Honors & Awards

  • MERIT Award, NIH (2014)
  • Election to Membership, American Association of University Pathologists (The Pluto Society) (2014)
  • Election to Membership, Association of American Physicians (2011)
  • Election to Membership, American Society for Clinical Investigation (2004)
  • Cancer Research Fund of the Damon Runyon-Walter Winchell Foundation, Damon-Runyon Scholar Award - Connie and Bob Lurie Scholar (1999)
  • Junior Faculty Scholars Award, Howard Hughes Medical Institute (1998)
  • Paul E. Strandjord Young Investigator Award, Academy of Clinical Laboratory Physicians and Scientists (1994)
  • Clinical Investigator Award, NIH (1993)

Professional Education

  • M.D., Ph.D., Washington University Sch of Med, Medicine and Molecular Biology (1991)
  • Sc.B., Brown University, Biochemistry (1981)

Current Research and Scholarly Interests

The overarching goal of the research in my lab is to understand how signal transduction pathways regulate morphogenesis - the emergence of spatial organization - during development. Development requires that cells differentiate to acquire the necessary complement of cell fates, and that they adopt the structure required to carry out their functions. In multicellular organisms, signal transduction is essential to these processes, yet while our understanding of how signals regulate gene expression is relatively advanced, our understanding of how signals direct the acquisition of specific shapes and forms is less advanced.

Our major project is to investige a pathway that controls the polarity of epithelial cells within the plane of the epithelium. Epithelia delimit compartments of differing composition, and are necessarily specialized on their apical and basal surfaces. In addition, many epithelial cells are overtly polarized along an axis orthogonal to the apical-basal axis, in a direction defined by the organization of the tissue or organ [referred to as planar cell polarity (PCP)]. In effect, therefore, cells acquire a global “knowledge” of which way is which, much as a compass tells us direction on the earth’s surface. Some examples include the specialized hair cells of the mammalian cochlea, that display a spectacularly polarized organization of kinocilia and stereocilia on their apical surfaces, the dynamic ciliated cells of the tracheal and reproductive tract epithelia, and cells in the gastrulating vertebrate embryo that display polarized migration and intercalation behaviors. In each case, PCP is critical to the function of these cells and tissues, and errors in the signaling system controlling PCP lead to human diseases and developmental defects, including congenital deafness, neural tube closure defects and cardiac outflow tract anomalies. The primary goal of my work on PCP has been to elucidate, at molecular and cell biological levels, the nature of the signals that induce subcellular asymmetry, and how cells then respond to this molecular asymmetry to orient their cytoskeletons.

We employ two principal model systems in our work. Because of the availability of remarkably powerful genetic, molecular and cell biological tools, we use the fruitfly, Drosophila melanogaster, as our primary model for investigating the fundamental mechanisms of PCP signaling. Importantly, flies have proven to be a remarkably well-conserved model for the molecular mechanisms of signaling events that direct vertebrate development. More recently, we have taken advantage of our experience in studying these mechanisms to extend our work to vertebrates, using primarily the mouse. To date, our work on vertebrates, along with the work of others, indicates a substantial conservation, but also reveals numerous differences and variations deserving of further study.

2023-24 Courses

Stanford Advisees

Graduate and Fellowship Programs

All Publications

  • Prickle isoforms determine handedness of helical morphogenesis. eLife Cho, B., Song, S., Axelrod, J. D. 2020; 9


    Subcellular asymmetry directed by the planar cell polarity (PCP) signaling pathway orients numerous morphogenetic events in both invertebrates and vertebrates. Here, we describe a morphogenetic movement in which the intertwined socket and shaft cells of the Drosophila anterior wing margin mechanosensory bristles undergo PCP-directed apical rotation, inducing twisting that results in a helical structure of defined chirality. We show that the Frizzled/Vang PCP signaling module coordinates polarity among and between bristles and surrounding cells to direct this rotation. Furthermore, we show that dynamic interplay between two isoforms of the Prickle protein determines right- or left-handed bristle morphogenesis. We provide evidence that, Frizzled/Vang signaling couples to the Fat/Dachsous PCP directional signal in opposite directions depending on whether Pkpk or Pksple predominates. Dynamic interplay between Pk isoforms is likely to be an important determinant of PCP outcomes in diverse contexts. Similar mechanisms may orient other lateralizing morphogenetic processes.

    View details for DOI 10.7554/eLife.51456

    View details for PubMedID 31934858

  • Cyclin-dependent kinase control of motile ciliogenesis ELIFE Vladar, E. K., Stratton, M. B., Saal, M. L., Salazar-De Simone, G., Wang, X., Wolgemuth, D., Stearns, T., Axelrod, J. D. 2018; 7


    Cycling cells maintain centriole number at precisely two per cell in part by limiting their duplication to S phase under the control of the cell cycle machinery. In contrast, postmitotic multiciliated cells (MCCs) uncouple centriole assembly from cell cycle progression and produce hundreds of centrioles in the absence of DNA replication to serve as basal bodies for motile cilia. Although some cell cycle regulators have previously been implicated in motile ciliogenesis, how the cell cycle machinery is employed to amplify centrioles is unclear. We use transgenic mice and primary airway epithelial cell culture to show that Cdk2, the kinase responsible for the G1 to S phase transition, is also required in MCCs to initiate motile ciliogenesis. While Cdk2 is coupled with cyclins E and A2 during cell division, cyclin A1 is required during ciliogenesis, contributing to an alternative regulatory landscape that facilitates centriole amplification without DNA replication.

    View details for PubMedID 30152757

  • Disruption of Core Planar Cell Polarity Signaling Regulates Renal Tubule Morphogenesis but Is Not Cystogenic CURRENT BIOLOGY Kunimoto, K., Bayly, R. D., Vladar, E. K., Vonderfecht, T., Gallagher, A., Axelrod, J. D. 2017; 27 (20): 3120-+


    Oriented cell division (OCD) and convergent extension (CE) shape developing renal tubules, and their disruption has been associated with polycystic kidney disease (PKD) genes, the majority of which encode proteins that localize to primary cilia. Core planar cell polarity (PCP) signaling controls OCD and CE in other contexts, leading to the hypothesis that disruption of PCP signaling interferes with CE and/or OCD to produce PKD. Nonetheless, the contribution of PCP to tubulogenesis and cystogenesis is uncertain, and two major questions remain unanswered. Specifically, the inference that mutation of PKD genes interferes with PCP signaling is untested, and the importance of PCP signaling for cystogenic PKD phenotypes has not been examined. We show that, during proliferative stages, PCP signaling polarizes renal tubules to control OCD. However, we find that, contrary to the prevailing model, PKD mutations do not disrupt PCP signaling but instead act independently and in parallel with PCP signaling to affect OCD. Indeed, PCP signaling that is normally downregulated once development is completed is retained in cystic adult kidneys. Disrupting PCP signaling results in inaccurate control of tubule diameter, a tightly regulated parameter with important physiological ramifications. However, we show that disruption of PCP signaling is not cystogenic. Our results suggest that regulating tubule diameter is a key function of PCP signaling but that loss of this control does not induce cysts.

    View details for PubMedID 29033332

    View details for PubMedCentralID PMC5683414

  • Clustering and negative feedback by endocytosis in planar cell polarity signaling is modulated by ubiquitinylation of prickle. PLoS genetics Cho, B., Pierre-Louis, G., Sagner, A., Eaton, S., Axelrod, J. D. 2015; 11 (5)


    The core components of the planar cell polarity (PCP) signaling system, including both transmembrane and peripheral membrane associated proteins, form asymmetric complexes that bridge apical intercellular junctions. While these can assemble in either orientation, coordinated cell polarization requires the enrichment of complexes of a given orientation at specific junctions. This might occur by both positive and negative feedback between oppositely oriented complexes, and requires the peripheral membrane associated PCP components. However, the molecular mechanisms underlying feedback are not understood. We find that the E3 ubiquitin ligase complex Cullin1(Cul1)/SkpA/Supernumerary limbs(Slimb) regulates the stability of one of the peripheral membrane components, Prickle (Pk). Excess Pk disrupts PCP feedback and prevents asymmetry. We show that Pk participates in negative feedback by mediating internalization of PCP complexes containing the transmembrane components Van Gogh (Vang) and Flamingo (Fmi), and that internalization is activated by oppositely oriented complexes within clusters. Pk also participates in positive feedback through an unknown mechanism promoting clustering. Our results therefore identify a molecular mechanism underlying generation of asymmetry in PCP signaling.

    View details for DOI 10.1371/journal.pgen.1005259

    View details for PubMedID 25996914

    View details for PubMedCentralID PMC4440771

  • Microtubules provide directional information for core PCP function ELIFE Matis, M., Russler-Germain, D. A., Hu, Q., Tomlin, C. J., Axelrod, J. D. 2014; 3


    Planar cell polarity (PCP) signaling controls the polarization of cells within the plane of an epithelium. Two molecular modules composed of Fat(Ft)/Dachsous(Ds)/Four-jointed(Fj) and a 'PCP-core' including Frizzled(Fz) and Dishevelled(Dsh) contribute to polarization of individual cells. How polarity is globally coordinated with tissue axes is unresolved. Consistent with previous results, we find that the Ft/Ds/Fj-module has an effect on a MT-cytoskeleton. Here, we provide evidence for the model that the Ft/Ds/Fj-module provides directional information to the core-module through this MT organizing function. We show Ft/Ds/Fj-dependent initial polarization of the apical MT-cytoskeleton prior to global alignment of the core-module, reveal that the anchoring of apical non-centrosomal MTs at apical junctions is polarized, observe that directional trafficking of vesicles containing Dsh depends on Ft, and demonstrate the feasibility of this model by mathematical simulation. Together, these results support the hypothesis that Ft/Ds/Fj provides a signal to orient core PCP function via MT polarization.

    View details for DOI 10.7554/eLife.02893.001

    View details for Web of Science ID 000341259200007

    View details for PubMedID 25124458

    View details for PubMedCentralID PMC4151085

  • Asymmetric homotypic interactions of the atypical cadherin Flamingo mediate intercellular polarity signaling CELL Chen, W., Antic, D., Matis, M., Logan, C. Y., Povelones, M., Anderson, G. A., Nusse, R., Axelrod, J. D. 2008; 133 (6): 1093-1105


    Acquisition of planar cell polarity (PCP) in epithelia involves intercellular communication, during which cells align their polarity with that of their neighbors. The transmembrane proteins Frizzled (Fz) and Van Gogh (Vang) are essential components of the intercellular communication mechanism, as loss of either strongly perturbs the polarity of neighboring cells. How Fz and Vang communicate polarity information between neighboring cells is poorly understood. The atypical cadherin, Flamingo (Fmi), is implicated in this process, yet whether Fmi acts permissively as a scaffold or instructively as a signal is unclear. Here, we provide evidence that Fmi functions instructively to mediate Fz-Vang intercellular signal relay, recruiting Fz and Vang to opposite sides of cell boundaries. We propose that two functional forms of Fmi, one of which is induced by and physically interacts with Fz, bind each other to create cadherin homodimers that signal bidirectionally and asymmetrically, instructing unequal responses in adjacent cell membranes to establish molecular asymmetry.

    View details for DOI 10.1016/j.cell.2008.04.048

    View details for Web of Science ID 000256693400023

    View details for PubMedID 18555784

    View details for PubMedCentralID PMC2446404

  • Mathematical modeling of planar cell polarity to understand domineering nonautonomy SCIENCE Amonlirdviman, K., Khare, N. A., Tree, D. R., Chen, W. S., Axelrod, J. D., Tomlin, C. J. 2005; 307 (5708): 423-426


    Planar cell polarity (PCP) signaling generates subcellular asymmetry along an axis orthogonal to the epithelial apical-basal axis. Through a poorly understood mechanism, cell clones that have mutations in some PCP signaling components, including some, but not all, alleles of the receptor frizzled, cause polarity disruptions of neighboring wild-type cells, a phenomenon referred to as domineering nonautonomy. Here, a contact-dependent signaling hypothesis, derived from experimental results, is shown by reaction-diffusion, partial differential equation modeling and simulation to fully reproduce PCP phenotypes, including domineering nonautonomy, in the Drosophila wing. The sufficiency of this model and the experimental validation of model predictions reveal how specific protein-protein interactions produce autonomy or domineering nonautonomy.

    View details for DOI 10.1126/science.1105471

    View details for Web of Science ID 000226492300047

    View details for PubMedID 15662015

  • Automated counting of Drosophila imaginal disc cell nuclei. Biology open Bosch, P. S., Axelrod, J. D. 2024


    Automated image quantification workflows have dramatically improved over the past decade, enriching image analysis and enhancing the ability to achieve statistical power. These analyses have proved especially useful for studies in organisms such as Drosophila melanogaster, where it is relatively simple to obtain high sample numbers for downstream analyses. However, the developing wing, an intensively utilized structure in developmental biology, has eluded efficient cell counting workflows due to its highly dense cellular population. Here, we present efficient automated cell counting workflows capable of quantifying cells in the developing wing. Our workflows can count the total number of cells or count cells in clones labeled with a fluorescent nuclear marker in imaginal discs. Moreover, by training a machine-learning algorithm we have developed a workflow capable of segmenting and counting twin-spot labeled nuclei, a challenging problem requiring distinguishing heterozygous and homozygous cells in a background of regionally varying intensity. Our workflows could potentially be applied to any tissue with high cellular density, as they are structure-agnostic, and only require a nuclear label to segment and count cells.

