIER SCIENCE LTD. 2015: S186
  • Mutations of GPR126 Are Responsible for Severe Arthrogryposis Multiplex Congenita AMERICAN JOURNAL OF HUMAN GENETICS Ravenscroft, G., Nolent, F., Rajagopalan, S., Meireles, A. M., Paavola, K. J., Gaillard, D., Alanio, E., Buckland, M., Arbuckle, S., Krivanek, M., Maluenda, J., Pannell, S., Gooding, R., Ong, R. W., Allcock, R. J., Carvalho, E. D., Carvalho, M. D., Kok, F., Talbot, W. S., Melki, J., Laing, N. G. 2015; 96 (6): 955-961

    Abstract

    Arthrogryposis multiplex congenita is defined by the presence of contractures across two or more major joints and results from reduced or absent fetal movement. Here, we present three consanguineous families affected by lethal arthrogryposis multiplex congenita. By whole-exome or targeted exome sequencing, it was shown that the probands each harbored a different homozygous mutation (one missense, one nonsense, and one frameshift mutation) in GPR126. GPR126 encodes G-protein-coupled receptor 126, which has been shown to be essential for myelination of axons in the peripheral nervous system in fish and mice. A previous study reported that Gpr126(-/-) mice have a lethal arthrogryposis phenotype. We have shown that the peripheral nerves in affected individuals from one family lack myelin basic protein, suggesting that this disease in affected individuals is due to defective myelination of the peripheral axons during fetal development. Previous work has suggested that autoproteolytic cleavage is important for activating GPR126 signaling, and our biochemical assays indicated that the missense substitution (p.Val769Glu [c.2306T>A]) impairs autoproteolytic cleavage of GPR126. Our data indicate that GPR126 is critical for myelination of peripheral nerves in humans. This study adds to the literature implicating defective axoglial function as a key cause of severe arthrogryposis multiplex congenita and suggests that GPR126 mutations should be investigated in individuals affected by this disorder.

    View details for DOI 10.1016/j.ajhg.2015.04.014

    View details for PubMedID 26004201

  • Glial Cell Development and Function in Zebrafish COLD SPRING HARBOR PERSPECTIVES IN BIOLOGY Lyons, D. A., Talbot, W. S. 2015; 7 (2)

    Abstract

    The zebrafish is a premier vertebrate model system that offers many experimental advantages for in vivo imaging and genetic studies. This review provides an overview of glial cell types in the central and peripheral nervous system of zebrafish. We highlight some recent work that exploited the strengths of the zebrafish system to increase the understanding of the role of Gpr126 in Schwann cell myelination and illuminate the mechanisms controlling oligodendrocyte development and myelination. We also summarize similarities and differences between zebrafish radial glia and mammalian astrocytes and consider the possibility that their distinct characteristics may represent extremes in a continuum of cell identity. Finally, we focus on the emergence of zebrafish as a model for elucidating the development and function of microglia. These recent studies have highlighted the power of the zebrafish system for analyzing important aspects of glial development and function.

    View details for DOI 10.1101/cshperspect.a020586

    View details for Web of Science ID 000351203700011

  • Differential Requirement for irf8 in Formation of Embryonic and Adult Macrophages in Zebrafish. PloS one Shiau, C. E., Kaufman, Z., Meireles, A. M., Talbot, W. S. 2015; 10 (1)

    Abstract

    Interferon regulatory factor 8 (Irf8) is critical for mammalian macrophage development and innate immunity, but its role in teleost myelopoiesis remains incompletely understood. In particular, genetic tools to analyze the role of Irf8 in zebrafish macrophage development at larval and adult stages are lacking. We generated irf8 null mutants in zebrafish using TALEN-mediated targeting. Our analysis defines different requirements for irf8 at different stages. irf8 is required for formation of all macrophages during primitive and transient definitive hematopoiesis, but not during adult-phase definitive hematopoiesis starting at 5-6 days postfertilization. At early stages, irf8 mutants have excess neutrophils and excess cell death in pu.1-expressing myeloid cells. Macrophage fates were recovered in irf8 mutants after wildtype irf8 expression in neutrophil and macrophage lineages, suggesting that irf8 regulates macrophage specification and survival. In juvenile irf8 mutant fish, mature macrophages are present, but at numbers significantly reduced compared to wildtype, indicating an ongoing requirement for irf8 after embryogenesis. As development progresses, tissue macrophages become apparent in zebrafish irf8 mutants, with the possible exception of microglia. Our study defines distinct requirement for irf8 in myelopoiesis before and after transition to the adult hematopoietic system.

    View details for DOI 10.1371/journal.pone.0117513

    View details for PubMedID 25615614

    View details for PubMedCentralID PMC4304715

  • Unique function of Kinesin Kif5A in localization of mitochondria in axons. journal of neuroscience Campbell, P. D., Shen, K., Sapio, M. R., Glenn, T. D., Talbot, W. S., Marlow, F. L. 2014; 34 (44): 14717-14732

    Abstract

    Mutations in Kinesin proteins (Kifs) are linked to various neurological diseases, but the specific and redundant functions of the vertebrate Kifs are incompletely understood. For example, Kif5A, but not other Kinesin-1 heavy-chain family members, is implicated in Charcot-Marie-Tooth disease (CMT) and Hereditary Spastic Paraplegia (HSP), but the mechanism of its involvement in the progressive axonal degeneration characteristic of these diseases is not well understood. We report that zebrafish kif5Aa mutants exhibit hyperexcitability, peripheral polyneuropathy, and axonal degeneration reminiscent of CMT and HSP. Strikingly, although kif5 genes are thought to act largely redundantly in other contexts, and zebrafish peripheral neurons express five kif5 genes, kif5Aa mutant peripheral sensory axons lack mitochondria and degenerate. We show that this Kif5Aa-specific function is cell autonomous and is mediated by its C-terminal tail, as only Kif5Aa and chimeric motors containing the Kif5Aa C-tail can rescue deficits. Finally, concurrent loss of the kinesin-3, kif1b, or its adaptor kbp, exacerbates axonal degeneration via a nonmitochondrial cargo common to Kif5Aa. Our results shed light on Kinesin complexity and reveal determinants of specific Kif5A functions in mitochondrial transport, adaptor binding, and axonal maintenance.

    View details for DOI 10.1523/JNEUROSCI.2770-14.2014

    View details for PubMedID 25355224

  • The Phosphate Exporter xpr1b Is Required for Differentiation of Tissue-Resident Macrophages CELL REPORTS Meireles, A. M., Shiau, C. E., Guenther, C. A., Sidik, H., Kingsley, D. M., Talbot, W. S. 2014; 8 (6): 1659-1667

    Abstract

    Phosphate concentration is tightly regulated at the cellular and organismal levels. The first metazoan phosphate exporter, XPR1, was recently identified, but its in vivo function remains unknown. In a genetic screen, we identified a mutation in a zebrafish ortholog of human XPR1, xpr1b. xpr1b mutants lack microglia, the specialized macrophages that reside in the brain, and also displayed an osteopetrotic phenotype characteristic of defects in osteoclast function. Transgenic expression studies indicated that xpr1b acts autonomously in developing macrophages. xpr1b mutants display no gross developmental defects that may arise from phosphate imbalance. We constructed a targeted mutation of xpr1a, a duplicate of xpr1b in the zebrafish genome, to determine whether Xpr1a and Xpr1b have redundant functions. Single mutants for xpr1a were viable, and double mutants for xpr1b;xpr1a were similar to xpr1b single mutants. Our genetic analysis reveals a specific role for the phosphate exporter Xpr1 in the differentiation of tissue macrophages.

    View details for DOI 10.1016/j.celrep.2014.08.018

    View details for Web of Science ID 000343867400007

    View details for PubMedCentralID PMC4177277

  • The phosphate exporter xpr1b is required for differentiation of tissue-resident macrophages. Cell reports Meireles, A. M., Shiau, C. E., Guenther, C. A., Sidik, H., Kingsley, D. M., Talbot, W. S. 2014; 8 (6): 1659-1667

    Abstract

    Phosphate concentration is tightly regulated at the cellular and organismal levels. The first metazoan phosphate exporter, XPR1, was recently identified, but its in vivo function remains unknown. In a genetic screen, we identified a mutation in a zebrafish ortholog of human XPR1, xpr1b. xpr1b mutants lack microglia, the specialized macrophages that reside in the brain, and also displayed an osteopetrotic phenotype characteristic of defects in osteoclast function. Transgenic expression studies indicated that xpr1b acts autonomously in developing macrophages. xpr1b mutants display no gross developmental defects that may arise from phosphate imbalance. We constructed a targeted mutation of xpr1a, a duplicate of xpr1b in the zebrafish genome, to determine whether Xpr1a and Xpr1b have redundant functions. Single mutants for xpr1a were viable, and double mutants for xpr1b;xpr1a were similar to xpr1b single mutants. Our genetic analysis reveals a specific role for the phosphate exporter Xpr1 in the differentiation of tissue macrophages.

    View details for DOI 10.1016/j.celrep.2014.08.018

    View details for PubMedID 25220463

  • Type IV collagen is an activating ligand for the adhesion G protein-coupled receptor GPR126. Science signaling Paavola, K. J., Sidik, H., Zuchero, J. B., Eckart, M., Talbot, W. S. 2014; 7 (338): ra76-?

    Abstract

    GPR126 is an orphan heterotrimeric guanine nucleotide-binding protein (G protein)-coupled receptor (GPCR) that is essential for the development of diverse organs. We found that type IV collagen, a major constituent of the basement membrane, binds to Gpr126 and activates its signaling function. Type IV collagen stimulated the production of cyclic adenosine monophosphate in rodent Schwann cells, which require Gpr126 activity to differentiate, and in human embryonic kidney (HEK) 293 cells expressing exogenous Gpr126. Type IV collagen specifically bound to the extracellular amino-terminal region of Gpr126 containing the CUB (complement, Uegf, Bmp1) and pentraxin domains. Gpr126 derivatives lacking the entire amino-terminal region were constitutively active, suggesting that this region inhibits signaling and that ligand binding relieves this inhibition to stimulate receptor activity. A new zebrafish mutation that truncates Gpr126 after the CUB and pentraxin domains disrupted development of peripheral nerves and the inner ear. Thus, our findings identify type IV collagen as an activating ligand for GPR126, define its mechanism of activation, and highlight a previously unrecognized signaling function of type IV collagen in basement membranes.

    View details for DOI 10.1126/scisignal.2005347

    View details for PubMedID 25118328

  • Glial cell development and function in zebrafish. Cold Spring Harbor perspectives in biology Lyons, D. A., Talbot, W. S. 2014; 7 (2)

    Abstract

    The zebrafish is a premier vertebrate model system that offers many experimental advantages for in vivo imaging and genetic studies. This review provides an overview of glial cell types in the central and peripheral nervous system of zebrafish. We highlight some recent work that exploited the strengths of the zebrafish system to increase the understanding of the role of Gpr126 in Schwann cell myelination and illuminate the mechanisms controlling oligodendrocyte development and myelination. We also summarize similarities and differences between zebrafish radial glia and mammalian astrocytes and consider the possibility that their distinct characteristics may represent extremes in a continuum of cell identity. Finally, we focus on the emergence of zebrafish as a model for elucidating the development and function of microglia. These recent studies have highlighted the power of the zebrafish system for analyzing important aspects of glial development and function.

    View details for DOI 10.1101/cshperspect.a020586

    View details for PubMedID 25395296

  • Signals regulating myelination in peripheral nerves and the Schwann cell response to injury. Current opinion in neurobiology Glenn, T. D., Talbot, W. S. 2013; 23 (6): 1041-1048

    Abstract

    In peripheral nerves, Schwann cells form myelin, which facilitates the rapid conduction of action potentials along axons in the vertebrate nervous system. Myelinating Schwann cells are derived from neural crest progenitors in a step-wise process that is regulated by extracellular signals and transcription factors. In addition to forming the myelin sheath, Schwann cells orchestrate much of the regenerative response that occurs after injury to peripheral nerves. In response to injury, myelinating Schwann cells dedifferentiate into repair cells that are essential for axonal regeneration, and then redifferentiate into myelinating Schwann cells to restore nerve function. Although this remarkable plasticity has long been recognized, many questions remain unanswered regarding the signaling pathways regulating both myelination and the Schwann cell response to injury.