    View details for DOI 10.1242/bio.060254

    View details for PubMedID 38345430

  • Protein phosphatase 1 regulates core PCP signaling. EMBO reports Song, S., Cho, B., Weiner, A. T., Nissen, S. B., Ojeda Naharros, I., Sanchez Bosch, P., Suyama, K., Hu, Y., He, L., Svinkina, T., Udeshi, N. D., Carr, S. A., Perrimon, N., Axelrod, J. D. 2023: e56997


    Planar cell polarity (PCP) signaling polarizes epithelial cells within the plane of an epithelium. Core PCP signaling components adopt asymmetric subcellular localizations within cells to both polarize and coordinate polarity between cells. Achieving subcellular asymmetry requires additional effectors, including some mediating post-translational modifications of core components. Identification of such proteins is challenging due to pleiotropy. We used mass spectrometry-based proximity labeling proteomics to identify such regulators in the Drosophila wing. We identified the catalytic subunit of protein phosphatase1, Pp1-87B, and show that it regulates core protein polarization. Pp1-87B interacts with the core protein Van Gogh and at least one serine/threonine kinase, Dco/CKIε, that is known to regulate PCP. Pp1-87B modulates Van Gogh subcellular localization and directs its dephosphorylation in vivo. PNUTS, a Pp1 regulatory subunit, also modulates PCP. While the direct substrate(s) of Pp1-87B in control of PCP is not known, our data support the model that cycling between phosphorylated and unphosphorylated forms of one or more core PCP components may regulate acquisition of asymmetry. Finally, our screen serves as a resource for identifying additional regulators of PCP signaling.

    View details for DOI 10.15252/embr.202356997

    View details for PubMedID 37975164

  • A WNT4 and DKK3 driven canonical to noncanonical Wnt signaling switch controls multiciliogenesis. Journal of cell science Cooney, R. A., Saal, M. L., Geraci, K. P., Maynard, C., Cleaver, O., Hoang, O. N., Moore, T. T., Hwang, R. F., Axelrod, J. D., Vladar, E. K. 2023


    Multiciliated cells contain hundreds of cilia whose directional movement powers the mucociliary clearance of the airways, a vital host defense mechanism. Multiciliated cell specification requires canonical Wnt signaling, which then must be turned off. Next, ciliogenesis and polarized ciliary orientation are regulated by noncanonical Wnt/planar cell polarity signaling. The mechanistic relationship between the Wnt pathways is unknown. We show that DKK3, a secreted canonical Wnt regulator and WNT4, a noncanonical Wnt ligand act together to facilitate a canonical to noncanonical Wnt signaling switch during multiciliated cell formation. In primary human airway epithelial cells, DKK3 and WNT4 CRISPR knockout blocks, while ectopic expression promotes multiciliated cell formation by inhibiting canonical Wnt signaling. Wnt4 and Dkk3 knockout mice also display defective ciliated cells. DKK3 and WNT4 are co-secreted from basal stem cells and act directly on multiciliated cells via KREMEN1 and FZD6, respectively. We provide a novel mechanism that links specification to cilium biogenesis and polarization for proper multiciliated cell formation.

    View details for DOI 10.1242/jcs.260807

    View details for PubMedID 37505110

  • Notch signaling inactivation by small molecule gamma-secretase inhibitors restores the multiciliated cell population in the airway epithelium. American journal of physiology. Lung cellular and molecular physiology Vladar, E. K., Kunimoto, K., Rojas-Hernandez, L. S., Spano, J. M., Sellers, Z. M., Joo, N. S., Cooney, R. A., Axelrod, J. D., Milla, C. E. 2023


    Multiciliated cell loss is a hallmark of airway epithelial remodeling in chronic inflammatory airway diseases including cystic fibrosis (CF), asthma, and chronic obstructive pulmonary disease. It disrupts mucociliary clearance, which fuels disease progression. Effective clearance requires an optimal proportion of multiciliated and secretory cells. This is controlled by Notch signaling such that between two adjacent cells the one that activates Notch becomes a secretory cell and the one that avoids Notch activation becomes a multiciliated cell. Consequently, blocking Notch by a small molecule inhibitor of the gamma-secretase enzyme that cleaves the Notch receptor for signal activation directs differentiation towards the multiciliated lineage. Thus, gamma-secretase inhibitor (GSI) treatment may alleviate multiciliated cell loss in lung disease. Here we demonstrate therapeutic restoration of multiciliated cells by the GSI LY450139 (semagacestat). LY450139 increased multiciliated cell numbers in a dose-dependent manner in healthy primary human nasal epithelial cells (HNECs) during differentiation and in mature cultures, but not when applied during early epithelialization of progenitors. LY450139 did not impact stem cell proliferation. Basal and apical administration were equally effective. In healthy adult mice, LY450139 increased multiciliated cell numbers without detectible toxicity. LY450139 also increased multiciliated cells and decreased excess mucus secretory cells in CF HNECs and IL-13 remodeled healthy HNECs. LY450139 normalized multiciliated cell numbers in CF HNECs without interfering with the activity of CFTR modulator compounds. In sum, we demonstrate that GSI administration is a promising therapeutic to restore multiciliated cells and potentially improve epithelial function in a wide range of chronic lung diseases.

    View details for DOI 10.1152/ajplung.00382.2022

    View details for PubMedID 37039381

  • A stem cell roadmap of ribosome heterogeneity reveals a function for RPL10A in mesoderm production. Nature communications Genuth, N. R., Shi, Z., Kunimoto, K., Hung, V., Xu, A. F., Kerr, C. H., Tiu, G. C., Oses-Prieto, J. A., Salomon-Shulman, R. E., Axelrod, J. D., Burlingame, A. L., Loh, K. M., Barna, M. 2022; 13 (1): 5491


    Recent findings suggest that the ribosome itself modulates gene expression. However, whether ribosomes change composition across cell types or control cell fate remains unknown. Here, employing quantitative mass spectrometry during human embryonic stem cell differentiation, we identify dozens of ribosome composition changes underlying cell fate specification. We observe upregulation of RPL10A/uL1-containing ribosomes in the primitive streak followed by progressive decreases during mesoderm differentiation. An Rpl10a loss-of-function allele in mice causes striking early mesodermal phenotypes, including posterior trunk truncations, and inhibits paraxial mesoderm production in culture. Ribosome profiling in Rpl10a loss-of-function mice reveals decreased translation of mesoderm regulators, including Wnt pathway mRNAs, which are also enriched on RPL10A/uL1-containing ribosomes. We further show that RPL10A/uL1 regulates canonical and non-canonical Wnt signaling during stem cell differentiation and in the developing embryo. These findings reveal unexpected ribosome composition modularity that controls differentiation and development through the specialized translation of key signaling networks.

    View details for DOI 10.1038/s41467-022-33263-3

    View details for PubMedID 36123354

  • Distinct overlapping functions for Prickle1 and Prickle2 in the polarization of the airway epithelium. Frontiers in cell and developmental biology Kunimoto, K., Weiner, A. T., Axelrod, J. D., Vladar, E. K. 2022; 10: 976182


    Planar cell polarity (PCP) signaling polarizes cells within the plane of an epithelium. In the airways, planar cell polarity signaling orients the directional beating of motile cilia required for effective mucociliary clearance. The planar cell polarity signaling mechanism is best understood from work in Drosophila, where it has been shown to both coordinate the axis of polarity between cells and to direct the morphological manifestations of polarization within cells. The 'core' planar cell polarity signaling mechanism comprises two protein complexes that segregate to opposite sides of each cell and interact with the opposite complex in neighboring cells. Proper subcellular localization of core planar cell polarity proteins correlates with, and is almost certainly responsible for, their ability to direct polarization. This mechanism is highly conserved from Drosophila to vertebrates, though for most of the core genes, mammals have multiple paralogs whereas Drosophila has only one. In the mouse airway epithelium, the core protein Prickle2 segregates asymmetrically, as is characteristic for core proteins, but is only present in multiciliated cells and is absent from other cell types. Furthermore, Prickle2 mutant mice show only modest ciliary polarity defects. These observations suggest that other Prickle paralogs might contribute to polarization. Here, we show that Prickle1 segregates asymmetrically in multiciliated and nonciliated airway epithelial cell types, that compared to Prickle2, Prickle1 has different spatial and temporal expression dynamics and a stronger ciliary polarity phenotype, and that Prickle1 and Prickle2 mutants genetically interact. We propose distinct and partially overlapping functions for the Prickle paralogs in polarization of the airway epithelium.

    View details for DOI 10.3389/fcell.2022.976182

    View details for PubMedID 36176272

    View details for PubMedCentralID PMC9513604

  • Prickle isoform participation in distinct polarization events in the Drosophila eye. PloS one Cho, B., Song, S., Wan, J. Y., Axelrod, J. D. 2022; 17 (2): e0262328


    Planar cell polarity (PCP) signaling regulates several polarization events during development of ommatidia in the Drosophila eye, including directing chirality by polarizing a cell fate choice and determining the direction and extent of ommatidial rotation. The pksple isoform of the PCP protein Prickle is known to participate in the R3/R4 cell fate decision, but the control of other polarization events and the potential contributions of the three Pk isoforms have not been clarified. Here, by characterizing expression and subcellular localization of individual isoforms together with re-analyzing isoform specific phenotypes, we show that the R3/R4 fate decision, its coordination with rotation direction, and completion of rotation to a final ±90° rotation angle are separable polarization decisions with distinct Pk isoform requirements and contributions. Both pksple and pkpk can enforce robust R3/R4 fate decisions, but only pksple can correctly orient them along the dorsal-ventral axis. In contrast, pksple and pkpk can fully and interchangeably sustain coordination of rotation direction and rotation to completion. We propose that expression dynamics and competitive interactions determine isoform participation in these processes. We propose that the selective requirement for pksple to orient the R3/R4 decision and their interchangeability for coordination and completion of rotation reflects their previously described differential interaction with the Fat/Dachsous system which is known to be required for orientation of R3/R4 decisions but not for coordination or completion of rotation.

    View details for DOI 10.1371/journal.pone.0262328

    View details for PubMedID 35148314

  • Endosomal Wnt signaling proteins control microtubule nucleation in dendrites. PLoS biology Weiner, A. T., Seebold, D. Y., Torres-Gutierrez, P. n., Folker, C. n., Swope, R. D., Kothe, G. O., Stoltz, J. G., Zalenski, M. K., Kozlowski, C. n., Barbera, D. J., Patel, M. A., Thyagarajan, P. n., Shorey, M. n., Nye, D. M., Keegan, M. n., Behari, K. n., Song, S. n., Axelrod, J. D., Rolls, M. M. 2020; 18 (3): e3000647


    Dendrite microtubules are polarized with minus-end-out orientation in Drosophila neurons. Nucleation sites concentrate at dendrite branch points, but how they localize is not known. Using Drosophila, we found that canonical Wnt signaling proteins regulate localization of the core nucleation protein γTubulin (γTub). Reduction of frizzleds (fz), arrow (low-density lipoprotein receptor-related protein [LRP] 5/6), dishevelled (dsh), casein kinase Iγ, G proteins, and Axin reduced γTub-green fluorescent protein (GFP) at branch points, and two functional readouts of dendritic nucleation confirmed a role for Wnt signaling proteins. Both dsh and Axin localized to branch points, with dsh upstream of Axin. Moreover, tethering Axin to mitochondria was sufficient to recruit ectopic γTub-GFP and increase microtubule dynamics in dendrites. At dendrite branch points, Axin and dsh colocalized with early endosomal marker Rab5, and new microtubule growth initiated at puncta marked with fz, dsh, Axin, and Rab5. We propose that in dendrites, canonical Wnt signaling proteins are housed on early endosomes and recruit nucleation sites to branch points.

    View details for DOI 10.1371/journal.pbio.3000647

    View details for PubMedID 32163403

  • Planar cell polarity signaling in the development of left-right asymmetry. Current opinion in cell biology Axelrod, J. D. 2019; 62: 61–69


    The planar cell polarity (PCP) signaling pathway, principally understood from work in Drosophila, is now known to contribute to development in a broad swath of the animal kingdom, and its impairment leads to developmental malformations and diseases affecting humans. The 'core' mechanism underlying PCP signaling polarizes sheets of cells, aligning them in a head-to-tail fashion within the sheet. Cells use the resulting directional information to guide a wide variety of processes. One such process is lateralization, the determination of left-right asymmetry that guides the asymmetric morphology and placement of internal organs. Recent evidence extends the idea that PCP signaling underlies the earliest steps in lateralizationand that PCP is invoked again during asymmetric morphogenesis of organs including the heart and gut.

    View details for DOI 10.1016/j.ceb.2019.09.002

    View details for PubMedID 31654871

  • VANGL2 regulates luminal epithelial organization and cell turnover in the mammary gland. Scientific reports Smith, P., Godde, N., Rubio, S., Tekeste, M., Vladar, E. K., Axelrod, J. D., Henderson, D. J., Milgrom-Hoffman, M., Humbert, P. O., Hinck, L. 2019; 9 (1): 7079


    The VANGL family of planar cell polarity proteins is implicated in breast cancer however its function in mammary gland biology is unknown. Here, we utilized a panel of Vang1 and Vangl2 mouse alleles to examine the requirement of VANGL family members in the murine mammary gland. We show that Vang1CKODelta/Delta glands display normal branching while Vangl2flox/flox and Vangl2Lp/Lp tissue exhibit several phenotypes. In MMTV-Cre;Vangl2flox/flox glands, cell turnover is reduced and lumens are narrowed. A Vangl2 missense mutation in the Vangl2Lp/Lp tissue leads to mammary anlage sprouting defects and deficient outgrowth with transplantation of anlage or secondary tissue fragments. In successful Vangl2Lp/Lp outgrowths, three morphological phenotypes are observed: distended ducts, supernumerary end buds, and ectopic acini. Layer specific defects are observed with loss of Vangl2 selectively in either basal or luminal layers of mammary cysts. Loss in the basal compartment inhibits cyst formation, but has the opposite effect in the luminal compartment. Candidate gene analysis on MMTV-Cre;Vangl2flox/flox and Vangl2Lp/Lp tissue reveals a significant reduction in Bmi1 expression, with overexpression of Bmi1 rescuing defects in Vangl2 knockdown cysts. Our results demonstrate that VANGL2 is necessary for normal mammary gland development and indicate differential functional requirements in basal versus luminal mammary compartments.

    View details for PubMedID 31068622

  • FijiWingsPolarity: An open source toolkit for semi-automated detection of cell polarity FLY Dobens, L. L., Shipman, A., Axelrod, J. D. 2018; 12 (1): 23–33


    Epithelial cells are defined by apical-basal and planar cell polarity (PCP) signaling, the latter of which establishes an orthogonal plane of polarity in the epithelial sheet. PCP signaling is required for normal cell migration, differentiation, stem cell generation and tissue repair, and defects in PCP have been associated with developmental abnormalities, neuropathologies and cancers. While the molecular mechanism of PCP is incompletely understood, the deepest insights have come from Drosophila, where PCP is manifest in hairs and bristles across the adult cuticle and organization of the ommatidia in the eye. Fly wing cells are marked by actin-rich trichome structures produced at the distal edge of each cell in the developing wing epithelium and in a mature wing the trichomes orient collectively in the distal direction. Genetic screens have identified key PCP signaling pathway components that disrupt trichome orientation, which has been measured manually in a tedious and error prone process. Here we describe a set of image processing and pattern-recognition macros that can quantify trichome arrangements in micrographs and mark these directly by color, arrow or colored arrow to indicate trichome location, length and orientation. Nearest neighbor calculations are made to exploit local differences in orientation to better and more reliably detect and highlight local defects in trichome polarity. We demonstrate the use of these tools on trichomes in adult wing preps and on actin-rich developing trichomes in pupal wing epithelia stained with phalloidin. FijiWingsPolarity is freely available and will be of interest to a broad community of fly geneticists studying the effect of gene function on PCP.

    View details for PubMedID 29189094

    View details for PubMedCentralID PMC5927706

  • Wnt Signaling in Chronic Rhinosinusitis with Nasal Polyps AMERICAN JOURNAL OF RESPIRATORY CELL AND MOLECULAR BIOLOGY Boeske, R., Vladar, E. K., Koennecke, M., Huesing, B., Linke, R., Pries, R., Reiling, N., Axelrod, J. D., Nayak, J. V., Wollenberg, B. 2017; 56 (5): 575-584


    The signaling pathways that sustain the disease process of chronic rhinosinusitis with nasal polyps (CRSwNP) remain poorly understood. We sought to determine the expression levels of Wnt signaling genes in CRSwNP and to study the role of the Wnt pathway in inflammation and epithelial remodeling in the nasal mucosa. Microarrays and real time-quantitative polymerase chain reaction comparing gene expression in matched NPs and inferior turbinates revealed that WNT2B, WNT3A, WNT4, WNT7A, WNT7B, and FZD2 were up-regulated and that FZD1, LRP5, LRP6, and WIF1 were down-regulated in NPs. Immunolabeling showed robust expression of Wnt ligands, nuclear β-catenin, and Axin-2 in NP tissue, suggesting that Wnt/β-catenin signaling is activated in NPs. We used primary human nasal epithelial cell (HNEpC) cultures to test the functional consequences of Wnt pathway activation. Monolayer HNEpCs treated with recombinant human WNT (rhWNT) 3A, but not with rhWNT4, had altered epithelial morphology and decreased adhesion, without loss of viability. We found that neither rhWNT3A nor rhWNT4 treatment induced proliferation. The expression and release of inflammatory cytokines IL-6 and granulocyte-macrophage colony-stimulating factor were increased after rhWNT3A exposure of HNEpCs. When differentiated at an air-liquid interface, rhWNT3A- and WNT agonist-, but not rhWNT4-treated HNEpCs, had abnormal epithelial architecture, failed to undergo motile ciliogenesis, and had defective noncanonical Wnt (planar cell polarity) signaling. On the basis of these results, we propose a model in which Wnt/β-catenin signaling sustains mucosal inflammation and leads to a spectrum of changes consistent with those seen during epithelial remodeling in NPs.

    View details for DOI 10.1165/rcmb.2016-0024OC

    View details for Web of Science ID 000400259400008

  • Wnt Signaling in Chronic Rhinosinusitis with Nasal Polyps. American journal of respiratory cell and molecular biology Böscke, R., Vladar, E. K., Könnecke, M., Hüsing, B., Linke, R., Pries, R., Reiling, N., Axelrod, J. D., Nayak, J. V., Wollenberg, B. 2017


    The signaling pathways that sustain the disease process of chronic rhinosinusitis with nasal polyps (CRSwNP) remain poorly understood. We sought to determine the expression levels of Wnt signaling genes in CRSwNP and to study the role of the Wnt pathway in inflammation and epithelial remodeling in the nasal mucosa. Microarrays and real time-quantitative polymerase chain reaction comparing gene expression in matched NPs and inferior turbinates revealed that WNT2B, WNT3A, WNT4, WNT7A, WNT7B, and FZD2 were up-regulated and that FZD1, LRP5, LRP6, and WIF1 were down-regulated in NPs. Immunolabeling showed robust expression of Wnt ligands, nuclear β-catenin, and Axin-2 in NP tissue, suggesting that Wnt/β-catenin signaling is activated in NPs. We used primary human nasal epithelial cell (HNEpC) cultures to test the functional consequences of Wnt pathway activation. Monolayer HNEpCs treated with recombinant human WNT (rhWNT) 3A, but not with rhWNT4, had altered epithelial morphology and decreased adhesion, without loss of viability. We found that neither rhWNT3A nor rhWNT4 treatment induced proliferation. The expression and release of inflammatory cytokines IL-6 and granulocyte-macrophage colony-stimulating factor were increased after rhWNT3A exposure of HNEpCs. When differentiated at an air-liquid interface, rhWNT3A- and WNT agonist-, but not rhWNT4-treated HNEpCs, had abnormal epithelial architecture, failed to undergo motile ciliogenesis, and had defective noncanonical Wnt (planar cell polarity) signaling. On the basis of these results, we propose a model in which Wnt/β-catenin signaling sustains mucosal inflammation and leads to a spectrum of changes consistent with those seen during epithelial remodeling in NPs.

    View details for DOI 10.1165/rcmb.2016-0024OC

    View details for PubMedID 28059551

  • Airway epithelial homeostasis and planar cell polarity signaling depend on multiciliated cell differentiation. JCI insight Vladar, E. K., Nayak, J. V., Milla, C. E., Axelrod, J. D. 2016; 1 (13)


    Motile airway cilia that propel contaminants out of the lung are oriented in a common direction by planar cell polarity (PCP) signaling, which localizes PCP protein complexes to opposite cell sides throughout the epithelium to orient cytoskeletal remodeling. In airway epithelia, PCP is determined in a 2-phase process. First, cell-cell communication via PCP complexes polarizes all cells with respect to the proximal-distal tissue axis. Second, during ciliogenesis, multiciliated cells (MCCs) undergo cytoskeletal remodeling to orient their cilia in the proximal direction. The second phase not only directs cilium polarization, but also consolidates polarization across the epithelium. Here, we demonstrate that in airway epithelia, PCP depends on MCC differentiation. PCP mutant epithelia have misaligned cilia, and also display defective barrier function and regeneration, indicating that PCP regulates multiple aspects of airway epithelial homeostasis. In humans, MCCs are often sparse in chronic inflammatory diseases, and these airways exhibit PCP dysfunction. The presence of insufficient MCCs impairs mucociliary clearance in part by disrupting PCP-driven polarization of the epithelium. Consistent with defective PCP, barrier function and regeneration are also disrupted. Pharmacological stimulation of MCC differentiation restores PCP and reverses these defects, suggesting its potential for broad therapeutic benefit in chronic inflammatory disease.

    View details for PubMedID 27570836

  • Airway epithelial homeostasis and planar cell polarity signaling depend on multiciliated cell differentiation JCI INSIGHT Vladar, E. K., Nayak, J. V., Milla, C. E., Axelrod, J. D. 2016; 1 (13)
  • Prickle1 mutation causes planar cell polarity and directional cell migration defects associated with cardiac outflow tract anomalies and other structural birth defects BIOLOGY OPEN Gibbs, B. C., Damerla, R. R., Vladar, E. K., Chatterjee, B., Wan, Y., Liu, X., Cui, C., Gabriel, G. C., Zahid, M., Yagi, H., Szabo-Rogers, H. L., Suyama, K. L., Axelrod, J. D., Lo, C. W. 2016; 5 (3): 323-335


    Planar cell polarity (PCP) is controlled by a conserved pathway that regulates directional cell behavior. Here, we show that mutant mice harboring a newly described mutation termed Beetlejuice (Bj) in Prickle1 (Pk1), a PCP component, exhibit developmental phenotypes involving cell polarity defects, including skeletal, cochlear and congenital cardiac anomalies. Bj mutants die neonatally with cardiac outflow tract (OFT) malalignment. This is associated with OFT shortening due to loss of polarized cell orientation and failure of second heart field cell intercalation mediating OFT lengthening. OFT myocardialization was disrupted with cardiomyocytes failing to align with the direction of cell invasion into the outflow cushions. The expression of genes mediating Wnt signaling was altered. Also noted were shortened but widened bile ducts and disruption in canonical Wnt signaling. Using an in vitro wound closure assay, we showed Bj mutant fibroblasts cannot establish polarized cell morphology or engage in directional cell migration, and their actin cytoskeleton failed to align with the direction of wound closure. Unexpectedly, Pk1 mutants exhibited primary and motile cilia defects. Given Bj mutant phenotypes are reminiscent of ciliopathies, these findings suggest Pk1 may also regulate ciliogenesis. Together these findings show Pk1 plays an essential role in regulating cell polarity and directional cell migration during development.

    View details for DOI 10.1242/bio.015750

    View details for Web of Science ID 000372304500014

    View details for PubMedID 26883626

  • Prickle isoforms control the direction of tissue polarity by microtubule independent and dependent mechanisms. Biology open Sharp, K. A., Axelrod, J. D. 2016; 5 (3): 229-236


    Planar cell polarity signaling directs the polarization of cells within the plane of many epithelia. While these tissues exhibit asymmetric localization of a set of core module proteins, in Drosophila, more than one mechanism links the direction of core module polarization to the tissue axes. One signaling system establishes a polarity bias in the parallel, apical microtubules upon which vesicles containing core proteins traffic. Swapping expression of the differentially expressed Prickle isoforms, Prickle and Spiny-legs, reverses the direction of core module polarization. Studies in the proximal wing and the anterior abdomen indicated that this results from their differential control of microtubule polarity. Prickle and Spiny-legs also control the direction of polarization in the distal wing (D-wing) and the posterior abdomen (P-abd). We report here that this occurs without affecting microtubule polarity in these tissues. The direction of polarity in the D-wing is therefore likely determined by a novel mechanism independent of microtubule polarity. In the P-abd, Prickle and Spiny-legs interpret at least two directional cues through a microtubule-polarity-independent mechanism.

    View details for DOI 10.1242/bio.016162

    View details for PubMedID 26863941

  • The Phenotypic Effects of Royal Jelly on Wild-Type D. melanogaster Are Strain-Specific. PloS one Morgan, S. L., Seggio, J. A., Nascimento, N. F., Huh, D. D., Hicks, J. A., Sharp, K. A., Axelrod, J. D., Wang, K. C. 2016; 11 (8)


    The role for royal jelly (RJ) in promoting caste differentiation of honeybee larvae into queens rather than workers is well characterized. A recent study demonstrated that this poorly understood complex nutrition drives strikingly similar phenotypic effects in Drosophila melanogaster, such as increased body size and reduced developmental time, making possible the use of D. melanogaster as a model system for the genetic analysis of the cellular mechanisms underlying RJ and caste differentiation. We demonstrate here that RJ increases the body size of some wild-type strains of D. melanogaster but not others, and report significant delays in developmental time in all flies reared on RJ. These findings suggest that cryptic genetic variation may be a factor in the D. melanogaster response to RJ, and should be considered when attempting to elucidate response mechanisms to environmental changes in non-honeybee species.