    View details for DOI 10.1016/j.conb.2013.06.010

    View details for PubMedID 23896313

    View details for PubMedCentralID PMC3830599

  • An Anti-inflammatory NOD-like Receptor Is Required for Microglia Development CELL REPORTS Shiau, C. E., Monk, K. R., Joo, W., Talbot, W. S. 2013; 5 (5): 1342-1352

    Abstract

    Microglia are phagocytic cells that form the basis of the brain's immune system. They derive from primitive macrophages that migrate into the brain during embryogenesis, but the genetic control of microglial development remains elusive. Starting with a genetic screen in zebrafish, we show that the noncanonical NOD-like receptor (NLR) nlrc3-like is essential for microglial formation. Although most NLRs trigger inflammatory signaling, nlrc3-like acts cell autonomously in microglia precursor cells to suppress unwarranted inflammation in the absence of overt immune challenge. In nlrc3-like mutants, primitive macrophages initiate a systemic inflammatory response with increased proinflammatory cytokines and actively aggregate instead of migrating into the brain to form microglia. NLRC3-like requires both its pyrin and NACHT domains, and it can bind the inflammasome component apoptosis-associated speck-like protein. Our studies suggest that NLRC3-like may regulate the inflammasome and other inflammatory pathways. Together, these results demonstrate that NLRC3-like prevents inappropriate macrophage activation, thereby allowing normal microglial development.

    View details for DOI 10.1016/j.celrep.2013.11.004

    View details for Web of Science ID 000328266400017

    View details for PubMedID 24316075

    View details for PubMedCentralID PMC3878655

  • notch3 is essential for oligodendrocyte development and vascular integrity in zebrafish DISEASE MODELS & MECHANISMS Zaucker, A., Mercurio, S., Sternheim, N., Talbot, W. S., Marlow, F. L. 2013; 6 (5): 1246-1259

    Abstract

    Mutations in the human NOTCH3 gene cause Cerebral Autosomal Dominant Arteriopathy with Subcortical Infarcts and Leukoencephalopathy (CADASIL). CADASIL is an inherited small vessel disease characterized by diverse clinical manifestations including vasculopathy, neurodegeneration, and dementia. Here we report two mutations in the zebrafish notch3 gene, one identified in a previous screen for mutations with reduced expression of myelin basic protein (mbp) and another caused by a retroviral insertion. Reduced mbp expression in embryos is associated with fewer oligodendrocyte precursor cells (OPCs) in notch3 mutants. Despite an early neurogenic phenotype, mbp expression recovered at later developmental stages and some of notch3 homozygous mutants survived to adulthood. Adult mutants for both notch3 alleles displayed a striking stress-associated accumulation of blood. Histological analysis of mutant vessels revealed vasculopathy, including an enlargement (dilation) of vessels in the telencephalon and fin, disorganization of the normal stereotyped arrangement of vessels in the fin, and an apparent loss of arterial morphological structure. Expression of hey1, a well-known transcriptional target of Notch signaling, was greatly reduced in notch3 mutants, suggesting Notch3 acts via a canonical Notch signaling pathway to promote normal vessel structure. Ultrastructural analysis confirmed the presence of dilated vessels in notch3 mutant fins and revealed that the vessel walls of presumed arteries showed signs of deterioration. Gaps evident in the arterial wall and the presence of blood cells outside of vessels in mutants indicated that compromised vessel structure led to hemorrhage. In notch3 heterozygotes, we found elevated expression of notch3 itself and target genes, indicating that specific alterations in gene expression caused by a partial loss of Notch3 function may contribute to pathologies observed in heterozygous larvae and adults. Our analysis of zebrafish notch3 mutants indicates that Notch3 regulates OPC development and mbp gene expression in larvae, and maintains vascular integrity in adults.

    View details for DOI 10.1242/dmm.012005

    View details for Web of Science ID 000325789600020

    View details for PubMedID 23720232

  • Analysis of Gpr126 function defines distinct mechanisms controlling the initiation and maturation of myelin DEVELOPMENT Glenn, T. D., Talbot, W. S. 2013; 140 (15): 3167-3175

    Abstract

    In peripheral nerves, Schwann cells form the myelin sheath, which allows the efficient propagation of action potentials along axons. The transcription factor Krox20 regulates the initiation of myelination in Schwann cells and is also required to maintain mature myelin. The adhesion G protein-coupled receptor (GPCR) Gpr126 is essential for Schwann cells to initiate myelination, but previous studies have not addressed the role of Gpr126 signaling in myelin maturation and maintenance. Through analysis of Gpr126 in zebrafish, we define two distinct mechanisms controlling the initiation and maturation of myelin. We show that gpr126 mutant Schwann cells elaborate mature myelin sheaths and maintain krox20 expression for months, provided that the early signaling defect is bypassed by transient elevation of cAMP. At the onset of myelination, Gpr126 and protein kinase A (PKA) function as a switch that allows Schwann cells to initiate krox20 expression and myelination. After myelination is initiated, krox20 expression is maintained and myelin maturation proceeds independently of Gpr126 signaling. Transgenic analysis indicates that the Krox20 cis-regulatory myelinating Schwann cell element (MSE) becomes active at the onset of myelination and that this activity is dependent on Gpr126 signaling. Activity of the MSE declines after initiation, suggesting that other elements are responsible for maintaining krox20 expression in mature nerves. We also show that elevated cAMP does not initiate myelination in the absence of functional Neuregulin 1 (Nrg1) signaling. These results indicate that the mechanisms regulating the initiation of myelination are distinct from those mediating the maturation and maintenance of myelin.

    View details for DOI 10.1242/dev.093401

    View details for Web of Science ID 000321864900010

    View details for PubMedID 23804499

  • Can't wait to myelinate. Developmental cell Glenn, T. D., Talbot, W. S. 2013; 25 (6): 549-550

    Abstract

    Oligodendrocytes myelinate axons in the central nervous system (CNS). In this issue of Developmental Cell, Czopka et al. (2013) shed light on the temporal control of myelination by individual cells. They demonstrate that oligodendrocytes in vivo have only a brief time window to initiate myelination, which has important implications for CNS plasticity.

    View details for DOI 10.1016/j.devcel.2013.06.003

    View details for PubMedID 23806613

  • Mutation of sec63 in zebrafish causes defects in myelinated axons and liver pathology DISEASE MODELS & MECHANISMS Monk, K. R., Voas, M. G., Franzini-Armstrong, C., Hakkinen, I. S., Talbot, W. S. 2013; 6 (1): 135-145

    Abstract

    Mutations in SEC63 cause polycystic liver disease in humans. Sec63 is a member of the endoplasmic reticulum (ER) translocon machinery, although it is unclear how mutations in SEC63 lead to liver cyst formation in humans. Here, we report the identification and characterization of a zebrafish sec63 mutant, which was discovered in a screen for mutations that affect the development of myelinated axons. Accordingly, we show that disruption of sec63 in zebrafish leads to abnormalities in myelinating glia in both the central and peripheral nervous systems. In the vertebrate nervous system, segments of myelin are separated by the nodes of Ranvier, which are unmyelinated regions of axonal membrane containing a high density of voltage-gated sodium channels. We show that sec63 mutants have morphologically abnormal and reduced numbers of clusters of voltage-gated sodium channels in the spinal cord and along peripheral nerves. Additionally, we observed reduced myelination in both the central and peripheral nervous systems, as well as swollen ER in myelinating glia. Markers of ER stress are upregulated in sec63 mutants. Finally, we show that sec63 mutants develop liver pathology. As in glia, the primary defect, detectable at 5 dpf, is fragmentation and swelling of the ER, indicative of accumulation of proteins in the lumen. At 8 dpf, ER swelling is severe; other pathological features include disrupted bile canaliculi, altered cytoplasmic matrix and accumulation of large lysosomes. Together, our analyses of sec63 mutant zebrafish highlight the possible role of ER stress in polycystic liver disease and suggest that these mutants will serve as a model for understanding the pathophysiology of this disease and other abnormalities involving ER stress.

    View details for DOI 10.1242/dmm.009217

    View details for Web of Science ID 000314865700015

    View details for PubMedID 22864019

    View details for PubMedCentralID PMC3529346

  • Scube/You activity mediates release of dually lipid-modified Hedgehog signal in soluble form GENES & DEVELOPMENT Creanga, A., Glenn, T. D., Mann, R. K., Saunders, A. M., Talbot, W. S., Beachy, P. A. 2012; 26 (12): 1312-1325

    Abstract

    Owing to their covalent modification by cholesterol and palmitate, Hedgehog (Hh) signaling proteins are localized predominantly to the plasma membrane of expressing cells. Yet Hh proteins are also capable of mobilizing to and eliciting direct responses from distant cells. The zebrafish you gene, identified genetically >15 years ago, was more recently shown to encode a secreted glycoprotein that acts cell-nonautonomously in the Hh signaling pathway by an unknown mechanism. We investigated the function of the protein encoded by murine Scube2, an ortholog of you, and found that it mediates release in soluble form of the mature, cholesterol- and palmitate-modified Sonic hedgehog protein signal (ShhNp) when added to cultured cells or purified detergent-resistant membrane microdomains containing ShhNp. The efficiency of Scube2-mediated release of ShhNp is enhanced by the palmitate adduct of ShhNp and by coexpression in ShhNp-producing cells of mDispatchedA (mDispA), a transporter-like protein with a previously defined role in the release of lipid-modified Hh signals. The structural determinants of Scube2 required for its activity in cultured cell assays match those required for rescue of you mutant zebrafish embryos, and we thus conclude that the role of Scube/You proteins in Hh signaling in vivo is to facilitate the release and mobilization of Hh proteins for distant action.

    View details for DOI 10.1101/gad.191866.112

    View details for Web of Science ID 000305485300006

    View details for PubMedID 22677548

    View details for PubMedCentralID PMC3387659

  • Neuronal Neuregulin 1 type III directs Schwann cell migration DEVELOPMENT Perlin, J. R., Lush, M. E., Stephens, W. Z., Piotrowski, T., Talbot, W. S. 2011; 138 (21): 4639-4648

    Abstract

    During peripheral nerve development, each segment of a myelinated axon is matched with a single Schwann cell. Tight regulation of Schwann cell movement, proliferation and differentiation is essential to ensure that these glial cells properly associate with axons. ErbB receptors are required for Schwann cell migration, but the operative ligand and its mechanism of action have remained unknown. We demonstrate that zebrafish Neuregulin 1 (Nrg1) type III, which signals through ErbB receptors, controls Schwann cell migration in addition to its previously known roles in proliferation and myelination. Chimera analyses indicate that ErbB receptors are required in all migrating Schwann cells, and that Nrg1 type III is required in neurons for migration. Surprisingly, expression of the ligand in a few axons is sufficient to induce migration along a chimeric nerve constituted largely of nrg1 type III mutant axons. These studies also reveal a mechanism that allows Schwann cells to fasciculate axons regardless of nrg1 type III expression. Time-lapse imaging of transgenic embryos demonstrated that misexpression of human NRG1 type III results in ectopic Schwann cell migration, allowing them to aberrantly enter the central nervous system. These results demonstrate that Nrg1 type III is an essential signal that controls Schwann cell migration to ensure that these glia are present in the correct numbers and positions in developing nerves.

    View details for DOI 10.1242/dev.068072

    View details for Web of Science ID 000296060100008

    View details for PubMedID 21965611

    View details for PubMedCentralID PMC3190382

  • ErbB Signaling Has a Role in Radial Sorting Independent of Schwann Cell Number GLIA Raphael, A. R., Lyons, D. A., Talbot, W. S. 2011; 59 (7): 1047-1055

    Abstract

    In the peripheral nervous system, Schwann cells make myelin, a specialized sheath that is essential for rapid axonal conduction of action potentials. Immature Schwann cells initially interact with many axons, but, through a process termed radial sorting, eventually interact with one segment of a single axon as promyelinating Schwann cells. Previous studies have identified genes that are required for Schwann cell process extension and proliferation during radial sorting. Previous analyses also show that ErbB signaling is required for Schwann cell proliferation, myelination, radial sorting, and the proper formation of unmyelinated Remak bundles. Because ErbB signaling and Schwann cell proliferation are both required during radial sorting, we sought to determine if the primary function of ErbB signaling in this process is to regulate Schwann cell proliferation or if ErbB signaling also controls other aspects of radial sorting. To address this question, we applied small molecule inhibitors in vivo in zebrafish to independently block ErbB signaling and proliferation. Ultrastructural analysis of treated animals revealed that both ErbB signaling and Schwann cell proliferation are required for radial sorting in vivo. ErbB signaling, however, is required for Schwann cell process extension, while Schwann cell proliferation is not. These results provide in vivo evidence that ErbB signaling plays a direct role in process extension during radial sorting, in addition to its role in regulating Schwann cell proliferation.

    View details for DOI 10.1002/glia.21175

    View details for Web of Science ID 000291434500004

    View details for PubMedID 21491500

    View details for PubMedCentralID PMC3094506

  • Gpr126 is essential for peripheral nerve development and myelination in mammals DEVELOPMENT Monk, K. R., Oshima, K., Joers, S., Heller, S., Talbot, W. S. 2011; 138 (13): 2673-2680

    Abstract

    In peripheral nerves, Schwann cells form the myelin sheath that insulates axons and allows rapid propagation of action potentials. Although a number of regulators of Schwann cell development are known, the signaling pathways that control myelination are incompletely understood. In this study, we show that Gpr126 is essential for myelination and other aspects of peripheral nerve development in mammals. A mutation in Gpr126 causes a severe congenital hypomyelinating peripheral neuropathy in mice, and expression of differentiated Schwann cell markers, including Pou3f1, Egr2, myelin protein zero and myelin basic protein, is reduced. Ultrastructural studies of Gpr126-/- mice showed that axonal sorting by Schwann cells is delayed, Remak bundles (non-myelinating Schwann cells associated with small caliber axons) are not observed, and Schwann cells are ultimately arrested at the promyelinating stage. Additionally, ectopic perineurial fibroblasts form aberrant fascicles throughout the endoneurium of the mutant sciatic nerve. This analysis shows that Gpr126 is required for Schwann cell myelination in mammals, and defines new roles for Gpr126 in axonal sorting, formation of mature non-myelinating Schwann cells and organization of the perineurium.

    View details for DOI 10.1242/dev.062224

    View details for Web of Science ID 000291348700005

    View details for PubMedID 21613327

    View details for PubMedCentralID PMC3109596

  • Schwann cell spectrins modulate peripheral nerve myelination PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Susuki, K., Raphael, A. R., Ogawa, Y., Stankewich, M. C., Peles, E., Talbot, W. S., Rasband, M. N. 2011; 108 (19): 8009-8014

    Abstract

    During peripheral nerve development, Schwann cells ensheathe axons and form myelin to enable rapid and efficient action potential propagation. Although myelination requires profound changes in Schwann cell shape, how neuron-glia interactions converge on the Schwann cell cytoskeleton to induce these changes is unknown. Here, we demonstrate that the submembranous cytoskeletal proteins αII and βII spectrin are polarized in Schwann cells and colocalize with signaling molecules known to modulate myelination in vitro. Silencing expression of these spectrins inhibited myelination in vitro, and remyelination in vivo. Furthermore, myelination was disrupted in motor nerves of zebrafish lacking αII spectrin. Finally, we demonstrate that loss of spectrin significantly reduces both F-actin in the Schwann cell cytoskeleton and the Nectin-like protein, Necl4, at the contact site between Schwann cells and axons. Therefore, we propose αII and βII spectrin in Schwann cells integrate the neuron-glia interactions mediated by membrane proteins into the actin-dependent cytoskeletal rearrangements necessary for myelination.

    View details for DOI 10.1073/pnas.1019600108

    View details for Web of Science ID 000290439500070

    View details for PubMedID 21518878

    View details for PubMedCentralID PMC3093478

  • Schwann cells reposition a peripheral nerve to isolate it from postembryonic remodeling of its targets DEVELOPMENT Raphael, A. R., Perlin, J. R., Talbot, W. S. 2010; 137 (21): 3643-3649

    Abstract

    Although much is known about the initial construction of the peripheral nervous system (PNS), less well understood are the processes that maintain the position and connections of nerves during postembryonic growth. Here, we show that the posterior lateral line nerve in zebrafish initially grows in the epidermis and then rapidly transitions across the epidermal basement membrane into the subepidermal space. Our experiments indicate that Schwann cells, which myelinate axons in the PNS, are required to reposition the nerve. In mutants lacking Schwann cells, the nerve is mislocalized and the axons remain in the epidermis. Transplanting wild-type Schwann cells into these mutants rescues the position of the nerve. Analysis of chimeric embryos suggests that the process of nerve relocalization involves two discrete steps - the degradation and recreation of the epidermal basement membrane. Although the outgrowth of axons is normal in mutants lacking Schwann cells, the nerve becomes severely disorganized at later stages. In wild-type embryos, exclusion of the nerve from the epidermis isolates axons from migration of their targets (sensory neuromasts) within the epidermis. Without Schwann cells, axons remain within the epidermis and are dragged along with the migrating neuromasts. Our analysis of the posterior lateral line system defines a new process in which Schwann cells relocate a nerve beneath the epidermal basement membrane to insulate axons from the postembryonic remodeling of their targets.

    View details for DOI 10.1242/dev.057521

    View details for Web of Science ID 000283669300012

    View details for PubMedID 20876648

    View details for PubMedCentralID PMC2964096

  • Schwann Cells Inhibit Ectopic Clustering of Axonal Sodium Channels JOURNAL OF NEUROSCIENCE Voas, M. G., Glenn, T. D., Raphael, A. R., Talbot, W. S. 2009; 29 (46): 14408-14414

    Abstract

    The clustering of voltage-gated sodium channels at the axon initial segment (AIS) and nodes of Ranvier is essential for the initiation and propagation of action potentials in myelinated axons. Sodium channels localize to the AIS through an axon-intrinsic mechanism driven by ankyrin G, while clustering at the nodes requires cues from myelinating glia that interact with axonal neurofascin186 (Sherman et al., 2005; Dzhashiashvili et al., 2007; Yang et al., 2007). Here, we report that in zebrafish mutants lacking Schwann cells in peripheral nerves (erbb2, erbb3, and sox10/colorless), axons form numerous aberrant sodium channel clusters throughout their length. Morpholino knockdown of ankyrin G, but not neurofascin, reduces the number of sodium channel clusters in Schwann cell-deficient mutants, suggesting that these aberrant clusters form by an axon-intrinsic mechanism. We also find that gpr126 mutants, in which Schwann cells are arrested at the promyelinating stage (Monk et al., 2009), are deficient in the clustering of neurofascin at the nodes of Ranvier. When Schwann cell migration in gpr126 mutants is blocked, there is an increase in the number of neurofascin clusters in peripheral axons. Our results suggest that Schwann cells inhibit the ability of ankyrin G to cluster sodium channels at ectopic locations, restricting its activity to the AIS and nodes of Ranvier.

    View details for DOI 10.1523/JNEUROSCI.0841-09.2009

    View details for Web of Science ID 000271944500004

    View details for PubMedID 19923275

    View details for PubMedCentralID PMC2826614

  • Genetic dissection of myellinated axons in zebrafish CURRENT OPINION IN NEUROBIOLOGY Monk, K. R., Talbot, W. S. 2009; 19 (5): 486-490

    Abstract

    In the vertebrate nervous system, the myelin sheath allows for rapid and efficient conduction of action potentials along axons. Despite the essential function of myelin, many questions remain unanswered about the mechanisms that govern the development of myelinated axons. The fundamental properties of myelin are widely shared among vertebrates, and the zebrafish has emerged as a powerful system to study myelination in vivo. This review will highlight recent advances from genetic screens in zebrafish, including the discovery of the role of kif1b in mRNA localization in myelinating oligodendrocytes.

    View details for DOI 10.1016/j.conb.2009.08.006

    View details for Web of Science ID 000272922900005

    View details for PubMedID 19740648

    View details for PubMedCentralID PMC2787740

  • A G Protein-Coupled Receptor Is Essential for Schwann Cells to Initiate Myelination SCIENCE Monk, K. R., Naylor, S. G., Glenn, T. D., Mercurio, S., Perlin, J. R., Dominguez, C., Moens, C. B., Talbot, W. S. 2009; 325 (5946): 1402-1405

    Abstract

    The myelin sheath allows axons to conduct action potentials rapidly in the vertebrate nervous system. Axonal signals activate expression of specific transcription factors, including Oct6 and Krox20, that initiate myelination in Schwann cells. Elevation of cyclic adenosine monophosphate (cAMP) can mimic axonal contact in vitro, but the mechanisms that regulate cAMP levels in vivo are unknown. Using mutational analysis in zebrafish, we found that the G protein-coupled receptor Gpr126 is required autonomously in Schwann cells for myelination. In gpr126 mutants, Schwann cells failed to express oct6 and krox20 and were arrested at the promyelinating stage. Elevation of cAMP in gpr126 mutants, but not krox20 mutants, could restore myelination. We propose that Gpr126 drives the differentiation of promyelinating Schwann cells by elevating cAMP levels, thereby triggering Oct6 expression and myelination.

    View details for DOI 10.1126/science.1173474

    View details for Web of Science ID 000269699100041

    View details for PubMedID 19745155

    View details for PubMedCentralID PMC2856697

  • Kif1b is essential for mRNA localization in oligodendrocytes and development of myelinated axons NATURE GENETICS Lyons, D. A., Naylor, S. G., Scholze, A., Talbot, W. S. 2009; 41 (7): 854-U121

    Abstract

    The kinesin motor protein Kif1b has previously been implicated in the axonal transport of mitochondria and synaptic vesicles. More recently, KIF1B has been associated with susceptibility to multiple sclerosis (MS). Here we show that Kif1b is required for the localization of mbp (myelin basic protein) mRNA to processes of myelinating oligodendrocytes in zebrafish. We observe the ectopic appearance of myelin-like membrane in kif1b mutants, coincident with the ectopic localization of myelin proteins in kif1b mutant oligodendrocyte cell bodies. These observations suggest that oligodendrocytes localize certain mRNA molecules, namely those encoding small basic proteins such as MBP, to prevent aberrant effects of these proteins elsewhere in the cell. We also find that Kif1b is required for outgrowth of some of the longest axons in the peripheral and central nervous systems. Our data demonstrate previously unknown functions of kif1b in vivo and provide insights into its possible roles in MS.

    View details for DOI 10.1038/ng.376

    View details for Web of Science ID 000267786200020

    View details for PubMedID 19503091

    View details for PubMedCentralID PMC2702462

  • Axonal domains: Role for paranodal junction in node of Ranvier assembly CURRENT BIOLOGY Lyons, D. A., Talbot, W. S. 2008; 18 (18): R876-R879

    Abstract

    A new study shows that communication between axons and glia at the paranodal junction can orchestrate the formation of the node of Ranvier.

    View details for DOI 10.1016/j.cub.2008.07.070

    View details for Web of Science ID 000259523600018

    View details for PubMedID 18812088

  • The ATPase-dependent chaperoning activity of Hsp90a regulates thick filament formation and integration during skeletal muscle myofibrillogenesis DEVELOPMENT Hawkins, T. A., Haramis, A., Etard, C., Prodromou, C., Vaughan, C. K., Ashworth, R., Ray, S., Behra, M., Holder, N., Talbot, W. S., Pearl, L. H., Strahle, U., Wilson, S. W. 2008; 135 (6): 1147-1156

    Abstract

    The mechanisms that regulate sarcomere assembly during myofibril formation are poorly understood. In this study, we characterise the zebrafish sloth(u45) mutant, in which the initial steps in sarcomere assembly take place, but thick filaments are absent and filamentous I-Z-I brushes fail to align or adopt correct spacing. The mutation only affects skeletal muscle and mutant embryos show no other obvious phenotypes. Surprisingly, we find that the phenotype is due to mutation in one copy of a tandemly duplicated hsp90a gene. The mutation disrupts the chaperoning function of Hsp90a through interference with ATPase activity. Despite being located only 2 kb from hsp90a, hsp90a2 has no obvious role in sarcomere assembly. Loss of Hsp90a function leads to the downregulation of genes encoding sarcomeric proteins and upregulation of hsp90a and several other genes encoding proteins that may act with Hsp90a during sarcomere assembly. Our studies reveal a surprisingly specific developmental role for a single Hsp90 gene in a regulatory pathway controlling late steps in sarcomere assembly.