    View details for DOI 10.1371/journal.pone.0159456

    View details for PubMedID 27486863

    View details for PubMedCentralID PMC4972316

  • Observing planar cell polarity in multiciliated mouse airway epithelial cells. Methods in cell biology Vladar, E. K., Lee, Y. L., Stearns, T., Axelrod, J. D. 2015; 127: 37-54


    The concerted movement of cilia propels inhaled contaminants out of the lungs, safeguarding the respiratory system from toxins, pathogens, pollutants, and allergens. Motile cilia on the multiciliated cells (MCCs) of the airway epithelium are physically oriented along the tissue axis for directional motility, which depends on the planar cell polarity (PCP) signaling pathway. The MCCs of the mouse respiratory epithelium have emerged as an important model for the study of motile ciliogenesis and the PCP signaling mechanism. Unlike other motile ciliated or planar polarized tissues, airway epithelial cells are relatively easily accessible and primary cultures faithfully model many of the essential features of the in vivo tissue. There is growing interest in understanding how cells acquire and polarize motile cilia due to the impact of mucociliary clearance on respiratory health. Here, we present methods for observing and quantifying the planar polarized orientation of motile cilia both in vivo and in primary culture airway epithelial cells. We describe how to acquire and evaluate electron and light microscopy images of ciliary ultrastructural features that reveal planar polarized orientation. Furthermore, we describe the immunofluorescence localization of PCP pathway components as a simple readout for airway epithelial planar polarization and ciliary orientation. These methods can be adapted to observe ciliary orientation in other multi- and monociliated cells and to detect PCP pathway activity in any tissue or cell type.

    View details for DOI 10.1016/bs.mcb.2015.01.016

    View details for PubMedID 25837385

  • Coordinating cell polarity: heading in the right direction? DEVELOPMENT Axelrod, J. D., Bergmann, D. C. 2014; 141 (17): 3298-3302


    A diverse group of researchers working on both plant and animal systems met at a Company of Biologists workshop to discuss 'Coordinating Cell Polarity'. The meeting included considerable free discussion as well as presentations exploring the ways that groups of cells in these various systems achieve coordinated cell polarity. Here, we discuss commonalities, differences and themes that emerged from these sessions that will serve to inform ongoing studies.

    View details for DOI 10.1242/dev.111484

    View details for Web of Science ID 000341305900003

  • Coordinating cell polarity: heading in the right direction? Development (Cambridge, England) Axelrod, J. D., Bergmann, D. C. 2014; 141 (17): 3298-302


    A diverse group of researchers working on both plant and animal systems met at a Company of Biologists workshop to discuss 'Coordinating Cell Polarity'. The meeting included considerable free discussion as well as presentations exploring the ways that groups of cells in these various systems achieve coordinated cell polarity. Here, we discuss commonalities, differences and themes that emerged from these sessions that will serve to inform ongoing studies.

    View details for DOI 10.1242/dev.111484

    View details for PubMedID 25139852

  • prickle modulates microtubule polarity and axonal transport to ameliorate seizures in flies. Proceedings of the National Academy of Sciences of the United States of America Ehaideb, S. N., Iyengar, A., Ueda, A., Iacobucci, G. J., Cranston, C., Bassuk, A. G., Gubb, D., Axelrod, J. D., Gunawardena, S., Wu, C., Manak, J. R. 2014; 111 (30): 11187-11192


    Recent analyses in flies, mice, zebrafish, and humans showed that mutations in prickle orthologs result in epileptic phenotypes, although the mechanism responsible for generating the seizures was unknown. Here, we show that Prickle organizes microtubule polarity and affects their growth dynamics in axons of Drosophila neurons, which in turn influences both anterograde and retrograde vesicle transport. We also show that enhancement of the anterograde transport mechanism is the cause of the seizure phenotype in flies, which can be suppressed by reducing the level of either of two Kinesin motor proteins responsible for anterograde vesicle transport. Additionally, we show that seizure-prone prickle mutant flies have electrophysiological defects similar to other fly mutants used to study seizures, and that merely altering the balance of the two adult prickle isoforms in neurons can predispose flies to seizures. These data reveal a previously unidentified pathway in the pathophysiology of seizure disorders and provide evidence for a more generalized cellular mechanism whereby Prickle mediates polarity by influencing microtubule-mediated transport.

    View details for DOI 10.1073/pnas.1403357111

    View details for PubMedID 25024231

    View details for PubMedCentralID PMC4121842

  • Prickle/spiny-legs isoforms control the polarity of the apical microtubule network in planar cell polarity. Development Olofsson, J., Sharp, K. A., Matis, M., Cho, B., Axelrod, J. D. 2014; 141 (14): 2866-2874


    Microtubules (MTs) are substrates upon which plus- and minus-end directed motors control the directional movement of cargos that are essential for generating cell polarity. Although centrosomal MTs are organized with plus-ends away from the MT organizing center, the regulation of non-centrosomal MT polarity is poorly understood. Increasing evidence supports the model that directional information for planar polarization is derived from the alignment of a parallel apical network of MTs and the directional MT-dependent trafficking of downstream signaling components. The Fat/Dachsous/Four-jointed (Ft/Ds/Fj) signaling system contributes to orienting those MTs. In addition to previously defined functions in promoting asymmetric subcellular localization of 'core' planar cell polarity (PCP) proteins, we find that alternative Prickle (Pk-Sple) protein isoforms control the polarity of this MT network. This function allows the isoforms of Pk-Sple to differentially determine the direction in which asymmetry is established and therefore, ultimately, the direction of tissue polarity. Oppositely oriented signals that are encoded by oppositely oriented Fj and Ds gradients produce the same polarity outcome in different tissues or compartments, and the tissue-specific activity of alternative Pk-Sple protein isoforms has been observed to rectify the interpretation of opposite upstream directional signals. The control of MT polarity, and thus the directionality of apical vesicle traffic, by Pk-Sple provides a mechanism for this rectification.

    View details for DOI 10.1242/dev.105932

    View details for PubMedID 25005476

  • Methods for studying planar cell polarity METHODS Olofsson, J., Axelrod, J. D. 2014; 68 (1): 97-104


    Planar cell polarity (PCP) is the polarity of epithelial cells in the plane orthogonal to the apical-basal axis, and is controlled by a partially defined signaling system. PCP related signaling also plays roles in cell migration, tissue re-organization and stem cell differentiation during embryonic development, and later, in regeneration and repair. Aberrant signaling has been linked to a broad range of pathophysiologies including cancer, developmental defects, and neurological disorders. The deepest mechanistic insights have come from studies of PCP in Drosophila. In this chapter we review tools and methods to study PCP signaling in Drosophila epithelia, where it was found to involve asymmetric protein localization that is coordinated between adjacent cells. Such signaling has been most extensively studied in wing, eye, and abdomen, but also in other tissues such as leg and notum. In the adult fly, PCP is manifested in the coordinated direction of hairs and bristles, as well as the organization of ommatidia in the eye. The polarity of these structures is preceded by asymmetric localization of PCP signaling proteins at the apical junctions of epithelial cells. Based on genetic and molecular criteria, the proteins that govern PCP can be divided into distinct modules, including the core module, the Fat/Dachsous/Four-jointed (Fat/Ds/Fj) module (often referred to as the 'global' module) as well as tissue specific effector modules. Different tissues and tissue regions differ in their sensitivity to disturbances in the various modules of the PCP signaling system, leading to controversies about the interactions among the modules, and emphasizing the value of studying PCP in multiple contexts. Here, we review methods including those generally applicable, as well as some that are selectively useful for analyses of PCP in eye (including eye discs), wing (including wing discs), pupal and adult abdomen, and the cuticle of larvae and embryos.

    View details for DOI 10.1016/j.ymeth.2014.03.017

    View details for Web of Science ID 000337551300011

    View details for PubMedID 24680701

    View details for PubMedCentralID PMC4048770

  • Myb promotes centriole amplification and later steps of the multiciliogenesis program DEVELOPMENT Tan, F. E., Vladar, E. K., Ma, L., Fuentealba, L. C., Hoh, R., Espinoza, F. H., Axelrod, J. D., Alvarez-Buylla, A., Stearns, T., Kintner, C., Krasnow, M. A. 2013; 140 (20): 4277-4286


    The transcriptional control of primary cilium formation and ciliary motility are beginning to be understood, but little is known about the transcriptional programs that control cilium number and other structural and functional specializations. One of the most intriguing ciliary specializations occurs in multiciliated cells (MCCs), which amplify their centrioles to nucleate hundreds of cilia per cell, instead of the usual monocilium. Here we report that the transcription factor MYB, which promotes S phase and drives cycling of a variety of progenitor cells, is expressed in postmitotic epithelial cells of the mouse airways and ependyma destined to become MCCs. MYB is expressed early in multiciliogenesis, as progenitors exit the cell cycle and amplify their centrioles, then switches off as MCCs mature. Conditional inactivation of Myb in the developing airways blocks or delays centriole amplification and expression of FOXJ1, a transcription factor that controls centriole docking and ciliary motility, and airways fail to become fully ciliated. We provide evidence that MYB acts in a conserved pathway downstream of Notch signaling and multicilin, a protein related to the S-phase regulator geminin, and upstream of FOXJ1. MYB can activate endogenous Foxj1 expression and stimulate a cotransfected Foxj1 reporter in heterologous cells, and it can drive the complete multiciliogenesis program in Xenopus embryonic epidermis. We conclude that MYB has an early, crucial and conserved role in multiciliogenesis, and propose that it promotes a novel S-like phase in which centriole amplification occurs uncoupled from DNA synthesis, and then drives later steps of multiciliogenesis through induction of Foxj1.

    View details for DOI 10.1242/dev.094102

    View details for Web of Science ID 000325153200017

    View details for PubMedID 24048590

    View details for PubMedCentralID PMC3787764

  • Regulation of PCP by the Fat signaling pathway GENES & DEVELOPMENT Matis, M., Axelrod, J. D. 2013; 27 (20): 2207-2220


    Planar cell polarity (PCP) in epithelia, orthogonal to the apical-basal axis, is essential for numerous developmental events and physiological functions. Drosophila model systems have been at the forefront of studies revealing insights into mechanisms regulating PCP and have revealed distinct signaling modules. One of these, involving the atypical cadherins Fat and Dachsous and the ectokinase Four-jointed, appears to link the direction of cell polarization to the tissue axes. We discuss models for the function of this signaling module as well as several unanswered questions that may guide future investigations.

    View details for DOI 10.1101/gad.228098.113

    View details for Web of Science ID 000325932500003

    View details for PubMedID 24142873

    View details for PubMedCentralID PMC3814641

  • Myb promotes centriole amplification and later steps of the multiciliogenesis program. Development Tan, F. E., Vladar, E. K., Ma, L., Fuentealba, L. C., Hoh, R., Espinoza, F. H., Axelrod, J. D., Alvarez-Buylla, A., Stearns, T., Kintner, C., Krasnow, M. A. 2013; 140 (20): 4277-4286


    The transcriptional control of primary cilium formation and ciliary motility are beginning to be understood, but little is known about the transcriptional programs that control cilium number and other structural and functional specializations. One of the most intriguing ciliary specializations occurs in multiciliated cells (MCCs), which amplify their centrioles to nucleate hundreds of cilia per cell, instead of the usual monocilium. Here we report that the transcription factor MYB, which promotes S phase and drives cycling of a variety of progenitor cells, is expressed in postmitotic epithelial cells of the mouse airways and ependyma destined to become MCCs. MYB is expressed early in multiciliogenesis, as progenitors exit the cell cycle and amplify their centrioles, then switches off as MCCs mature. Conditional inactivation of Myb in the developing airways blocks or delays centriole amplification and expression of FOXJ1, a transcription factor that controls centriole docking and ciliary motility, and airways fail to become fully ciliated. We provide evidence that MYB acts in a conserved pathway downstream of Notch signaling and multicilin, a protein related to the S-phase regulator geminin, and upstream of FOXJ1. MYB can activate endogenous Foxj1 expression and stimulate a cotransfected Foxj1 reporter in heterologous cells, and it can drive the complete multiciliogenesis program in Xenopus embryonic epidermis. We conclude that MYB has an early, crucial and conserved role in multiciliogenesis, and propose that it promotes a novel S-like phase in which centriole amplification occurs uncoupled from DNA synthesis, and then drives later steps of multiciliogenesis through induction of Foxj1.