    View details for DOI 10.1242/dev.018150

    View details for Web of Science ID 000253426300016

    View details for PubMedID 18256191

    View details for PubMedCentralID PMC2358948

  • KBP is essential for axonal structure, outgrowth and maintenance in zebrafish, providing insight into the cellular basis of Goldberg-Shprintzen syndrome DEVELOPMENT Lyons, D. A., Naylor, S. G., Mercurio, S., Dominguez, C., Talbot, W. S. 2008; 135 (3): 599-608

    Abstract

    Mutations in Kif1-binding protein/KIAA1279 (KBP) cause the devastating neurological disorder Goldberg-Shprintzen syndrome (GSS) in humans. The cellular function of KBP and the basis of the symptoms of GSS, however, remain unclear. Here, we report the identification and characterization of a zebrafish kbp mutant. We show that kbp is required for axonal outgrowth and maintenance. In vivo time-lapse analysis of neuronal development shows that the speed of early axonal outgrowth is reduced in both the peripheral and central nervous systems in kbp mutants. Ultrastructural studies reveal that kbp mutants have disruption to axonal microtubules during outgrowth. These results together suggest that kbp is an important regulator of the microtubule dynamics that drive the forward propulsion of axons. At later stages, we observe that many affected axons degenerate. Ultrastructural analyses at these stages demonstrate mislocalization of axonal mitochondria and a reduction in axonal number in the peripheral, central and enteric nervous systems. We propose that kbp is an important regulator of axonal development and that axonal cytoskeletal defects underlie the nervous system defects in GSS.

    View details for DOI 10.1242/dev.012377

    View details for Web of Science ID 000252254900019

    View details for PubMedID 18192286

  • Laminin alpha 5 is essential for the formation of the zebrafish fins DEVELOPMENTAL BIOLOGY Webb, A. E., Sanderford, J., Frank, D., Talbot, W. S., Driever, W., Kimelman, D. 2007; 311 (2): 369-382

    Abstract

    The vertebrate fin fold, the presumptive evolutionary antecedent of the paired fins, consists of two layers of epidermal cells extending dorsally and ventrally over the trunk and tail of the embryo, facilitating swimming during the embryonic and larval stages. Development of the fin fold requires dramatic changes in cell shape and adhesion during early development, but the proteins involved in this process are completely unknown. In a screen of mutants defective in fin fold morphogenesis, we identified a mutant with a severe fin fold defect, which also displays malformed pectoral fins. We find that the cause of the defect is a non-sense mutation in the zebrafish lama5 gene that truncates laminin alpha5 before the C-terminal laminin LG domains, thereby preventing laminin alpha5 from interacting with its cell surface receptors. Laminin is mislocalized in this mutant, as are the membrane-associated proteins, actin and beta-catenin, that normally form foci within the fin fold. Ultrastructural analysis revealed severe morphological abnormalities and defects in cell-cell adhesion within the epidermis of the developing fin fold at 36 hpf, resulting in an epidermal sheet that can not extend away from the body. Examining the pectoral fins, we find that the lama5 mutant is the first zebrafish mutant identified in which the pectoral fins fail to make the transition from an apical epidermal ridge to an apical fold, a transformation that is essential for pectoral fin morphogenesis. We propose that laminin alpha5, which is concentrated at the distal ends of the fins, organizes the distal cells of the fin fold and pectoral fins in order to promote the morphogenesis of the epidermis. The lama5 mutant provides novel insight into the role of laminins in the zebrafish epidermis, and the molecular mechanisms driving fin formation in vertebrates.

    View details for DOI 10.1016/j.ydbio.2007.08.034

    View details for Web of Science ID 000251123900009

    View details for PubMedID 17919534

  • Zebrafish bmp4 functions during late gastrulation to specify ventroposterior cell fates DEVELOPMENTAL BIOLOGY Stickney, H. L., Imai, Y., Draper, B., Moens, C., Talbot, W. S. 2007; 310 (1): 71-84

    Abstract

    Bone morphogenetic proteins (BMPs) are key mediators of dorsoventral patterning in vertebrates and are required for the induction of ventral fates in fish and frogs. A widely accepted model of dorsoventral patterning postulates that a morphogenetic BMP activity gradient patterns cell fates along the dorsoventral axis. Recent work in zebrafish suggests that the role of BMP signaling changes over time, with BMPs required for global dorsoventral patterning during early gastrulation and for tail patterning during late gastrulation and early somitogenesis. Key questions remain about the late phase, including which BMP ligands are required and how the functions of BMPs differ during the early and late gastrula stages. In a screen for dominant enhancers of mutations in the homeobox genes vox and vent, which function in parallel to bmp signaling, we identified an insertion mutation in bmp4. We then performed a reverse genetic screen to isolate a null allele of bmp4. We report the characterization of these two alleles and demonstrate that BMP4 is required during the later phase of BMP signaling for the specification of ventroposterior cell fates. Our results indicate that different bmp genes are essential at different stages. In addition, we present genetic evidence supporting a role for a morphogenetic BMP gradient in establishing mesodermal fates during the later phase of BMP signaling.

    View details for DOI 10.1016/j.ydbio.2007.07.027

    View details for Web of Science ID 000250080000007

    View details for PubMedID 17727832

    View details for PubMedCentralID PMC2683675

  • Signals on the move: chemokine receptors and organogenesis in zebrafish. Science's STKE : signal transduction knowledge environment Perlin, J. R., Talbot, W. S. 2007; 2007 (400): pe45-?

    Abstract

    The chemokine SDF1 (stromal cell-derived factor 1) directs cell migration in many different contexts, ranging from embryogenesis to inflammation. SDF1a is the guidance cue for the zebrafish lateral line primordium, a tissue that moves along the flank of the embryo and deposits cells that form mechanosensory organs. The SDF1a receptor CXCR4b acts in cells at the leading edge of the primordium to direct its migration. Two new studies show that a second SDF1 receptor, CXCR7, is required only in the trailing cells of the primordium, and they explore how these two receptors orchestrate migration of the primordium. CXCR4b and CXCR7 are expressed in complementary domains, possibly through mutual repression in which each receptor inhibits expression of the other. These studies illustrate how the entire primordium can respond to a single signal, yet generate cell type-specific responses by using different receptors.

    View details for PubMedID 17712137

  • Putting the glue in glia: Necls mediate Schwann cell-axon adhesion JOURNAL OF CELL BIOLOGY Perlin, J. R., Talbot, W. S. 2007; 178 (5): 721-723

    Abstract

    Interactions between Schwann cells and axons are critical for the development and function of myelinated axons. Two recent studies (see Maurel et al. on p. 861 of this issue; Spiegel et al., 2007) report that the nectin-like (Necl) proteins Necl-1 and -4 are internodal adhesion molecules that are critical for myelination. These studies suggest that Necl proteins mediate a specific interaction between Schwann cells and axons that allows proper communication of the signals that trigger myelination.

    View details for DOI 10.1083/jcb.200708019

    View details for Web of Science ID 000249240800002

    View details for PubMedID 17724116

    View details for PubMedCentralID PMC2064536

  • alpha II-spectrin is essential for assembly of the nodes of Ranvier in myelinated axons CURRENT BIOLOGY Voas, M. G., Lyons, D. A., Naylor, S. G., Arana, N., Rasband, M. N., Talbot, W. S. 2007; 17 (6): 562-568

    Abstract

    Saltatory conduction in myelinated axons requires organization of the nodes of Ranvier, where voltage-gated sodium channels are prominently localized [1]. Previous results indicate that alphaII-spectrin, a component of the cortical cytoskeleton [2], is enriched at the paranodes [3, 4], which flank the node of Ranvier, but alphaII-spectrin's function has not been investigated. Starting with a genetic screen in zebrafish, we discovered in alphaII-spectrin (alphaII-spn) a mutation that disrupts nodal sodium-channel clusters in myelinated axons of the PNS and CNS. In alphaII-spn mutants, the nodal sodium-channel clusters are reduced in number and disrupted at early stages. Analysis of chimeric animals indicated that alphaII-spn functions autonomously in neurons. Ultrastructural studies show that myelin forms in the posterior lateral line nerve and in the ventral spinal cord in alphaII-spn mutants and that the node is abnormally long; these findings indicate that alphaII-spn is required for the assembly of a mature node of the correct length. We find that alphaII-spectrin is enriched in nodes and paranodes at early stages and that the nodal expression diminishes as nodes mature. Our results provide functional evidence that alphaII-spectrin in the axonal cytoskeleton is essential for stabilizing nascent sodium-channel clusters and assembling the mature node of Ranvier.

    View details for DOI 10.1016/j.cub.2007.01.071

    View details for Web of Science ID 000245225500031

    View details for PubMedID 17331725

  • A genetic screen identifies genes essential for development of myelinated axons in zebrafish DEVELOPMENTAL BIOLOGY Pogoda, H., Sternheim, N., Lyons, D. A., Diamond, B., Hawkins, T. A., Woods, I. G., Bhatt, D. H., Franzini-Armstrong, C., Dominguez, C., Arana, N., Jacobs, J., Nix, R., Fetcho, J. R., Talbot, W. S. 2006; 298 (1): 118-131

    Abstract

    The myelin sheath insulates axons in the vertebrate nervous system, allowing rapid propagation of action potentials via saltatory conduction. Specialized glial cells, termed Schwann cells in the PNS and oligodendrocytes in the CNS, wrap axons to form myelin, a compacted, multilayered sheath comprising specific proteins and lipids. Disruption of myelinated axons causes human diseases, including multiple sclerosis and Charcot-Marie-Tooth peripheral neuropathies. Despite the progress in identifying human disease genes and other mutations disrupting glial development and myelination, many important unanswered questions remain about the mechanisms that coordinate the development of myelinated axons. To address these questions, we began a genetic dissection of myelination in zebrafish. Here we report a genetic screen that identified 13 mutations, which define 10 genes, disrupting the development of myelinated axons. We present the initial characterization of seven of these mutations, defining six different genes, along with additional characterization of mutations that we have described previously. The different mutations affect the PNS, the CNS, or both, and phenotypic analyses indicate that the genes affect a wide range of steps in glial development, from fate specification through terminal differentiation. The analysis of these mutations will advance our understanding of myelination, and the mutants will serve as models of human diseases of myelin.

    View details for DOI 10.1016/j.ydbio.2006.06.021

    View details for Web of Science ID 000241071100011

    View details for PubMedID 16875686

  • nsf is essential for organization of myelinated axons in zebrafish CURRENT BIOLOGY Woods, I. G., Lyons, D. A., Voas, M. G., Pogoda, H. M., Talbot, W. S. 2006; 16 (7): 636-648

    Abstract

    Myelinated axons are essential for rapid conduction of action potentials in the vertebrate nervous system. Of particular importance are the nodes of Ranvier, sites of voltage-gated sodium channel clustering that allow action potentials to be propagated along myelinated axons by saltatory conduction. Despite their critical role in the function of myelinated axons, little is known about the mechanisms that organize the nodes of Ranvier.Starting with a forward genetic screen in zebrafish, we have identified an essential requirement for nsf (N-ethylmaleimide sensitive factor) in the organization of myelinated axons. Previous work has shown that NSF is essential for membrane fusion in eukaryotes and has a critical role in vesicle fusion at chemical synapses. Zebrafish nsf mutants are paralyzed and have impaired response to light, reflecting disrupted nsf function in synaptic transmission and neural activity. In addition, nsf mutants exhibit defects in Myelin basic protein expression and in localization of sodium channel proteins at nodes of Ranvier. Analysis of chimeric larvae indicates that nsf functions autonomously in neurons, such that sodium channel clusters are evident in wild-type neurons transplanted into the nsf mutant hosts. Through pharmacological analyses, we show that neural activity and function of chemical synapses are not required for sodium channel clustering and myelination in the larval nervous system.Zebrafish nsf mutants provide a novel vertebrate system to investigate Nsf function in vivo. Our results reveal a previously unknown role for nsf, independent of its function in synaptic vesicle fusion, in the formation of the nodes of Ranvier in the vertebrate nervous system.

    View details for DOI 10.1016/j.cub.2006.02.067

    View details for Web of Science ID 000236649400021

    View details for PubMedID 16581508

  • Essential and opposing roles of zebrafish beta-catenins in the formation of dorsal axial structures and neurectoderm DEVELOPMENT Bellipanni, G., Varga, M. T., Maegawa, S., Imai, Y., Kelly, C., Myers, A. P., Chu, F., Talbot, W. S., Weinberg, E. S. 2006; 133 (7): 1299-1309

    Abstract

    In Xenopus, Wnt signals and their transcriptional effector beta-catenin are required for the development of dorsal axial structures. In zebrafish, previous loss-of-function studies have not identified an essential role for beta-catenin in dorsal axis formation, but the maternal-effect mutation ichabod disrupts beta-catenin accumulation in dorsal nuclei and leads to a reduction of dorsoanterior derivatives. We have identified and characterized a second zebrafish beta-catenin gene, beta-catenin-2, located on a different linkage group from the previously studied beta-catenin-1, but situated close to the ichabod mutation on LG19. Although the ichabod mutation does not functionally alter the beta-catenin-2 reading frame, the level of maternal beta-catenin-2, but not beta-catenin-1, transcript is substantially lower in ichabod, compared with wild-type, embryos. Reduction of beta-catenin-2 function in wild-type embryos by injection of morpholino antisense oligonucleotides (MOs) specific for this gene (MO2) results in the same ventralized phenotypes as seen in ichabod embryos, and administration of MO2 to ichabod embryos increases the extent of ventralization. MOs directed against beta-catenin-1 (MO1), by contrast, had no ventralizing effect on wild-type embryos. beta-catenin-2 is thus specifically required for organizer formation and this function is apparently required maternally, because the ichabod mutation causes a reduction in maternal transcription of the gene and a reduced level of beta-catenin-2 protein in the early embryo. A redundant role of beta-catenins in suppressing formation of neurectoderm is revealed when both beta-catenin genes are inhibited. Using a combination of MO1 and MO2 in wild-type embryos, or by injecting solely MO1 in ichabod embryos, we obtain expression of a wide spectrum of neural markers in apparently appropriate anteroposterior pattern. We propose that the early, dorsal-promoting function of beta-catenin-2 is essential to counteract a later, dorsal- and neurectoderm-repressing function that is shared by both beta-catenin genes.

    View details for DOI 10.1242/dev.02295

    View details for Web of Science ID 000236764100009

    View details for PubMedID 16510506

  • lessen encodes a zebrafish trap100 required for enteric nervous system development DEVELOPMENT Pietsch, J., Delalande, J. M., Jakaitis, B., Stensby, J. D., Dohle, S., Talbot, W. S., Raible, D. W., Shepherd, I. T. 2006; 133 (3): 395-406

    Abstract

    The zebrafish enteric nervous system (ENS), like those of all other vertebrate species, is principally derived from the vagal neural crest. The developmental controls that govern the specification and patterning of the ENS are not well understood. To identify genes required for the formation of the vertebrate ENS, we preformed a genetic screen in zebrafish. We isolated the lessen (lsn) mutation that has a significant reduction in the number of ENS neurons as well as defects in other cranial neural crest derived structures. We show that the lsn gene encodes a zebrafish orthologue of Trap100, one of the subunits of the TRAP/mediator transcriptional regulation complex. A point mutation in trap100 causes a premature stop codon that truncates the protein, causing a loss of function. Antisense-mediated knockdown of trap100 causes an identical phenotype to lsn. During development trap100 is expressed in a dynamic tissue-specific expression pattern consistent with its function in ENS and jaw cartilage development. Analysis of neural crest markers revealed that the initial specification and migration of the neural crest is unaffected in lsn mutants. Phosphohistone H3 immunocytochemistry revealed that there is a significant reduction in proliferation of ENS precursors in lsn mutants. Using cell transplantation studies, we demonstrate that lsn/trap100 acts cell autonomously in the pharyngeal mesendoderm and influences the development of neural crest derived cartilages secondarily. Furthermore, we show that endoderm is essential for ENS development. These studies demonstrate that lsn/trap100 is not required for initial steps of cranial neural crest development and migration, but is essential for later proliferation of ENS precursors in the intestine.

    View details for DOI 10.1242/dev.02215

    View details for Web of Science ID 000235910500002

    View details for PubMedID 16396911

    View details for PubMedCentralID PMC2651469

  • The zebrafish gene map defines ancestral vertebrate chromosomes GENOME RESEARCH Woods, I. G., Wilson, C., Friedlander, B., Chang, P., Reyes, D. K., Nix, R., Kelly, P. D., Chu, F., Postlethwait, J. H., Talbot, W. S. 2005; 15 (9): 1307-1314

    Abstract

    Genetic screens in zebrafish (Danio rerio) have identified mutations that define the roles of hundreds of essential vertebrate genes. Genetic maps can link mutant phenotype with gene sequence by providing candidate genes for mutations and polymorphic genetic markers useful in positional cloning projects. Here we report a zebrafish genetic map comprising 4073 polymorphic markers, with more than twice the number of coding sequences localized in previously reported zebrafish genetic maps. We use this map in comparative studies to identify numerous regions of synteny conserved among the genomes of zebrafish, Tetraodon, and human. In addition, we use our map to analyze gene duplication in the zebrafish and Tetraodon genomes. Current evidence suggests that a whole-genome duplication occurred in the teleost lineage after it split from the tetrapod lineage, and that only a subset of the duplicates have been retained in modern teleost genomes. It has been proposed that differential retention of duplicate genes may have facilitated the isolation of nascent species formed during the vast radiation of teleosts. We find that different duplicated genes have been retained in zebrafish and Tetraodon, although similar numbers of duplicates remain in both genomes. Finally, we use comparative mapping data to address the proposal that the common ancestor of vertebrates had a genome consisting of 12 chromosomes. In a three-way comparison between the genomes of zebrafish, Tetraodon, and human, our analysis delineates the gene content for 11 of these 12 proposed ancestral chromosomes.

    View details for DOI 10.1101/gr.4134305

    View details for PubMedID 16109975

  • erbb3 and erbb2 are essential for Schwann cell migration and myelination in zebrafish CURRENT BIOLOGY Lyons, D. A., Pogoda, H. M., Voas, M. G., Woods, I. G., Diamond, B., Nix, R., Arana, N., Jacobs, J., Talbot, W. S. 2005; 15 (6): 513-524

    Abstract

    Myelin is critical for efficient axonal conduction in the vertebrate nervous system. Neuregulin (Nrg) ligands and their ErbB receptors are required for the development of Schwann cells, the glial cells that form myelin in the peripheral nervous system. Previous studies have not determined whether Nrg-ErbB signaling is essential in vivo for Schwann cell fate specification, proliferation, survival, migration, or the onset of myelination.In genetic screens for mutants with disruptions in myelinated nerves, we identified mutations in erbb3 and erbb2, which together encode a heteromeric tyrosine kinase receptor for Neuregulin ligands. Phenotypic analysis shows that both genes are essential for development of Schwann cells. BrdU-incorporation studies and time-lapse analysis reveal that Schwann cell proliferation and migration, but not survival, are disrupted in erbb3 mutants. We show that Schwann cells can migrate in the absence of DNA replication. This uncoupling of proliferation and migration indicates that erbb gene function is required independently for these two processes. Pharmacological inhibition of ErbB signaling at different stages reveals a continuing requirement for ErbB function during migration and also provides evidence that ErbB signaling is required after migration for proliferation and the terminal differentiation of myelinating Schwann cells.These results provide in vivo evidence that Neuregulin-ErbB signaling is essential for directed Schwann cell migration and demonstrate that this pathway is also required for the onset of myelination in postmigratory Schwann cells.

    View details for DOI 10.1016/j.cub.2005.02.030

    View details for PubMedID 15797019

  • The you gene encodes an EGF-CUB protein essential for hedgehog signaling in zebrafish PLOS BIOLOGY Woods, I. G., Talbot, W. S. 2005; 3 (3): 476-487

    Abstract

    Hedgehog signaling is required for many aspects of development in vertebrates and invertebrates. Misregulation of the Hedgehog pathway causes developmental abnormalities and has been implicated in certain types of cancer. Large-scale genetic screens in zebrafish have identified a group of mutations, termed you-class mutations, that share common defects in somite shape and in most cases disrupt Hedgehog signaling. These mutant embryos exhibit U-shaped somites characteristic of defects in slow muscle development. In addition, Hedgehog pathway mutations disrupt spinal cord patterning. We report the positional cloning of you, one of the original you-class mutations, and show that it is required for Hedgehog signaling in the development of slow muscle and in the specification of ventral fates in the spinal cord. The you gene encodes a novel protein with conserved EGF and CUB domains and a secretory pathway signal sequence. Epistasis experiments support an extracellular role for You upstream of the Hedgehog response mechanism. Analysis of chimeras indicates that you mutant cells can appropriately respond to Hedgehog signaling in a wild-type environment. Additional chimera analysis indicates that wild-type you gene function is not required in axial Hedgehog-producing cells, suggesting that You is essential for transport or stability of Hedgehog signals in the extracellular environment. Our positional cloning and functional studies demonstrate that You is a novel extracellular component of the Hedgehog pathway in vertebrates.

    View details for DOI 10.1371/journal.pbio.0030066

    View details for Web of Science ID 000227984000015

    View details for PubMedID 15660164

    View details for PubMedCentralID PMC544551

  • Monorail/Foxa2 regulates floorplate differentiation and specification of oligodendrocytes, serotonergic raphe neurones and cranial motoneurones DEVELOPMENT Norton, W. H., Mangoli, M., Lele, Z., Pogoda, H. M., Diamond, B., Mercurio, S., Russell, C., Teraoka, H., Stickney, H. L., Rauch, G. J., HEISENBERG, C. P., Houart, C., Schilling, T. F., Frohnhoefer, H. G., Rastegar, S., Neumann, C. J., Gardiner, R. M., Strahle, U., Geisler, R., Rees, M., Talbot, W. S., Wilson, S. W. 2005; 132 (4): 645-658

    Abstract

    In this study, we elucidate the roles of the winged-helix transcription factor Foxa2 in ventral CNS development in zebrafish. Through cloning of monorail (mol), which we find encodes the transcription factor Foxa2, and phenotypic analysis of mol-/- embryos, we show that floorplate is induced in the absence of Foxa2 function but fails to further differentiate. In mol-/- mutants, expression of Foxa and Hh family genes is not maintained in floorplate cells and lateral expansion of the floorplate fails to occur. Our results suggest that this is due to defects both in the regulation of Hh activity in medial floorplate cells as well as cell-autonomous requirements for Foxa2 in the prospective laterally positioned floorplate cells themselves. Foxa2 is also required for induction and/or patterning of several distinct cell types in the ventral CNS. Serotonergic neurones of the raphenucleus and the trochlear motor nucleus are absent in mol-/- embryos, and oculomotor and facial motoneurones ectopically occupy ventral CNS midline positions in the midbrain and hindbrain. There is also a severe reduction of prospective oligodendrocytes in the midbrain and hindbrain. Finally, in the absence of Foxa2, at least two likely Hh pathway target genes are ectopically expressed in more dorsal regions of the midbrain and hindbrain ventricular neuroepithelium, raising the possibility that Foxa2 activity may normally be required to limit the range of action of secreted Hh proteins.

    View details for DOI 10.1242/dev.01611

    View details for Web of Science ID 000227427100003

    View details for PubMedID 15677724

    View details for PubMedCentralID PMC2790417

  • Molecular genetics of axis formation in zebrafish ANNUAL REVIEW OF GENETICS Schier, A. F., Talbot, W. S. 2005; 39: 561-613

    Abstract

    The basic vertebrate body plan of the zebrafish embryo is established in the first 10 hours of development. This period is characterized by the formation of the anterior-posterior and dorsal-ventral axes, the development of the three germ layers, the specification of organ progenitors, and the complex morphogenetic movements of cells. During the past 10 years a combination of genetic, embryological, and molecular analyses has provided detailed insights into the mechanisms underlying this process. Maternal determinants control the expression of transcription factors and the location of signaling centers that pattern the blastula and gastrula. Bmp, Nodal, FGF, canonical Wnt, and retinoic acid signals generate positional information that leads to the restricted expression of transcription factors that control cell type specification. Noncanonical Wnt signaling is required for the morphogenetic movements during gastrulation. We review how the coordinated interplay of these molecules determines the fate and movement of embryonic cells.

    View details for DOI 10.1146/annurev.genet.37.110801.143752

    View details for Web of Science ID 000234685200024

    View details for PubMedID 16285872

  • Axon sorting in the optic tract requires HSPG synthesis by ext2 (dackel) and extl3 (boxer) NEURON Lee, J. S., von der Hardt, S., Rusch, M. A., Stringer, S. E., Stickney, H. L., Talbot, W. S., Geisler, R., Nusslein-Volhard, C., Selleck, S. B., Chien, C. B., Roehl, H. 2004; 44 (6): 947-960

    Abstract

    Retinal ganglion cell (RGC) axons are topographically ordered in the optic tract according to their retinal origin. In zebrafish dackel (dak) and boxer (box) mutants, some dorsal RGC axons missort in the optic tract but innervate the tectum topographically. Molecular cloning reveals that dak and box encode ext2 and extl3, glycosyltransferases implicated in heparan sulfate (HS) biosynthesis. Both genes are required for HS synthesis, as shown by biochemical and immunohistochemical analysis, and are expressed maternally and then ubiquitously, likely playing permissive roles. Missorting in box can be rescued by overexpression of extl3. dak;box double mutants show synthetic pathfinding phenotypes that phenocopy robo2 mutants, suggesting that Robo2 function requires HS in vivo; however, tract sorting does not require Robo function, since it is normal in robo2 null mutants. This genetic evidence that heparan sulfate proteoglycan function is required for optic tract sorting provides clues to begin understanding the underlying molecular mechanisms.