    View details for DOI 10.1242/dev.094102

    View details for PubMedID 24048590

  • Absolute requirement of cholesterol binding for Hedgehog gradient formation in Drosophila. Biology open Ducuing, A., Mollereau, B., Axelrod, J. D., Vincent, S. 2013; 2 (6): 596-604


    How morphogen gradients are shaped is a major question in developmental biology, but remains poorly understood. Hedgehog (Hh) is a locally secreted ligand that reaches cells at a distance and acts as a morphogen to pattern the Drosophila wing and the vertebrate neural tube. The proper patterning of both structures relies on the precise control over the slope of Hh activity gradient. A number of hypotheses have been proposed to explain Hh movement and hence graded activity of Hh. A crux to all these models is that the covalent binding of cholesterol to Hh N-terminus is essential to achieve the correct slope of the activity gradient. Still, the behavior of cholesterol-free Hh (Hh-N) remains controversial: cholesterol has been shown to either increase or restrict Hh range depending on the experimental setting. Here, in fly embryos and wing imaginal discs, we show that cholesterol-free Hh diffuses at a long-range. This unrestricted diffusion of cholesterol-free Hh leads to an absence of gradient while Hh signaling strength remains uncompromised. These data support a model where cholesterol addition restricts Hh diffusion and can transform a leveled signaling activity into a gradient. In addition, our data indicate that the receptor Patched is not able to sequester cholesterol-free Hh. We propose that a morphogen gradient does not necessarily stem from the active transfer of a poorly diffusing molecule, but can be achieved by the restriction of a highly diffusible ligand.

    View details for DOI 10.1242/bio.20134952

    View details for PubMedID 23789110

  • Microtubules Enable the Planar Cell Polarity of Airway Cilia CURRENT BIOLOGY Vladar, E. K., Bayly, R. D., Sangoram, A. M., Scott, M. P., Axelrod, J. D. 2012; 22 (23): 2203-2212


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

    View details for DOI 10.1016/j.cub.2012.09.046

    View details for Web of Science ID 000312115300016

    View details for PubMedID 23122850

    View details for PubMedCentralID PMC3518597

  • Remodeling a Tissue: Subtraction Adds Insight SCIENCE SIGNALING Axelrod, J. D. 2012; 5 (252)


    Sculpting a body plan requires both patterning of gene expression and translating that pattern into morphogenesis. Developmental biologists have made remarkable strides in understanding gene expression patterning, but despite a long history of fascination with the mechanics of morphogenesis, knowledge of how patterned gene expression drives the emergence of even simple shapes and forms has grown at a slower pace. The successful merging of approaches from cell biology, developmental biology, imaging, engineering, and mathematical and computational sciences is now accelerating progress toward a fuller and better integrated understanding of the forces shaping morphogenesis.

    View details for DOI 10.1126/scisignal.2003620

    View details for Web of Science ID 000311749700003

    View details for PubMedID 23193158

  • A Mathematical Model to Study the Dynamics of Epithelial Cellular Networks IEEE-ACM TRANSACTIONS ON COMPUTATIONAL BIOLOGY AND BIOINFORMATICS Abate, A., Vincent, S., Dobbe, R., Silletti, A., Master, N., Axelrod, J. D., Tomlin, C. J. 2012; 9 (6): 1607-1620


    Epithelia are sheets of connected cells that are essential across the animal kingdom. Experimental observations suggest that the dynamical behavior of many single-layered epithelial tissues has strong analogies with that of specific mechanical systems, namely large networks consisting of point masses connected through spring-damper elements and undergoing the influence of active and dissipating forces. Based on this analogy, this work develops a modeling framework to enable the study of the mechanical properties and of the dynamic behavior of large epithelial cellular networks. The model is built first by creating a network topology that is extracted from the actual cellular geometry as obtained from experiments, then by associating a mechanical structure and dynamics to the network via spring-damper elements. This scalable approach enables running simulations of large network dynamics: the derived modeling framework in particular is predisposed to be tailored to study general dynamics (for example, morphogenesis) of various classes of single-layered epithelial cellular networks. In this contribution, we test the model on a case study of the dorsal epithelium of the Drosophila melanogaster embryo during early dorsal closure (and, less conspicuously, germband retraction).

    View details for DOI 10.1109/TCBB.2012.126

    View details for Web of Science ID 000312558400006

    View details for PubMedID 23221083

    View details for PubMedCentralID PMC3558995

  • Planar Polarized Protrusions Break the Symmetry of EGFR Signaling during Drosophila Bract Cell Fate Induction DEVELOPMENTAL CELL Peng, Y., Han, C., Axelrod, J. D. 2012; 23 (3): 507-518


    Secreted signaling molecules typically float in the outer leaflet of the plasma membrane or freely diffuse away from the signaling cell, suggesting that a signal should be sensed equally by all neighboring cells. However, we demonstrate that Spitz (Spi)-mediated epidermal growth factor receptor (EGFR) signaling is spatially biased to selectively determine the induction of a single bract cell on the proximal side of each mechanosensory organ on the Drosophila leg. Dynamic and oriented cellular protrusions emanating from the socket cell, the source of Spi, robustly favor the Spi/EGFR signaling response in a particular cell among equally competent neighbors. We propose that these protrusive structures enhance signaling by increasing contact between the signaling and responding cells. The planar polarized direction of the protrusions determines the direction of the signaling outcome. This asymmetric cell signaling serves as a developmental mechanism to generate spatially patterned cell fates.

    View details for DOI 10.1016/j.devcel.2012.07.016

    View details for Web of Science ID 000308776400009

    View details for PubMedID 22921201

    View details for PubMedCentralID PMC3482102

  • A universal analysis tool for the detection of asymmetric signal distribution in microscopic images DEVELOPMENTAL DYNAMICS Matis, M., Axelrod, J. D., Galic, M. 2012; 241 (8): 1301-1309


    Polarization of tissue is achieved by asymmetric distribution of proteins and organelles within individual cells. However, existing quantitative assays to measure this asymmetry in an automated and unbiased manner suffer from significant limitations.Here, we report a new way to assess protein and organelle localization in tissue based on correlative fluorescence analysis. As a proof of principle, we successfully characterized planar cell polarity dependent asymmetry in developing Drosophila melanogaster tissues on the single cell level using fluorescence cross-correlation.Systematic modulation of signal strength and distribution show that fluorescence cross-correlation reliably detects asymmetry over a broad parameter space. The novel method described here produces robust, rapid, and unbiased measurement of biometrical properties of cell components in live tissue that is readily applicable in other model systems.

    View details for DOI 10.1002/dvdy.23818

    View details for Web of Science ID 000306490100007

    View details for PubMedID 22689329

    View details for PubMedCentralID PMC3469164

  • Nuclear localization of Prickle2 is required to establish cell polarity during early mouse embryogenesis DEVELOPMENTAL BIOLOGY Tao, H., Inoue, K., Kiyonari, H., Bassuk, A. G., Axelrod, J. D., Sasaki, H., Aizawa, S., Ueno, N. 2012; 364 (2): 138-148


    The establishment of trophectoderm (TE) manifests as the formation of epithelium, and is dependent on many structural and regulatory components that are commonly found and function in many epithelial tissues. However, the mechanism of TE formation is currently not well understood. Prickle1 (Pk1), a core component of the planar cell polarity (PCP) pathway, is essential for epiblast polarization before gastrulation, yet the roles of Pk family members in early mouse embryogenesis are obscure. Here we found that Pk2(-/-) embryos died at E3.0-3.5 without forming the blastocyst cavity and not maintained epithelial integrity of TE. These phenotypes were due to loss of the apical-basal (AB) polarity that underlies the asymmetric redistribution of microtubule networks and proper accumulation of AB polarity components on each membrane during compaction. In addition, we found GTP-bound active form of nuclear RhoA was decreased in Pk2(-/-) embryos during compaction. We further show that the first cell fate decision was disrupted in Pk2(-/-) embryos. Interestingly, Pk2 localized to the nucleus from the 2-cell to around the 16-cell stage despite its cytoplasmic function previously reported. Inhibiting farnesylation blocked Pk2's nuclear localization and disrupted AB cell polarity, suggesting that Pk2 farnesylation is essential for its nuclear localization and function. The cell polarity phenotype was efficiently rescued by nuclear but not cytoplasmic Pk2, demonstrating the nuclear localization of Pk2 is critical for its function.

    View details for DOI 10.1016/j.ydbio.2012.01.025

    View details for Web of Science ID 000301827500005

    View details for PubMedID 22333836

    View details for PubMedCentralID PMC3299875

  • Multicilin promotes centriole assembly and ciliogenesis during multiciliate cell differentiation NATURE CELL BIOLOGY Stubbs, J. L., Vladar, E. K., Axelrod, J. D., Kintner, C. 2012; 14 (2): 140-147


    Multiciliate cells function prominently in the respiratory system, brain ependyma and female reproductive tract to produce vigorous fluid flow along epithelial surfaces. These specialized cells form during development when epithelial progenitors undergo an unusual form of ciliogenesis, in which they assemble and project hundreds of motile cilia. Notch inhibits multiciliate cell formation in diverse epithelia, but how progenitors overcome lateral inhibition and initiate multiciliate cell differentiation is unknown. Here we identify a coiled-coil protein, termed multicilin, which is regulated by Notch and highly expressed in developing epithelia where multiciliate cells form. Inhibiting multicilin function specifically blocks multiciliate cell formation in Xenopus skin and kidney, whereas ectopic expression induces the differentiation of multiciliate cells in ectopic locations. Multicilin localizes to the nucleus, where it directly activates the expression of genes required for multiciliate cell formation, including foxj1 and genes mediating centriole assembly. Multicilin is also necessary and sufficient to promote multiciliate cell differentiation in mouse airway epithelial cultures. These findings indicate that multicilin initiates multiciliate cell differentiation in diverse tissues, by coordinately promoting the transcriptional changes required for motile ciliogenesis and centriole assembly.

    View details for DOI 10.1038/ncb2406

    View details for Web of Science ID 000300332500008

    View details for PubMedID 22231168

    View details for PubMedCentralID PMC3329891

  • Asymmetric Protein Localization in Planar Cell Polarity: Mechanisms, Puzzles, and Challenges PLANAR CELL POLARITY DURING DEVELOPMENT Peng, Y., Axelrod, J. D. 2012; 101: 33-53


    The polarization of epithelial cells along an axis orthogonal to their apical-basal axis is increasingly recognized for roles in a variety of developmental events and physiological functions. While now studied in many model organisms, mechanistic understanding is rooted in intensive investigations of planar cell polarity (PCP) in Drosophila. Consensus has emerged that two molecular modules, referred to here as the global and core modules, operate upstream of effector proteins to produce morphological PCP. Proteins of the core module develop subcellular asymmetry, accumulating in two groups on opposite sides of cells, consistent with proposed functions in producing cell polarity and in communicating that polarity between neighboring cells. Less clear are the molecular and cell biological mechanisms underlying core module function in the generation and communication of subcellular asymmetry and the relationship between the global and the core modules. In this review, we discuss these two unresolved questions, highlighting important studies and potentially enlightening avenues for further investigation. It is likely that results from Drosophila will continue to inform our views of the growing list of examples of PCP in vertebrate systems.

    View details for DOI 10.1016/B978-0-12-394592-1.00002-8

    View details for Web of Science ID 000314133400003

    View details for PubMedID 23140624

  • Modeling the control of planar cell polarity WILEY INTERDISCIPLINARY REVIEWS-SYSTEMS BIOLOGY AND MEDICINE Axelrod, J. D., Tomlin, C. J. 2011; 3 (5): 588-605


    A growing list of medically important developmental defects and disease mechanisms can be traced to disruption of the planar cell polarity (PCP) pathway. The PCP system polarizes cells in epithelial sheets along an axis orthogonal to their apical-basal axis. Studies in the fruitfly, Drosophila, have suggested that components of the PCP signaling system function in distinct modules, and that these modules and the effector systems with which they interact function together to produce emergent patterns. Experimental methods allow the manipulation of individual PCP signaling molecules in specified groups of cells; these interventions not only perturb the polarization of the targeted cells at a subcellular level, but also perturb patterns of polarity at the multicellular level, often affecting nearby cells in characteristic ways. These kinds of experiments should, in principle, allow one to infer the architecture of the PCP signaling system, but the relationships between molecular interactions and tissue-level pattern are sufficiently complex that they defy intuitive understanding. Mathematical modeling has been an important tool to address these problems. This article explores the emergence of a local signaling hypothesis, and describes how a local intercellular signal, coupled with a directional cue, can give rise to global pattern. We will discuss the critical role mathematical modeling has played in guiding and interpreting experimental results, and speculate about future roles for mathematical modeling of PCP. Mathematical models at varying levels of inhibition have and are expected to continue contributing in distinct ways to understanding the regulation of PCP signaling.

    View details for DOI 10.1002/wsbm.138

    View details for Web of Science ID 000294351300006

    View details for PubMedID 21755606

  • Pointing in the right direction: new developments in the field of planar cell polarity NATURE REVIEWS GENETICS Bayly, R., Axelrod, J. D. 2011; 12 (6): 385-391


    Planar cell polarity (PCP) is observed in an array of developmental processes that involve collective cell movement and tissue organization, and its disruption can lead to severe developmental defects. Recent studies in flies and vertebrates have identified new functions for PCP as well as new signalling components, and have proposed new mechanistic models. However, despite this progress, the search to simplify principles of understanding continues and important mechanistic uncertainties still pose formidable challenges.