    View details for Web of Science ID 000225842300009

    View details for PubMedID 15603738

  • The role of the zebrafish nodal-related genes squint and cyclops in patterning of mesendoderm DEVELOPMENT Dougan, S. T., Warga, R. M., Kane, D. A., Schier, A. F., Talbot, W. S. 2003; 130 (9): 1837-1851

    Abstract

    Nodal signals, a subclass of the TGFbeta superfamily of secreted factors, induce formation of mesoderm and endoderm in vertebrate embryos. We have examined the possible dorsoventral and animal-vegetal patterning roles for Nodal signals by using mutations in two zebrafish nodal-related genes, squint and cyclops, to manipulate genetically the levels and timing of Nodal activity. squint mutants lack dorsal mesendodermal gene expression at the late blastula stage, and fate mapping and gene expression studies in sqt(-/-); cyc(+/+) and sqt(-/-); cyc(+/-) mutants show that some dorsal marginal cells inappropriately form hindbrain and spinal cord instead of dorsal mesendodermal derivatives. The effects on ventrolateral mesendoderm are less severe, although the endoderm is reduced and muscle precursors are located nearer to the margin than in wild type. Our results support a role for Nodal signals in patterning the mesendoderm along the animal-vegetal axis and indicate that dorsal and ventrolateral mesoderm require different levels of squint and cyclops function. Dorsal marginal cells were not transformed toward more lateral fates in either sqt(-/-); cyc(+/-) or sqt(-/-); cyc(+/+) embryos, arguing against a role for the graded action of Nodal signals in dorsoventral patterning of the mesendoderm. Differential regulation of the cyclops gene in these cells contributes to the different requirements for nodal-related gene function in these cells. Dorsal expression of cyclops requires Nodal-dependent autoregulation, whereas other factors induce cyclops expression in ventrolateral cells. In addition, the differential timing of dorsal mesendoderm induction in squint and cyclops mutants suggests that dorsal marginal cells can respond to Nodal signals at stages ranging from the mid-blastula through the mid-gastrula.

    View details for DOI 10.1242/dev.00400

    View details for Web of Science ID 000182811600011

    View details for PubMedID 12642489

  • Genetic analysis of zebrafish gli1 and gli2 reveals divergent requirements for gli genes in vertebrate development DEVELOPMENT Karlstrom, R. O., Tyurina, O. V., Kawakami, A., Nishioka, N., Talbot, W. S., SASAKI, H., Schier, A. F. 2003; 130 (8): 1549-1564

    Abstract

    Gli proteins regulate the transcription of Hedgehog (Hh) target genes. Genetic studies in mouse have shown that Gli1 is not essential for embryogenesis, whereas Gli2 acts as an activator of Hh target genes. In contrast, misexpression studies in Xenopus and cultured cells have suggested that Gli1 can act as an activator of Hh-regulated genes, whereas Gli2 might function as a repressor of a subset of Hh targets. To clarify the roles of gli genes during vertebrate development, we have analyzed the requirements for gli1 and gli2 during zebrafish embryogenesis. We report that detour (dtr) mutations encode loss-of-function alleles of gli1. In contrast to mouse Gli1 mutants, dtr mutants and embryos injected with gli1 antisense morpholino oligonucleotides display defects in the activation of Hh target genes in the ventral neuroectoderm. Mutations in you-too (yot) encode C-terminally truncated Gli2. We find that these truncated proteins act as dominant repressors of Hh signaling, in part by blocking Gli1 function. In contrast, blocking Gli2 function by eliminating full-length Gli2 results in minor Hh signaling defects and uncovers a repressor function of Gli2 in the telencephalon. In addition, we find that Gli1 and Gli2 have activator functions during somite and neural development. These results reveal divergent requirements for Gli1 and Gli2 in mouse and zebrafish and indicate that zebrafish Gli1 is an activator of Hh-regulated genes, while zebrafish Gli2 has minor roles as a repressor or activator of Hh targets.

    View details for DOI 10.1242/dev.00364

    View details for Web of Science ID 000182592500007

    View details for PubMedID 12620981

  • Zebrafish comparative genomics and the origins of vertebrate chromosomes GENOME RESEARCH Postlethwait, J. H., Woods, I. G., Ngo-Hazelett, P., Yan, Y. L., Kelly, P. D., Chu, F., Huang, H., Hill-Force, A., Talbot, W. S. 2000; 10 (12): 1890-1902

    Abstract

    To help understand mechanisms of vertebrate genome evolution, we have compared zebrafish and tetrapod gene maps. It has been suggested that translocations are fixed more frequently than inversions in mammals. Gene maps showed that blocks of conserved syntenies between zebrafish and humans were large, but gene orders were frequently inverted and transposed. This shows that intrachromosomal rearrangements have been fixed more frequently than translocations. Duplicated chromosome segments suggest that a genome duplication occurred in ray-fin phylogeny, and comparative studies suggest that this event happened deep in the ancestry of teleost fish. Consideration of duplicate chromosome segments shows that at least 20% of duplicated gene pairs may be retained from this event. Despite genome duplication, zebrafish and humans have about the same number of chromosomes, and zebrafish chromosomes are mosaically orthologous to several human chromosomes. Is this because of an excess of chromosome fissions in the human lineage or an excess of chromosome fusions in the zebrafish lineage? Comparative analysis suggests that an excess of chromosome fissions in the tetrapod lineage may account for chromosome numbers and provides histories for several human chromosomes.

    View details for Web of Science ID 000165833900007

    View details for PubMedID 11116085

  • fast1 is required for the development of dorsal axial structures in zebrafish CURRENT BIOLOGY Sirotkin, H. I., GATES, M. A., Kelly, P. D., Schier, A. F., Talbot, W. S. 2000; 10 (17): 1051-1054

    Abstract

    Nodal-related signals comprise a subclass of the transforming growth factor-beta (TGF-beta) superfamily and regulate key events in vertebrate embryogenesis, including mesoderm formation, establishment of left-right asymmetry and neural patterning [1-8]. Nodal ligands are thought to act with EGF-CFC protein co-factors to activate activin type I and II or related receptors, which phosphorylate Smad2 and trigger nuclear translocation of a Smad2/4 complex [8-12]. The winged-helix transcription factor forkhead activin signal transducer-1 (Fast-1) acts as a co-factor for Smad2 [12-20]. Xenopus Fast-1 is thought to function as a transcriptional effector of Nodal signals during mesoderm formation [17], but no mutations in the Fast-1 gene have been identified. We report the identification of the zebrafish fast1 gene and show that it is disrupted in schmalspur (sur) mutants, which have defects in the development of dorsal midline cell types and establishment of left-right asymmetry [21-25]. We find that prechordal plate and notochord are strongly reduced in maternal-zygotic sur mutants, whereas other mesendodermal structures are present - a less severe phenotype than that caused by complete loss of Nodal signaling. These results show that fast1 is required for development of dorsal axial structures and left-right asymmetry, and suggest that Nodal signals act through Fast1-dependent and independent pathways.

    View details for PubMedID 10996072

  • Bozozok and squint act in parallel to specify dorsal mesoderm and anterior neuroectoderm in zebrafish DEVELOPMENT Sirotkin, H. I., Dougan, S. T., Schier, A. F., Talbot, W. S. 2000; 127 (12): 2583-2592

    Abstract

    In vertebrate embryos, maternal (beta)-catenin protein activates the expression of zygotic genes that establish the dorsal axial structures. Among the zygotically acting genes with key roles in the specification of dorsal axial structures are the homeobox gene bozozok (boz) and the nodal-related (TGF-(beta) family) gene squint (sqt). Both genes are expressed in the dorsal yolk syncytial layer, a source of dorsal mesoderm inducing signals, and mutational analysis has indicated that boz and sqt are required for dorsal mesoderm development. Here we examine the regulatory interactions among boz, sqt and a second nodal-related gene, cyclops (cyc). Three lines of evidence indicate that boz and sqt act in parallel to specify dorsal mesoderm and anterior neuroectoderm. First, boz requires sqt function to induce high levels of ectopic dorsal mesoderm, consistent with sqt acting either downstream or in parallel to boz. Second, sqt mRNA is expressed in blastula stage boz mutants, indicating that boz is not essential for activation of sqt transcription, and conversely, boz mRNA is expressed in blastula stage sqt mutants. Third, boz;sqt double mutants have a much more severe phenotype than boz and sqt single mutants. Double mutants consistently lack the anterior neural tube and axial mesoderm, and ventral fates are markedly expanded. Expression of chordin and noggin1 is greatly reduced in boz;sqt mutants, indicating that the boz and sqt pathways have overlapping roles in activating secreted BMP antagonists. In striking contrast to boz;sqt double mutants, anterior neural fates are specified in boz;sqt;cyc triple mutants. This indicates that cyc represses anterior neural development, and that boz and sqt counteract this repressive function. Our results support a model in which boz and sqt act in parallel to induce dorsalizing BMP-antagonists and to counteract the repressive function of cyc in neural patterning.

    View details for Web of Science ID 000087948900007

    View details for PubMedID 10821757

  • The role of one-eyed pinhead and nodal signaling in left-right axis determination Burdine, R. D., Gritsman, K., Corrales, J., Talbot, W. S., Schier, A. F. ACADEMIC PRESS INC. 2000: 262
  • Analysis of chromosomal rearrangements induced by postmeiotic mutagenesis with ethylnitrosourea in zebrafish GENETICS Imai, Y., Feldman, B., Schier, A. F., Talbot, W. S. 2000; 155 (1): 261-272

    Abstract

    Mutations identified in zebrafish genetic screens allow the dissection of a wide array of problems in vertebrate biology. Most screens have examined mutations induced by treatment of spermatogonial (premeiotic) cells with the chemical mutagen N-ethyl-N-nitrosourea (ENU). Treatment of postmeiotic gametes with ENU induces specific-locus mutations at a higher rate than premeiotic regimens, suggesting that postmeiotic mutagenesis protocols could be useful in some screening strategies. Whereas there is extensive evidence that ENU induces point mutations in premeiotic cells, the range of mutations induced in postmeiotic zebrafish germ cells has been less thoroughly characterized. Here we report the identification and analysis of five mutations induced by postmeiotic ENU treatment. One mutation, snh(st1), is a translocation involving linkage group (LG) 11 and LG 14. The other four mutations, oep(st2), kny(st3), Df(LG 13)(st4), and cyc(st5), are deletions, ranging in size from less than 3 cM to greater than 20 cM. These results show that germ cell stage is an important determinant of the type of mutations induced. The induction of chromosomal rearrangements may account for the elevated frequency of specific-locus mutations observed after treatment of postmeiotic gametes with ENU.

    View details for Web of Science ID 000086869200022

    View details for PubMedID 10790400

  • Genetic linkage mapping of zebrafish genes and ESTs GENOME RESEARCH Kelly, P. D., Chu, F., Woods, I. G., Ngo-Hazelett, P., Cardozo, T., Huang, H., Kimm, F., Liao, L. Y., Yan, Y. L., Zhou, Y. Y., Johnson, S. L., Abagyan, R., Schier, A. F., Postlethwait, J. H., Talbot, W. S. 2000; 10 (4): 558-567

    Abstract

    Genetic screens in zebrafish (Danio rerio) have isolated mutations in hundreds of genes essential for vertebrate development, physiology, and behavior. We have constructed a genetic linkage map that will facilitate the identification of candidate genes for these mutations and allow comparisons among the genomes of zebrafish and other vertebrates. On this map, we have localized 771 zebrafish genes and expressed sequence tags (ESTs) by scoring single-stranded conformational polymorphisms (SSCPs) in a meiotic mapping panel. Of these sequences, 642 represent previously unmapped genes and ESTs. The mapping panel was comprised of 42 homozygous diploid individuals produced by heat shock treatment of haploid embryos at the one-cell stage (HS diploids). This "doubled haploid" strategy combines the advantages of mapping in haploid and standard diploid systems, because heat shock diploid individuals have only one allele at each locus and can survive to adulthood, enabling a relatively large quantity of genomic DNA to be prepared from each individual in the mapping panel. To integrate this map with others, we also scored 593 previously mapped simple-sequence length polymorphisms (SSLPs) in the mapping panel. This map will accelerate the molecular analysis of zebrafish mutations and facilitate comparative analysis of vertebrate genomes.

    View details for PubMedID 10779498

  • Nodal signaling patterns the organizer DEVELOPMENT Gritsman, K., Talbot, W. S., Schier, A. F. 2000; 127 (5): 921-932

    Abstract

    Spemann's organizer plays an essential role in patterning the vertebrate embryo. During gastrulation, organizer cells involute and form the prechordal plate anteriorly and the notochord more posteriorly. The fate mapping and gene expression analyses in zebrafish presented in this study reveal that this anteroposterior polarity is already initiated in the organizer before gastrulation. Prechordal plate progenitors reside close to the blastoderm margin and express the homeobox gene goosecoid, whereas notochord precursors are located further from the margin and express the homeobox gene floating head. The nodal-related genes cyclops and squint are expressed at the blastoderm margin and are required for prechordal plate and notochord formation. We show that differential activation of the Nodal signaling pathway is essential in establishing anteroposterior pattern in the organizer. First, overexpression of cyclops and squint at different doses leads to the induction of floating head at low doses and the induction of both goosecoid and floating head at higher doses. Second, decreasing Nodal signaling using different concentrations of the antagonist Antivin inhibits goosecoid expression at low doses and blocks expression of both goosecoid and floating head at higher doses. Third, attenuation of Nodal signaling in zygotic mutants for the EGF-CFC gene one-eyed pinhead, an essential cofactor for Nodal signaling, leads to the loss of goosecoid expression and expansion of floating head expression in the organizer. Concomitantly, cells normally fated to become prechordal plate are transformed into notochord progenitors. Finally, activation of Nodal signaling at different times suggests that prechordal plate specification requires sustained Nodal signaling, whereas transient signaling is sufficient for notochord development. Together, these results indicate that differential Nodal signaling patterns the organizer before gastrulation, with the highest level of activity required for anterior fates and lower activity essential for posterior fates.

    View details for Web of Science ID 000086086200001

    View details for PubMedID 10662632

  • The EGF-CFC protein one-eyed pinhead is essential for nodal signaling CELL Gritsman, K., Zhang, J. J., Cheng, S., Heckscher, E., Talbot, W. S., Schier, A. F. 1999; 97 (1): 121-132

    Abstract

    The zebrafish EGF-CFC gene one-eyed pinhead (oep) is required zygotically for the formation of the ventral neuroectoderm, endoderm, and prechordal plate. Here we report that embryos lacking both maternal and zygotic Oep activity are defective in germ layer formation, organizer development, and the positioning of the anterior-posterior axis. An identical phenotype is displayed by double mutants for the nodal-related genes squint and cyclops. Mutations in oep eliminate the response to Squint and Cyclops overexpression but are suppressed by expression of Activin and activated forms of the type I receptor ActRIB and Smad2. Expression of the murine EGF-CFC gene cripto rescues oep mutants. These results suggest a conserved role for EGF-CFC proteins as essential extracellular cofactors for Nodal signaling during vertebrate development.

    View details for Web of Science ID 000079843900014

    View details for PubMedID 10199408

  • A genetic linkage map for zebrafish: Comparative analysis and localization of genes and expressed sequences GENOME RESEARCH GATES, M. A., Kim, L., Egan, E. S., Cardozo, T., Sirotkin, H. I., Dougan, S. T., Lashkari, D., Abagyan, R., Schier, A. F., Talbot, W. S. 1999; 9 (4): 334-347

    Abstract

    Genetic screens in zebrafish (Danio rerio) have isolated mutations in hundreds of genes with essential functions. To facilitate the identification of candidate genes for these mutations, we have genetically mapped 104 genes and expressed sequence tags by scoring single-strand conformational polymorphisms in a panel of haploid siblings. To integrate this map with existing genetic maps, we also scored 275 previously mapped genes, microsatellites, and sequence-tagged sites in the same haploid panel. Systematic phylogenetic analysis defined likely mammalian orthologs of mapped zebrafish genes, and comparison of map positions in zebrafish and mammals identified significant conservation of synteny. This comparative analysis also identified pairs of zebrafish genes that appear to be orthologous to single mammalian genes, suggesting that these genes arose in a genome duplication that occurred in the teleost lineage after the divergence of fish and mammal ancestors. This comparative map analysis will be useful in predicting the locations of zebrafish genes from mammalian gene maps and in understanding the evolution of the vertebrate genome.

    View details for Web of Science ID 000079904300003

    View details for PubMedID 10207156

  • The zebrafish bozozok locus encodes Dharma, a homeodomain protein essential for induction of gastrula organizer and dorsoanterior embryonic structures DEVELOPMENT Fekany, K., Yamanaka, Y., Leung, T. C., Sirotkin, H. I., Topczewski, J., GATES, M. A., Hibi, M., Renucci, A., Stemple, D., Radbill, A., Schier, A. F., Driever, W., Hirano, T., Talbot, W. S., Solnica-Krezel, L. 1999; 126 (7): 1427-1438

    Abstract

    The dorsal gastrula organizer plays a fundamental role in establishment of the vertebrate axis. We demonstrate that the zebrafish bozozok (boz) locus is required at the blastula stages for formation of the embryonic shield, the equivalent of the gastrula organizer and expression of multiple organizer-specific genes. Furthermore, boz is essential for specification of dorsoanterior embryonic structures, including notochord, prechordal mesendoderm, floor plate and forebrain. We report that boz mutations disrupt the homeobox gene dharma. Overexpression of boz in the extraembryonic yolk syncytial layer of boz mutant embryos is sufficient for normal development of the overlying blastoderm, revealing an involvement of extraembryonic structures in anterior patterning in fish similarly to murine embryos. Epistatic analyses indicate that boz acts downstream of beta-catenin and upstream to TGF-beta signaling or in a parallel pathway. These studies provide genetic evidence for an essential function of a homeodomain protein in beta-catenin-mediated induction of the dorsal gastrula organizer and place boz at the top of a hierarchy of zygotic genes specifying the dorsal midline of a vertebrate embryo.

    View details for Web of Science ID 000079848200008

    View details for PubMedID 10068636

  • Positional cloning of mutated zebrafish genes METHODS IN CELL BIOLOGY, VOL 60 Talbot, W. S., Schier, A. F. 1999; 60: 259-286

    View details for Web of Science ID 000087260100015

    View details for PubMedID 9891342

  • Zebrafish organizer development and germ-layer formation require nodal-related signals NATURE Feldman, B., GATES, M. A., Egan, E. S., Dougan, S. T., Rennebeck, G., Sirotkin, H. I., Schier, A. F., Talbot, W. S. 1998; 395 (6698): 181-185

    Abstract

    The vertebrate body plan is established during gastrulation, when cells move inwards to form the mesodermal and endodermal germ layers. Signals from a region of dorsal mesoderm, which is termed the organizer, pattern the body axis by specifying the fates of neighbouring cells. The organizer is itself induced by earlier signals. Although members of the transforming growth factor-beta (TGF-beta) and Wnt families have been implicated in the formation of the organizer, no endogenous signalling molecule is known to be required for this process. Here we report that the zebrafish squint (sqt) and cyclops (cyc) genes have essential, although partly redundant, functions in organizer development and also in the formation of mesoderm and endoderm. We show that the sqt gene encodes a member of the TGF-beta superfamily that is related to mouse nodal. cyc encodes another nodal-related proteins, which is consistent with our genetic evidence that sqt and cyc have overlapping functions. The sqt gene is expressed in a dorsal region of the blastula that includes the extraembryonic yolk syncytial layer (YSL). The YSL has been implicated as a source of signals that induce organizer development and mesendoderm formation. Misexpression of sqt RNA within the embryo or specifically in the YSL induces expanded or ectopic dorsal mesoderm. These results establish an essential role for nodal-related signals in organizer development and mesendoderm formation.

    View details for Web of Science ID 000075829900044

    View details for PubMedID 9744277

  • The zebrafish organizer CURRENT OPINION IN GENETICS & DEVELOPMENT Schier, A. F., Talbot, W. S. 1998; 8 (4): 464-471

    Abstract

    Signals from the organizer play a crucial role in patterning the vertebrate embryo. Recent molecular analysis of zebrafish mutations has established an essential role for BMP2 and chordin in organizer function and has identified one-eyed pinhead as a novel EGF-like gene involved in prechordal plate and endoderm formation. In addition, embryological studies have suggested that the zebrafish organizer is induced by extraembryonic cues and have defined two novel organizing centers that pattern the nervous system along the anterior-posterior axes.

    View details for Web of Science ID 000075603800014

    View details for PubMedID 9729724

  • Mutant rescue by BAC clone injection in zebrafish GENOMICS Yan, Y. L., Talbot, W. S., Egan, E. S., Postlethwait, J. H. 1998; 50 (2): 287-289

    Abstract

    Genes essential for vertebrate body plan specification, organ development, and organ function are likely to be shared between mammals and zebrafish, but only in zebrafish have large-scale, genome-wide mutagenesis screens been conducted to isolate embryonic lethal mutations. Discovering the roles played by these disrupted genes requires their molecular characterization, which would be facilitated by assaying large cloned genomic DNAs for their potential to rescue mutant phenotypes. Here we demonstrate that bacterial artificial chromosomes can rescue the phenotype of floating head (flh) mutants. Homozygous flh embryos lack a differentiated notochord and have a reduced, discontinuous floor plate. Mutant embryos injected with genomic clones containing the flh+ gene often had stretches of several to many notochord cells overlaid by a row of floor-plate cells. In contrast, control mutant embryos injected with artificial chromosomes lacking the flh+ gene failed to form notochord. We conclude that the injection of large-insert genomic clones will speed the isolation of zebrafish genes disrupted by mutation and hence the identification of gene functions necessary for development of vertebrate embryos.

    View details for Web of Science ID 000074367700018

    View details for PubMedID 9653657

  • The cloche and spadetail genes differentially affect hematopoiesis and vasculogenesis DEVELOPMENTAL BIOLOGY Thompson, M. A., Ransom, D. G., Pratt, S. J., MacLennan, H., Kieran, M. W., Detrich, H. W., Vail, B., Huber, T. L., Paw, B., Brownlie, A. J., Oates, A. C., Fritz, A., GATES, M. A., Amores, A., Bahary, N., Talbot, W. S., Her, H., Beier, D. R., Postlethwait, J. H., Zon, L. I. 1998; 197 (2): 248-269

    Abstract

    In vertebrates, hematopoietic and vascular progenitors develop from ventral mesoderm. The first primitive wave of hematopoiesis yields embryonic red blood cells, whereas progenitor cells of subsequent definitive waves form all hematopoietic cell lineages. In this report we examine the development of hematopoietic and vasculogenic cells in normal zebrafish and characterize defects in cloche and spadetail mutant embryos. The zebrafish homologs of lmo2, c-myb, fli1, flk1, and flt4 have been cloned and characterized in this study. Expression of these genes identifies embryonic regions that contain hematopoietic and vascular progenitor cells. The expression of c-myb also identifies definitive hematopoietic cells in the ventral wall of the dorsal aorta. Analysis of b316 mutant embryos that carry a deletion of the c-myb gene demonstrates that c-myb is not required for primitive erythropoiesis in zebrafish even though it is expressed in these cells. Both cloche and spadetail mutant embryos have defects in primitive hematopoiesis and definitive hematopoiesis. The cloche mutants also have significant decreases in vascular gene expression, whereas spadetail mutants expressed normal levels of these genes. These studies demonstrate that the molecular mechanisms that regulate hematopoiesis and vasculogenesis have been conserved throughout vertebrate evolution and the clo and spt genes are key regulators of these programs.