    View details for DOI 10.1038/nrg2956

    View details for Web of Science ID 000290714000009

    View details for PubMedID 21502960

  • Versatile spectral methods for point set matching PATTERN RECOGNITION LETTERS Silletti, A., Abate, A., Axelrod, J. D., Tomlin, C. J. 2011; 32 (5): 731-739
  • Mutations in Prickle Orthologs Cause Seizures in Flies, Mice, and Humans AMERICAN JOURNAL OF HUMAN GENETICS Tao, H., Manak, J. R., Sowers, L., Mei, X., Kiyonari, H., Abe, T., Dandaleh, N. S., Yang, T., Wu, S., Chen, S., Fox, M. H., Gurnett, C., Montine, T., Bird, T., Shaffer, L. G., Rosenfeld, J. A., McConne, J., Madan-Khetarpal, S., Berry-Kravis, E., Griesbach, H., Saneto, R. P., Scott, M. P., Antic, D., Reed, J., Boland, R., Ehaideb, S. N., El-Shanti, H., Mahajan, V. B., Ferguson, P. J., Axelrod, J. D., Lehesjoki, A., Fritzsch, B., Slusarski, D. C., Wemmie, J., Ueno, N., Bassuk, A. G. 2011; 88 (2): 138-149


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

    View details for DOI 10.1016/j.ajhg.2010.12.012

    View details for Web of Science ID 000287684100002

    View details for PubMedID 21276947

    View details for PubMedCentralID PMC3035715

  • STED Super-resolution Microscopy in Drosophila Tissue and in Mammalian Cells. Proceedings of SPIE--the International Society for Optical Engineering Lau, L., Lee, Y. L., Matis, M., Axelrod, J., Stearns, T., Moerner, W. E. 2011; 7910


    Far-field super-resolution microscopy is a rapidly emerging method that is opening up opportunities for biological imaging beyond the optical diffraction limit. We have implemented a Stimulated Emission Depletion (STED) microscope to image single dye, cell, and tissue samples with 50-80 nm resolution. First, we compare the STED performance imaging single molecules of several common dyes and report a novel STED dye. Then we apply STED to image planar cell polarity protein complexes in intact fixed Drosophila tissue for the first time. Finally, we present a preliminary study of the centrosomal protein Cep164 in mammalian cells. Our images suggest that Cep164 is arranged in a nine-fold symmetric pattern around the centriole, consistent with findings suggested by cryoelectron tomography. Our work demonstrates that STED microscopy can be used for superresolution imaging in intact tissue and provides ultrastructural information in biological samples as an alternative to immuno-electron microscopy.

    View details for PubMedID 23447411

  • STED Super-resolution Microscopy in Drosophila Tissue and in Mammalian Cells Conference on Reporters, Markers, Dyes, Nanoparticles, and Molecular Probes for Biomedical Applications III Lau, L., Lee, Y. L., Matis, M., Axelrod, J., Stearns, T., Moerner, W. E. SPIE-INT SOC OPTICAL ENGINEERING. 2011

    View details for DOI 10.1117/12.881221

    View details for Web of Science ID 000297729300032

  • Delivering the Lateral Inhibition Punchline: It's All About the Timing SCIENCE SIGNALING Axelrod, J. D. 2010; 3 (145)


    Experimental and theoretical biologists have long been fascinated with the emergence of self-organizing patterns in developing organisms, and much attention has focused on Notch-mediated lateral inhibition. Within sheets of cells that may adopt either of two possible cell fates, lateral inhibition establishes patterns through the activity of a negative intercellular feedback loop involving the receptor, Notch, and its ligands Delta or Serrate. Despite a long history of intensive study in Drosophila, where the mechanism was first described, as well as in other organisms, new work continues to yield important insights. Mathematical modeling, combined with biological analyses, has now shed light on two features of the process: how antagonistic and activating ligand-receptor interactions work together to accelerate inhibition and ensure fidelity, and how filopodial dynamics contribute to the observed pattern refinement and spacing.

    View details for DOI 10.1126/scisignal.3145pe38

    View details for Web of Science ID 000283733600002

    View details for PubMedID 20978236

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


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

    View details for DOI 10.1371/journal.pone.0008999

    View details for Web of Science ID 000274207200003

    View details for PubMedID 20126399

    View details for PubMedCentralID PMC2814853

  • Progress and challenges in understanding planar cell polarity signaling SEMINARS IN CELL & DEVELOPMENTAL BIOLOGY Axelrod, J. D. 2009; 20 (8): 964-971


    During development, epithelial cells in some tissues acquire a polarity orthogonal to their apical-basal axis. This polarity, referred to as planar cell polarity (PCP), or tissue polarity, is essential for the normal physiological function of many epithelia. Early studies of PCP focused on insect epithelia (Lawrence, 1966 [1]), and the earliest genetic analyses were carried out in Drosophila (Held et al., 1986; Gubb and Garcia-Bellido, 1982 [2,3]). Indeed, most of our mechanistic understanding of PCP derives from the ongoing use of Drosophila as a model system. However, a range of medically important developmental defects and physiological processes are under the control of PCP mechanisms that appear to be at least partially conserved, driving considerable interest in studying PCP both in Drosophila and in vertebrate model systems. Here, I present a model of the PCP signaling mechanism based on studies in Drosophila. I highlight two areas in which our understanding is deficient, and which lead to current confusion in the literature. Future studies that shed light on these areas will substantially enhance our understanding of the fascinating yet challenging problem of understanding the mechanisms that generate PCP.

    View details for DOI 10.1016/j.semcdb.2009.08.001

    View details for Web of Science ID 000274300800012

    View details for PubMedID 19665570

  • Studies of epithelial PCP. Seminars in cell & developmental biology Axelrod, J. D. 2009; 20 (8): 956-?

    View details for DOI 10.1016/j.semcdb.2009.08.002

    View details for PubMedID 19665569

  • Planar Cell Polarity Signaling: The Developing Cell's Compass COLD SPRING HARBOR PERSPECTIVES IN BIOLOGY Vladar, E. K., Antic, D., Axelrod, J. D. 2009; 1 (3)


    Cells of many tissues acquire cellular asymmetry to execute their physiologic functions. The planar cell polarity system, first characterized in Drosophila, is important for many of these events. Studies in Drosophila suggest that an upstream system breaks cellular symmetry by converting tissue gradients to subcellular asymmetry, whereas a downstream system amplifies subcellular asymmetry and communicates polarity between cells. In this review, we discuss apparent similarities and differences in the mechanism that controls PCP as it has been adapted to a broad variety of morphological cellular asymmetries in various organisms.

    View details for DOI 10.1101/cshperspect.a002964

    View details for Web of Science ID 000279879100006

    View details for PubMedID 20066108

    View details for PubMedCentralID PMC2773631

  • Bone morphogenetic protein 2 induces pulmonary angiogenesis via Wnt-beta-catenin and Wnt-RhoA-Rac1 pathways JOURNAL OF CELL BIOLOGY Perez, V. A., Alastalo, T., Wu, J. C., Axelrod, J. D., Cooke, J. P., Amieva, M., Rabinovitch, M. 2009; 184 (1): 83-99


    Mutations in bone morphogenetic protein (BMP) receptor II (BMPRII) are associated with pulmonary artery endothelial cell (PAEC) apoptosis and the loss of small vessels seen in idiopathic pulmonary arterial hypertension. Given the low penetrance of BMPRII mutations, abnormalities in other converging signaling pathways may be necessary for disease development. We hypothesized that BMPRII supports normal PAEC function by recruiting Wingless (Wnt) signaling pathways to promote proliferation, survival, and motility. In this study, we report that BMP-2, via BMPRII-mediated inhibition of GSK3-beta, induces beta-catenin (beta-C) accumulation and transcriptional activity necessary for PAEC survival and proliferation. At the same time, BMP-2 mediates phosphorylated Smad1 (pSmad1) or, with loss of BMPRII, pSmad3-dependent recruitment of Disheveled (Dvl) to promote RhoA-Rac1 signaling necessary for motility. Finally, using an angiogenesis assay in severe combined immunodeficient mice, we demonstrate that both beta-C- and Dvl-mediated RhoA-Rac1 activation are necessary for vascular growth in vivo. These findings suggest that the recruitment of both canonical and noncanonical Wnt pathways is required in BMP-2-mediated angiogenesis.

    View details for DOI 10.1083/jcb.200806049

    View details for Web of Science ID 000262867000010

    View details for PubMedID 19139264

    View details for PubMedCentralID PMC2615088

  • Cell packing influences planar cell polarity signaling PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Ma, D., Amonlirdviman, K., Raffard, R. L., Abate, A., Tomlin, C. J., Axelrod, J. D. 2008; 105 (48): 18800-18805


    Some epithelial cells display asymmetry along an axis orthogonal to the apical-basal axis, referred to as planar cell polarity (PCP). A Frizzled-mediated feedback loop coordinates PCP between neighboring cells, and the cadherin Fat transduces a global directional cue that orients PCP with respect to the tissue axes. The feedback loop can propagate polarity across clones of cells that lack the global directional signal, although this polarity propagation is error prone. Here, we show that, in the Drosophila wing, a combination of cell geometry and nonautonomous signaling at clone boundaries determines the correct or incorrect polarity propagation in clones that lack Fat mediated global directional information. Pattern elements, such as veins, and sporadic occurrences of irregular geometry are obstacles to polarity propagation. Hence, in the wild type, broad distribution of the global directional cue combines with a local feedback mechanism to overcome irregularities in cell packing geometry during PCP signaling.

    View details for DOI 10.1073/pnas.0808868105

    View details for Web of Science ID 000261489100036

    View details for PubMedID 19022903

    View details for PubMedCentralID PMC2585485

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


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

    View details for DOI 10.1016/j.ajhg.2008.10.003

    View details for Web of Science ID 000261006900003

    View details for PubMedID 18976727

    View details for PubMedCentralID PMC2668041

  • Dishevelled links basal body docking and orientation in ciliated epithelial cells TRENDS IN CELL BIOLOGY Vladar, E. K., Axelrod, J. D. 2008; 18 (11): 517-520


    Some epithelia contain cells with multiple motile cilia that beat in a concerted manner. New tools and experimental systems have facilitated molecular studies of cilium biogenesis and the coordinated planar polarization of cilia that leads to their concerted motility. A recent elegant study using embryonic frog epidermis demonstrates that Dishevelled, a key regulator of both the Wnt-beta-catenin and planar cell polarity pathways, controls both the docking and planar polarization of ciliary basal bodies.

    View details for DOI 10.1016/j.tcb.2008.08.004

    View details for Web of Science ID 000261074400001

    View details for PubMedID 18819800

  • Bad hair days for mouse PCP mutants NATURE CELL BIOLOGY Axelrod, J. D. 2008; 10 (11): 1251-1253

    View details for Web of Science ID 000260586700006

    View details for PubMedID 18978834

  • Hedgehog and Wingless stabilize but do not induce cell fate during Drosophila dorsal embryonic epidermal patterning DEVELOPMENT Vincent, S., Perrimon, N., Axelrod, J. D. 2008; 135 (16): 2767-2775


    A fundamental concept in development is that secreted molecules such as Wingless (Wg) and Hedgehog (Hh) generate pattern by inducing cell fate. By following markers of cellular identity posterior to the Wg- and Hh-expressing cells in the Drosophila dorsal embryonic epidermis, we provide evidence that neither Wg nor Hh specifies the identity of the cell types they pattern. Rather, they maintain pre-existing cellular identities that are otherwise unstable and progress stepwise towards a default fate. Wg and Hh therefore generate pattern by inhibiting specific switches in cell identity, showing that the specification and the patterning of a given cell are uncoupled. Sequential binary decisions without induction of cell identity give rise to both the groove cells and their posterior neighbors. The combination of independent progression of cell identity and arrest of progression by signals facilitates accurate patterning of an extremely plastic developing epidermis.

    View details for DOI 10.1242/dev.017814

    View details for Web of Science ID 000257922600010

    View details for PubMedID 18614578

    View details for PubMedCentralID PMC2585068

  • An adjoint-based parameter identification algorithm applied to planar cell polarity signaling IEEE TRANSACTIONS ON AUTOMATIC CONTROL Raffard, R. L., Amonlirdviman, K., Axelrod, J. D., Tomlin, C. J. 2008: 109-121
  • An adjoint-based parameter identification algorithm applied to planar cell polarity signaling IEEE TRANSACTIONS ON CIRCUITS AND SYSTEMS I-REGULAR PAPERS Raffard, R. L., Amonlirdviman, K., Axelrod, J. D., Tomlin, C. J. 2008: 109-121
  • Basal bodies, kinocilia and planar cell polarity NATURE GENETICS Axelrod, J. D. 2008; 40 (1): 10-11

    View details for DOI 10.1038/ng0108-10

    View details for Web of Science ID 000252118600005

    View details for PubMedID 18163128

  • Biology by numbers: mathematical modelling in developmental biology NATURE REVIEWS GENETICS Tomlin, C. J., Axelrod, J. D. 2007; 8 (5): 331-340


    In recent years, mathematical modelling of developmental processes has earned new respect. Not only have mathematical models been used to validate hypotheses made from experimental data, but designing and testing these models has led to testable experimental predictions. There are now impressive cases in which mathematical models have provided fresh insight into biological systems, by suggesting, for example, how connections between local interactions among system components relate to their wider biological effects. By examining three developmental processes and corresponding mathematical models, this Review addresses the potential of mathematical modelling to help understand development.