    View details for Web of Science ID 000074064000009

    View details for PubMedID 9630750

  • Positional cloning identifies zebrafish one-eyed pinhead as a permissive EGF-related ligand required during gastrulation CELL Zhang, J. J., Talbot, W. S., Schier, A. F. 1998; 92 (2): 241-251

    Abstract

    The zebrafish one-eyed pinhead (oep) mutation disrupts embryonic development, resulting in cyclopia and defects in endoderm, prechordal plate, and ventral neuroectoderm formation. We report the molecular isolation of oep using a positional cloning approach. The oep gene encodes a novel EGF-related protein with similarity to the EGF-CFC proteins cripto, cryptic, and FRL-1. Wild-type oep protein contains a functional signal sequence and is membrane-associated. Following ubiquitous maternal and zygotic expression, highest levels of oep mRNA are found in the gastrula margin and in axial structures and forebrain. Widespread misexpression of both membrane-attached and secreted forms of oep rescues prechordal plate and forebrain development in mutant embryos but does not lead to the ectopic induction of these cell types in wild-type fish. These results establish an essential but permissive role for an EGF-related ligand during vertebrate gastrulation.

    View details for Web of Science ID 000071672600013

    View details for PubMedID 9458048

  • Genetic analysis of chromosomal rearrangements in the cyclops region of the zebrafish genome GENETICS Talbot, W. S., Egan, E. S., GATES, M. A., Walker, C., Ullmann, B., Neuhauss, S. C., Kimmel, C. B., Postlethwait, J. H. 1998; 148 (1): 373-380

    Abstract

    Genetic screens in zebrafish have provided mutations in hundreds of genes with essential functions in the developing embryo. To investigate the possible uses of chromosomal rearrangements in the analysis of these mutations, we genetically characterized three gamma-ray induced alleles of cyclops (cyc), a gene required for development of midline structures. We show that cyc maps near one end of Linkage Group 12 (LG 12) and that this region is involved in a reciprocal translocation with LG 2 in one gamma-ray induced mutation, cyc(b213). The translocated segments together cover approximately 5% of the genetic map, and we show that this rearrangement is useful for mapping cloned genes that reside in the affected chromosomal regions. The other two alleles, cyc(b16) and cyc(b229), have deletions in the distal region of LG 12. Interestingly, both of these mutations suppress recombination between genetic markers in LG 12, including markers at a distance from the deletion. This observation raises the possibility that these deletions affect a site required for meiotic recombination on the LG 12 chromosome. The cyc(b16) and cyc(b229) mutations may be useful for balancing other lethal mutations located in the distal region of LG 12. These results show that chromosomal rearrangements can provide useful resources for mapping and genetic analyses in zebrafish.

    View details for Web of Science ID 000071494000034

    View details for PubMedID 9475747

    View details for PubMedCentralID PMC1459804

  • Genetic interactions in zebrafish midline development DEVELOPMENTAL BIOLOGY Halpern, M. E., Hatta, K., Amacher, S. L., Talbot, W. S., Yan, Y. L., Thisse, B., Thisse, C., Postlethwait, J. H., Kimmel, C. B. 1997; 187 (2): 154-170

    Abstract

    Mutational analyses have shown that the genes no tail (ntl, Brachyury homolog), floating head (flh, a Not homeobox gene), and cyclops (cyc) play direct and essential roles in the development of midline structures in the zebrafish. In both ntl and flh mutants a notochord does not develop, and in cyc mutants the floor plate is nearly entirely missing. We made double mutants to learn how these genes might interact. Midline development is disrupted to a greater extent in cyc;flh double mutants than in either cyc or flh single mutants; their effects appear additive. Both the notochord and floor plate are completely lacking, and other phenotypic disturbances suggest that midline signaling functions are severely reduced. On the other hand, trunk midline defects in flh;ntl double mutants are not additive, but are most often similar to those in ntl single mutants. This finding reveals that loss of ntl function can suppress phenotypic defects due to mutation at flh, and we interpret it to mean that the wild-type allele of ntl (ntl+) functions upstream to flh in a regulatory hierarchy. Loss of function of ntl also strongly suppresses the floor plate deficiency in cyc mutants, for we found trunk floor plate to be present in cyc;ntl double mutants. From these findings we propose that ntl+ plays an early role in cell fate choice at the dorsal midline, mediated by the Ntl protein acting to antagonize floor plate development as well as to promote notochord development.

    View details for Web of Science ID A1997XM60100003

    View details for PubMedID 9242414

  • Zebrafish genomics: From mutants to genes TRENDS IN GENETICS Postlethwait, J. H., Talbot, W. S. 1997; 13 (5): 183-190

    Abstract

    Exquisite embryonic lethal mutations have been isolated in hundreds of genes necessary for zebrafish development. Analysis of this resource promises to enhance our understanding of the molecular genetic mechanisms of vertebrate development. This review discusses the state of the zebrafish genome project and the genetic trickery that can expedite molecular isolation of genes disrupted by these mutations.

    View details for Web of Science ID A1997WX39200007

    View details for PubMedID 9154001

  • The one-eyed pinhead gene functions in mesoderm and endoderm formation in zebrafish and interacts with no tail DEVELOPMENT Schier, A. F., Neuhauss, S. C., Helde, K. A., Talbot, W. S., Driever, W. 1997; 124 (2): 327-342

    Abstract

    The zebrafish locus one-eyed pinhead (oep) is essential for the formation of anterior axial mesoderm, endoderm and ventral neuroectoderm. At the beginning of gastrulation anterior axial mesoderm cells form the prechordal plate and express goosecoid (gsc) in wild-type embryos. In oep mutants the prechordal plate does not form and gsc expression is not maintained. Exposure to lithium, a dorsalizing agent, leads to the ectopic induction and maintenance of gsc expression in wild-type embryos. Lithium treatment of oep mutants still leads to ectopic gsc induction but not maintenance, suggesting that oep acts downstream of inducers of dorsal mesoderm. In genetic mosaics, wild-type cells are capable of forming anterior axial mesoderm in oep embryos, suggesting that oep is required in prospective anterior axial mesoderm cells before gastrulation. The oep gene is also essential for endoderm formation and the early development of ventral neuroectoderm, including the floor plate. The loss of endoderm is already manifest during gastrulation by the absence of axial-expressing cells in the hypoblast of oep mutants. These findings suggest that oep is also required in lateral and ventral regions of the gastrula margin. The sonic hedgehog (shh).gene is expressed in the notochord of oep animals. Therefore, the impaired floor plate development in oep mutants is not caused by the absence of the floor plate inducer shh. This suggests that oep is required downstream or in parallel to shh signaling. The ventral region of the forebrain is also absent in oep mutants, leading to severe cyclopia. In contrast, anterior-posterior brain patterning appears largely unaffected, suggesting that underlying prechordal plate is not required for anterior-posterior pattern formation but might be involved in dorsoventral brain patterning. To test if oep has a wider, partially redundant role, we constructed double mutants with two other zebrafish loci essential for patterning during gastrulation. Double mutants with floating head, the zebrafish Xnot homologue, display enhanced floor plate and adaxial muscle phenotypes. Double mutants with no tail (ntl), the zebrafish homologue of the mouse Brachyury locus, display severe defects in midline and mesoderm formation including absence of most of the somitic mesoderm. These results reveal a redundant function of oep and ntl in mesoderm formation. Our data suggest that both oep and ntl act in the blastoderm margin to specify mesendodermal cell fates.

    View details for Web of Science ID A1997WH00400007

    View details for PubMedID 9053309

  • ECDYSONE RECEPTOR EXPRESSION IN THE CNS CORRELATES WITH STAGE-SPECIFIC RESPONSES TO ECDYSTEROIDS DURING DROSOPHILA AND MANDUCA DEVELOPMENT DEVELOPMENT Truman, J. W., Talbot, W. S., Fahrbach, S. E., Hogness, D. S. 1994; 120 (1): 219-234

    Abstract

    In insects, the ecdysteroids act to transform the CNS from its larval to its adult form. A key gene in this response is the ecdysone receptor (EcR), which has been shown in Drosophila to code for 3 protein isoforms. Two of these isoforms, EcR-A and EcR-B1, are prominently expressed in the CNS and we have used isoform-specific antibodies to examine their fluctuations through postembryonic life. EcR expression at the onset of metamorphosis is extremely diverse but specific patterns of EcR expression correlate with distinct patterns of steroid response. Most larval neurons show high levels of EcR-B1 at the start of metamorphosis, a time when they lose larval features in response to ecdysteroids. Earlier, during the larval molts, the same cells have no detectable receptors and show no response to circulating ecdysteroids; later, during the pupal-adult transformation, they switch to EcR-A expression and respond by maturing to their adult form. During the latter period, a subset of the larval neurons hyperexpress EcR-A and these cells are fated to die after the emergence of the adult. The stem cells for the imaginal neurons show prominent EcR-B1 expression during the last larval stage correlated with their main proliferative period. Most imaginal neurons, by contrast, express only EcR-A when they subsequently initiate maturation at the start of metamorphosis. The imaginal neurons of the mushroom bodies are unusual amongst imaginal neurons in expressing the B1 isoform at the start of metamorphosis but they also show regressive changes at this time as they lose their larval axons. Imaginal neurons of the optic lobe show a delayed expression of EcR-B1 through the period when cell-cell interactions are important for establishing connections within this region of the CNS. Overall, the appearance of the two receptor isoforms in cells correlates with different types of steroid responses: EcR-A predominates when cells are undergoing maturational responses whereas EcR-B1 predominates during proliferative activity or regressive responses. The heterogeneity of EcR expression at the start of metamorphosis presumably reflects the diverse origins and requirements of the neurons that nevertheless are all exposed to a common hormonal signal.

    View details for Web of Science ID A1994MT59500021

    View details for PubMedID 8119129

  • PROGRAMMED CELL-DEATH IN THE DROSOPHILA CNS IS ECDYSONE-REGULATED AND COUPLED WITH A SPECIFIC ECDYSONE RECEPTOR ISOFORM DEVELOPMENT Robinow, S., Talbot, W. S., Hogness, D. S., Truman, J. W. 1993; 119 (4): 1251-1259

    Abstract

    At adult emergence, the ventral CNS of Drosophila shows a group of approximately 300 neurons, which are unique in that they express 10-fold higher levels of the A isoform of the ecdysone receptor (EcR-A) than do other central neurons. This expression pattern is established early in metamorphosis and persists throughout the remainder of the pupal stage. Although these cells represent a heterogeneous group of neurons, they all share the same fate of undergoing rapid degeneration after the adult emerges from the pupal case. One prerequisite for this death is the decline of ecdysteroids at the end of metamorphosis. Treatment of flies with 20-hydroxyecdysone blocks the death of the cells, but only if given at least 3 hours before the normal time of degeneration. The correlation of a unique pattern of receptor isoform expression with a particular steroid-regulated fate suggests that variations in the pattern of receptor isoform expression may serve as important switches during development.

    View details for Web of Science ID A1993MM50900023

    View details for PubMedID 8306887

  • THE ORIGIN OF MHC CLASS-II GENE POLYMORPHISM WITHIN THE GENUS MUS NATURE McConnell, T. J., Talbot, W. S., McIndoe, R. A., Wakeland, E. K. 1988; 332 (6165): 651-654

    Abstract

    The I region of the major histocompatibility complex (MHC) of the mouse (H-2) contains a tightly-linked cluster of highly polymorphic genes (class II MHC genes) which control immune responsiveness. Speculation on the origin of this polymorphism, which is believed to be essential for the function of the class II proteins in immune responses to disease, has given rise to two hypotheses. The first is that hypermutational mechanisms (gene conversion or segmental exchange) promote the rapid generation of diversity in MHC genes. The alternative is that polymorphism has arisen from the steady accumulation of mutations over long evolutionary periods, and multiple specific alleles have survived speciation (trans-species evolution). We have looked for evidence of 'segmental exchange' and/or 'trans-species evolution' in the class II genes of the genus Mus by molecular genetic analysis of I-A beta alleles. The results indicate that greater than 90% (28 out of 31) of the alleles examined can be organized into two evolutionary groups both on the basis of restriction site polymorphisms and by the presence or absence of a short interspersed nucleotide element (SINE). Using this SINE sequence as an evolutionary tag, we demonstrate that I-A beta alleles in these two evolutionary groups diverged at least three million years ago and have survived the speciation events leading to several modern Mus species. Nucleotide sequence comparisons of eight Mus m. domesticus I-A beta alleles representing all three evolutionary groups indicate that most of the divergence in exon sequences is due to the steady accumulation of mutations that are maintained independently in the different alleles. But segmental exchanges between alleles from different evolutionary groups have also played a role in the diversification of beta 1 exons.

    View details for Web of Science ID A1988M921800068

    View details for PubMedID 2895893