    View details for DOI 10.1038/nrg2098

    View details for Web of Science ID 000245906500013

    View details for PubMedID 17440530

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


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

    View details for DOI 10.1523/JNEUROSCI.5151-06.2007

    View details for Web of Science ID 000245103700010

    View details for PubMedID 17376975

  • The Drosophila casein kinase I epsilon/delta Discs overgrown promotes cell survival via activation of DIAP1 expression DEVELOPMENTAL BIOLOGY Guan, J., Li, H., Rogulja, A., Axelrod, J. D., Cadigan, K. M. 2007; 303 (1): 16-28


    The proper number of cells in developing tissues is achieved by coordinating cell division with apoptosis. In Drosophila, the adult wing is derived from wing imaginal discs, which undergo a period of growth and proliferation during larval stages without much programmed cell death. In this report, we demonstrate that the Drosophila casein kinase Iepsilon/delta, known as Discs overgrown (Dco), is required for maintaining this low level of apoptosis. Expression of dco can suppress the apoptotic activity of Head involution defective (Hid) in the developing eye. Loss of dco in the wing disc results in a dramatic reduction in expression of the caspase inhibitor DIAP1 and a concomitant activation of caspases. The regulation of DIAP1 by Dco occurs by a post-transcriptional mechanism that is independent of hid. Mutant clones of dco are considerably smaller than controls even when apoptosis is inhibited, suggesting that Dco promotes cell division/growth in addition to its role in cell survival. The dco phenotype cannot be explained by defects Wingless (Wg) signaling. We propose that Dco coordinates tissue size by stimulating cell division/growth and blocking apoptosis via activation of DIAP1 expression.

    View details for DOI 10.1016/j.ydbio.2006.10.028

    View details for Web of Science ID 000244542800002

    View details for PubMedID 17134692

    View details for PubMedCentralID PMC2892850

  • Cell shape in proliferating epithelia: A multifaceted problem CELL Axelrod, J. D. 2006; 126 (4): 643-645


    A specific and unexpected distribution pattern of polygonal cell shapes in proliferating epithelia is revealed in a recent study that combines mathematical modeling with experimental data (Gibson et. al., 2006). This pattern is conserved in epithelia from diverse species, suggesting that this distribution is a fundamental property of proliferating epithelial sheets.

    View details for DOI 10.1016/j.cell.2006.07.018

    View details for Web of Science ID 000240276700009

    View details for PubMedID 16923381

  • A WNTer wonderland in Snowbird DEVELOPMENT He, X., Axelrod, J. D. 2006; 133 (14): 2597-2603


    The Keystone Symposium on ;Wnt and beta-catenin signaling in development and disease' was held recently in Snowbird, UT, USA. Organized by Mariann Bienz and Hans Clevers, this meeting covered a wide range of topics, including Wnt protein biogenesis, Wnt receptors and signaling pathways, beta-catenin/Tcf complexes and gene expression, Wnt signaling in development, cancer, stem cell biology and regeneration, and therapeutics that target the Wnt/beta-catenin pathway.

    View details for DOI 10.1242/dev.02452

    View details for Web of Science ID 000238475500001

    View details for PubMedID 16794030

  • Automatic parameter identification via the adjoint method, with application to understanding planar cell polarity 45th IEEE Conference on Decision and Control Raffard, R., Amonlirdviman, K., Axelrod, J. D., Tomlin, C. J. IEEE. 2006: 13–18
  • Understanding biology by reverse engineering the control PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Tomlin, C. J., Axelrod, J. D. 2005; 102 (12): 4219-4220

    View details for DOI 10.1073/pnas.0500276102

    View details for Web of Science ID 000227854800001

    View details for PubMedID 15767568

    View details for PubMedCentralID PMC555517

  • D-2 dopamine receptors colocalize regulator of G-protein signaling 9-2 (RGS9-2) via the RGS9 DEP domain, and RGS9 knock-out mice develop dyskinesias associated with dopamine pathways JOURNAL OF NEUROSCIENCE Kovoor, A., Seyffarth, P., Ebert, J., Barghshoon, S., Chen, C. K., Schwarz, S., Axelrod, J. D., Cheyette, B. N., Simon, M. I., Lester, H. A., Schwarz, J. 2005; 25 (8): 2157-2165


    Regulator of G-protein signaling 9-2 (RGS9-2), a member of the RGS family of G GTPase accelerating proteins, is expressed specifically in the striatum, which participates in antipsychotic-induced tardive dyskinesia and in levodopa-induced dyskinesia. We report that RGS9 knock-out mice develop abnormal involuntary movements when inhibition of dopaminergic transmission is followed by activation of D2-like dopamine receptors (DRs). These abnormal movements resemble drug-induced dyskinesia more closely than other rodent models. Recordings from striatal neurons of these mice establish that activation of D2-like DRs abnormally inhibits glutamate-elicited currents. We show that RGS9-2, via its DEP domain (for Disheveled, EGL-10, Pleckstrin homology), colocalizes with D2DRs when coexpressed in mammalian cells. Recordings from oocytes coexpressing D2DR or the m2 muscarinic receptor and G-protein-gated inward rectifier potassium channels show that RGS9-2, via its DEP domain, preferentially accelerates the termination of D2DR signals. Thus, alterations in RGS9-2 may be a key factor in the pathway leading from D2DRs to the side effects associated with the treatment both of psychoses and Parkinson's disease.

    View details for DOI 10.1523/JNEUROSCI.2840-04.2005

    View details for Web of Science ID 000227211000030

    View details for PubMedID 15728856

  • A second canon: Functions and mechanisms of beta-catenin-independent wnt signaling DEVELOPMENTAL CELL Veeman, M. T., Axelrod, J. D., Moon, R. T. 2003; 5 (3): 367-377


    More is becoming known about so-called noncanonical Wnt pathways that signal independently of beta-catenin. Here we review recent developments in both the functions and mechanisms of noncanonical Wnt signaling. We also discuss some unresolved and vexing questions. How many noncanonical Wnt pathways are there? How extensive are the parallels between Drosophila planar polarization and vertebrate convergence and extension? Last, we will outline some challenges and difficulties we foresee for this exciting but still very young field.

    View details for Web of Science ID 000185309600006

    View details for PubMedID 12967557

  • Fidelity in planar cell polarity signalling NATURE Ma, D., Yang, C. H., McNeill, H., Simon, M. A., Axelrod, J. D. 2003; 421 (6922): 543-547


    The polarity of Drosophila wing hairs displays remarkable fidelity. Each of the approximately 30,000 wing epithelial cells constructs an actin-rich prehair that protrudes from its distal vertex and points distally. The distal location and orientation of the hairs is virtually error free, thus forming a nearly perfect parallel array. This process is controlled by the planar cell polarity signalling pathway. Here we show that interaction between two tiers of the planar cell polarity signalling mechanism results in the observed high fidelity. The first tier, mediated by the cadherin Fat, dictates global orientation by transducing a directional signal to individual cells. The second tier, orchestrated by the 7-pass transmembrane receptor Frizzled, aligns each cell's polarity with that of its neighbours through the action of an intercellular feedback loop, enabling polarity to propagate from cell to cell. We show that all cells need not respond correctly to the presumably subtle signal transmitted by Fat. Subsequent action of the Frizzled feedback loop is sufficient to align all the cells cooperatively. This economical system is therefore highly robust, and produces virtually error-free arrays.

    View details for DOI 10.1038/nature01366

    View details for Web of Science ID 000180670600048

    View details for PubMedID 12540853

  • A three-tiered mechanism for regulation of planar cell polarity SEMINARS IN CELL & DEVELOPMENTAL BIOLOGY Tree, D. R., Ma, D. L., Axelrod, J. D. 2002; 13 (3): 217-224


    Some epithelial cells are polarized along an axis orthogonal to their apical-basal axes. Recent studies in Drosophila lead to the view that three classes of signaling molecules govern the planar cell polarity (PCP) pathway. The first class, or module, functions across whole tissues, providing directional information to individual cells. The second module, apparently shared by all planar polarized tissues, and related to the canonical Wnt signaling pathway, interprets the directional signal to produce subcellular asymmetries. The third modules are tissue specific, acting to translate subcellular asymmetry into the appropriate morphological manifestations in the different cell types.

    View details for DOI 10.1016/S1084-9521(02)00042-3

    View details for Web of Science ID 000177976000009

    View details for PubMedID 12137730

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


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

    View details for Web of Science ID 000175412100012

    View details for PubMedID 12015986

  • Regulation of Frizzled by fat-like cadherins during planar polarity signaling in the Drosophila compound eye CELL Yang, C. H., Axelrod, J. D., Simon, M. A. 2002; 108 (5): 675-688


    Planar polarity is evident in the coordinated orientation of ommatidia in the Drosophila eye. This process requires that the R3 photoreceptor precursor of each ommatidium have a higher level of Frizzled signaling than its neighboring R4 precursor. We show that two cadherin superfamily members, Fat and Dachsous, and the transmembrane/secreted protein Four-jointed play important roles in this process. Our data support a model in which the bias of Frizzled signaling between the R3/R4 precursors results from higher Fat function in the precursor cell closer to the equator, which becomes R3. We also provide evidence that positional information regulating Fat action is provided by graded expression of Dachsous across the eye and the action of Four-jointed, which is expressed in an opposing expression gradient and appears to modulate Dachsous function.

    View details for Web of Science ID 000174314800011

    View details for PubMedID 11893338

  • Coupling planar cell polarity signaling to morphogenesis. TheScientificWorldJournal Axelrod, J. D., McNeill, H. 2002; 2: 434-454


    Epithelial cells and other groups of cells acquire a polarity orthogonal to their apical-basal axes, referred to as Planar Cell Polarity (PCP). The process by which these cells become polarized requires a signaling pathway using Frizzled as a receptor. Responding cells sense cues from their environment that provide directional information, and they translate this information into cellular asymmetry. Most of what is known about PCP derives from studies in the fruit fly, Drosophila. We review what is known about how cells translate an unknown signal into asymmetric cytoskeletal reorganization. We then discuss how the vertebrate processes of convergent extension and cochlear hair-cell development may relate to Drosophila PCP signaling.

    View details for PubMedID 12806028

  • Strabismus comes into focus NATURE CELL BIOLOGY Axelrod, J. D. 2002; 4 (1): E6-E8

    View details for Web of Science ID 000173381500003

    View details for PubMedID 11780132

  • Unipolar membrane association of dishevelled mediates frizzled planar cell polarity signaling GENES & DEVELOPMENT Axelrod, J. D. 2001; 15 (10): 1182-1187


    Drosophila epithelia acquire a planar cell polarity (PCP) orthogonal to their apical-basal axes. Frizzled (Fz) is the receptor for the PCP signal, and Dishevelled (Dsh) transduces the signal. Here, I demonstrate that unipolar relocalization of Dsh to the membrane is required to mediate PCP, but not Wingless (Wg) signaling. Dsh membrane localization reflects the activation of Fz/PCP signaling, revealing that the initially symmetric signal evolves to one that displays unipolar asymmetry, specifying the cells' ultimate polarity. This transition from symmetric to asymmetric Dsh localization requires Dsh function, and reflects an amplification process that generates a steep intracellular activity gradient necessary to determine PCP.

    View details for Web of Science ID 000168930600002

    View details for PubMedID 11358862

  • Drosophila Rho-associated kinase (Drok) links frizzled-mediated planar cell polarity signaling to the actin cytoskeleton CELL Winter, C. G., Wang, B., Ballew, A., Royou, A., Karess, R., Axelrod, J. D., Luo, L. Q. 2001; 105 (1): 81-91


    Frizzled (Fz) and Dishevelled (Dsh) are components of an evolutionarily conserved signaling pathway that regulates planar cell polarity. How this signaling pathway directs asymmetric cytoskeletal reorganization and polarized cell morphology remains unknown. Here, we show that Drosophila Rho-associated kinase (Drok) works downstream of Fz/Dsh to mediate a branch of the planar polarity pathway involved in ommatidial rotation in the eye and in restricting actin bundle formation to a single site in developing wing cells. The primary output of Drok signaling is regulating the phosphorylation of nonmuscle myosin regulatory light chain, and hence the activity of myosin II. Drosophila myosin VIIA, the homolog of the human Usher Syndrome 1B gene, also functions in conjunction with this newly defined portion of the Fz/Dsh signaling pathway to regulate the actin cytoskeleton.

    View details for Web of Science ID 000168063300009

    View details for PubMedID 11301004

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


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

    View details for Web of Science ID 000167821300003

    View details for PubMedID 11274052

  • Frizzled signaling and the developmental control of cell polarity TRENDS IN GENETICS Shulman, J. M., Perrimon, N., Axelrod, J. D. 1998; 14 (11): 452-458


    Within the last three years, Frizzled receptors have risen from obscurity to celebrity status owing to their functional identification as receptors for the ubiquitous family of secreted WNT signaling factors. However, the founding member of the Frizzled family, Drosophila Frizzled (FZ), was cloned almost a decade ago because of its role in regulating cell polarity within the plane of an epithelium. In this review, we consider the role of FZ in this intriguing context. We discuss recent progress towards elucidating mechanisms for the intracellular specification of planar polarity, and further review evidence for models of global polarity regulation at the tissue level. The data suggest that a genetic 'cassette', encoding a set of core signaling components, could pattern hair, bristle and ommatidial planar polarity in Drosophila, and that additional tissue-specific factors might explain the diversity of signal responses. Recently described examples from the nematode and frog suggest that the developmental control of cell polarity by FZ receptors might represent a functionally conserved signaling mechanism.

    View details for Web of Science ID 000076946100005

    View details for PubMedID 9825673

  • Differential recruitment of Dishevelled provides signaling specificity in the planar cell polarity and Wingless signaling pathways GENES & DEVELOPMENT Axelrod, J. D., Miller, J. R., Shulman, J. M., MOON, R. T., Perrimon, N. 1998; 12 (16): 2610-2622


    In Drosophila, planar cell polarity (PCP) signaling is mediated by the receptor Frizzled (Fz) and transduced by Dishevelled (Dsh). Wingless (Wg) signaling also requires Dsh and may utilize DFz2 as a receptor. Using a heterologous system, we show that Dsh is recruited selectively to the membrane by Fz but not DFz2, and this recruitment depends on the DEP domain but not the PDZ domain in Dsh. A mutation in the DEP domain impairs both membrane localization and the function of Dsh in PCP signaling, indicating that translocation is important for function. Further genetic and molecular analyses suggest that conserved domains in Dsh function differently during PCP and Wg signaling, and that divergent intracellular pathways are activated. We propose that Dsh has distinct roles in PCP and Wg signaling. The PCP signal may selectively result in focal Fz activation and asymmetric relocalization of Dsh to the membrane, where Dsh effects cytoskeletal reorganization to orient prehair initiation.

    View details for Web of Science ID 000075604900015

    View details for PubMedID 9716412

    View details for PubMedCentralID PMC317102

  • wingless refines its own expression domain on the Drosophila wing margin NATURE Rulifson, E. J., Micchelli, C. A., Axelrod, J. D., Perrimon, N., Blair, S. S. 1996; 384 (6604): 72-74


    The imaginal discs of Drosophila, which give rise to the adult appendages, are patterned during a period of intense cell proliferation. The specification of differing regions occurs in some cases by subdividing the disc epithelium into lineage compartments. However, in most cases precise boundaries are formed between different cell types without early compartmentalization. One such boundary occurs between the wingless (wg)-expressing cells of the wing margin and the adjacent proneural cells, which give rise to margin sensory bristles. Here we show that this boundary arises in part by a mechanism of 'self-refinement', by which wingless protein (Wg) represses wg expression in adjacent cells. Cells unable to receive the Wg signal do not resolve the boundary between wg-expressing and proneural cells.

    View details for Web of Science ID A1996VR21900056

    View details for PubMedID 8900280

  • Conservation of dishevelled structure and function between flies and mice: Isolation and characterization of Dvl2 MECHANISMS OF DEVELOPMENT Klingensmith, J., Yang, Y., Axelrod, J. D., Beier, D. R., Perrimon, N., Sussman, D. J. 1996; 58 (1-2): 15-26


    The segment polarity gene dishevelled (dsh) of Drosophila is required for pattern formation of the embryonic segments and the adult imaginal discs. dsh encodes the earliest-acting and most specific known component of the signal transduction pathway of Wingless, an extracellular signal homologous to Wnt1 in mice. We have previously described the isolation and characterization of the Dvl1 mouse dsh homolog. We report here the isolation of a second mouse dsh homolog, Dvl2, which maps to chromosome 11. The Dvl2 amino acid sequence is equally related to the dsh sequence as is that of Dvl1, but Dvl2 is most similar to the Xenopus homolog Xdsh. However, unlike the other vertebrate dsh homologs. Like the other genes, Dvl2 is ubiquitously expressed throughout most of embryogenesis and is expressed in many adult organs. We have developed an assay for dsh function in fly embryos, and show that Dvl2 can partially rescue the segmentation defects of embryos devoid of dsh. Thus, Dvl2 encodes a mammalian homolog of dsh which can transduce the Wingless signal.

    View details for Web of Science ID A1996VJ10700002

    View details for PubMedID 8887313

  • The wingless signaling pathway is directly involved in Drosophila heart development DEVELOPMENTAL BIOLOGY Park, M. Y., Wu, X. S., GOLDEN, K., Axelrod, J. D., Bodmer, R. 1996; 177 (1): 104-116


    Heart development in both vertebrates and Drosophila is initiated by bilaterally symmetrical primordia that may be of equivalent embryological origin: the anterior lateral plate mesoderm in vertebrates and the dorsal-most mesoderm in arthropods. These mesodermal progenitors then merge into a heart tube at the ventral midline (vertebrates) or the dorsal midline (Drosophila). These observations suggest that there may be similarities between vertebrate and invertebrate heart development. The homeobox gene, tinman, is required for heart as well as visceral mesoderm formation in Drosophila, and at least one of several vertebrate genes with similarities in protein sequence and cardiac expression to tinman is crucial for heart development in vertebrates. Inductive signals are also required for Drosophila heart development: The secreted gene product of wingless (wg) is critical for heart development during a time period distinct from its function in segmentation and neurogenesis. Here, we show that wg is epistatic to hedgehog (hh), another secreted segmentation gene product, in its requirement for heart formation. We also provide evidence show that downstream of wg in the signal transduction cascade, dishevelled (dsh, a pioneer protein) and armadillo (arm, beta-catenin homolog) are mediating the cardiogenic Wg signal. In particular, overexpression of dsh can restore heart formation in the absence of wg function. We discuss the possibility that Wg signaling is part of a combinatorial mechanism to specify the cardiac mesoderm.

    View details for Web of Science ID A1996VA13600010

    View details for PubMedID 8660881

  • Interaction between wingless and notch signaling pathways mediated by dishevelled SCIENCE Axelrod, J. D., Matsuno, K., ARTAVANISTSAKONAS, S., Perrimon, N. 1996; 271 (5257): 1826-1832


    In Drosophila, the Wingless and Notch signaling pathways function in m any of the same developmental patterning events. Genetic analysis demonstrates that the dishevelled gene, which encodes a molecule previously implicated in implementation of the Winglass signal, interacts antagonistically with Notch and one of its known ligands, Delta. A direct physical interaction between Dishevelled and the Notch carboxyl terminus, distal to the cdc10/ankyrin repeats, suggests a mechanism for this interaction. It is proposed that Dishevelled, in addition to transducing the Wingless signal, blocks Notch signaling directly, thus providing a molecular mechanism for the inhibitory cross talk observed between these pathways.

    View details for Web of Science ID A1996UC77800035

    View details for PubMedID 8596950

  • THE COLLECTION AND EVALUATION OF PERIPHERAL-BLOOD PROGENITOR CELLS SUFFICIENT FOR REPETITIVE CYCLES OF HIGH-DOSE CHEMOTHERAPY SUPPORT TRANSFUSION Benjamin, R. J., LINSLEY, L., Axelrod, J. D., Churchill, W. H., Sieff, C., Shulman, L. N., Elias, A., Ayash, L., MALACHOWSKI, M. E., Uhl, L., GAYNES, L., McCauley, M., Thompson, L., Mazanet, R., Antman, K., Schnipper, L., Tepler, I., Antin, J. H., WHEELER, C. 1995; 35 (10): 837-844


    The development of an optimized peripheral blood progenitor cell (PBPC) harvest protocol to provide support for repetitive chemotherapy cycles is described.PBPCs mobilized by cyclophosphamide plus granulocyte-colony-stimulating factor (G-CSF) were studied in 163 leukapheresis harvests from 26 lymphoma patients. Harvested cells were transfused with two chemotherapy cycles and with an autologous bone marrow transplant. Progenitor cell content was examined in the context of hematopoietic engraftment.Mobilization allowed the harvest of large numbers of PBPCs. Peak harvests tended to occur after the recovering white cell count exceeded 10 x 10(9) per L. CD34+ lymphomononuclear cell (MNC) and colony-forming units-granulocyte-macrophage (CFU-GM) counts correlated poorly, but both measures peaked within 24 hours of each other in 21 of 26 patients, which demonstrated PBPC mobilization. Engraftment of platelets (> 50 x 10(9)/L) and granulocytes (> 500 x 10(6)/L) was achieved in a median of 20.5 and 16 days, respectively. A minimum number of progenitors necessary to ensure engraftment could be derived.Cyclophosphamide and G-CSF allowed the harvest of sufficient PBPCs to support multiple rounds of chemotherapy. Harvest should commence when the recovery white cell count exceeds 10 x 10(9) per L. PBPC harvest CD34+MNC counts are as useful as CFU-GM results in the assessment of PBPC content, and they may allow harvest protocols to be tailored to individual patients.

    View details for Web of Science ID A1995TA44800008

    View details for PubMedID 7570914



    Photofootprinting in vivo of GAL1 reveals an activation-dependent pattern between the UASG and the TATA box, in a sequence not required for transcriptional activation by GAL4. The pattern results from a nucleosome whose position depends on sequences within the UASG. In the wild-type gene, activation by GAL4 and derivatives disrupts this nucleosome. This activity is independent of interactions with DNA-bound core transcription factors and is proportional to the strength of the activator. Presence of the nucleosome correlates with low basal transcription levels under various conditions, suggesting a role in limiting basal expression. We propose a role for the GAL4 activation domain in displacing a nucleosome and suggest that this is part of the mechanism by which GAL4 activates transcription in vivo.

    View details for Web of Science ID A1993LC49800012

    View details for PubMedID 8491382



    The PUT3 gene product is a transcriptional activator required for expression of the enzymes of the proline utilization pathway. Using two methods of footprinting in vivo, we have determined that PUT3 protein is poised at the promoters of the genes encoding these enzymes and that proline-mediated induction modulates the activity of constitutively bound PUT3.

    View details for Web of Science ID A1991ER08100060

    View details for PubMedID 1986247



    We have developed an improved method for photofootprinting in vivo which utilizes the thermostable DNA polymerase from T. aquaticus (Taq) in a primer extension assay. UV light is used to introduce photoproducts into the genomic DNA of intact yeast cells. The photoproducts are then detected and mapped at the nucleotide level by multiple rounds of annealing and extension using Taq polymerase, which is blocked by photoproducts in the template DNA. The method is more rapid, sensitive, and reproducible than the previously described chemical photofootprinting procedure developed in this laboratory (Nature 325. 173-177), and detects photoproducts with a specificity which is similar, but not identical to that of the previously described procedure. Binding of GAL4 protein to its binding sites within the GAL1-10 upstream activating sequence is demonstrated using the primer extension photofootprinting method. The primer extension assay can also be used to map DNA strand breakage generated by other footprinting methods, and to determine DNA sequence directly from the yeast genome.

    View details for Web of Science ID A1989R859800014

    View details for PubMedID 2643080

  • VITAMIN-D AFFECTS PROLIFERATION OF A MURINE T-HELPER CELL CLONE JOURNAL OF IMMUNOLOGY Lacey, D. L., Axelrod, J., Chappel, J. C., Kahn, A. J., Teitelbaum, S. L. 1987; 138 (6): 1680-1686


    1,25-Dihydroxyvitamin D3 (1,25(OH)2D3), the biologically active form of vitamin D3, has been shown to inhibit the activation of T cell hybridomas and heterogeneous populations of mononuclear leukocytes. Because the response of various clones to 1,25(OH)2D3 may differ, we have examined the proliferative effects of the steroid on an antigen-specific cloned, nontransformed T helper cell line (D10.G4.1 [D10 cells]), and find that in contrast to these previous studies, the steroid is a potent stimulator of lectin-induced proliferation. In these experiments, D10 cells were incubated with concanavalin A and 1,25(OH)2D3, and although the lectin or steroid alone has minimal proliferative effects, their co-addition prompts up to a 50-fold increase in 3H-TdR incorporation at a concentration of 2.5 to 5 X 10(-9) M 1,25(OH)2D3, with significant mitogenesis occurring at 0.1 to 0.3 X 10(-9) M 1,25(OH)2D3. 25-Hydroxyvitamin D3 and 24,25(OH)2D3 have similar activity, but at concentrations two to three times greater than that of 1,25(OH)2D3, reflecting their relative affinities for the 1,25(OH)2D3 receptor. In addition, lectin treatment enhances 1,25(OH)2D3 receptor capacity fourfold to fivefold, an event coupled with the appearance of positive cooperativity. Although the steroid does not affect the quantity of bioassayable T cell growth factors as assessed by HT-2 cell proliferation, the expression of immunoreactive IL 2 receptors by lectin-activated D10 cells exposed to 1,25(OH)2D3 is enhanced. In contrast to its proliferative effect in the absence of IL 1, 1,25(OH)2D3 exerts biphasic effects on D10 replication when this monokine is present. Specifically, this steroid augments D10 proliferation at low concentrations of recombinant IL 1, but as the abundance of the monokine increases in the presence of 10(-10) to 10(-8) M 1,25(OH)2D3, the peak response of D10 cells to optimal IL 1 concentrations is diminished. Therefore, in this clone, 1,25(OH)2D3 presents itself as a regulator of T helper cell proliferation.

    View details for Web of Science ID A1987G388300005

    View details for PubMedID 3029220



    Cytochalasin B is shown to inhibit uptake of 3-O-methylglucose into isolated rat hepatocytes with a Ki = 1.9 microM. The nature of this inhibition was characterized by studies of [3H]cytochalasin B binding to liver plasma membranes. Scatchard analysis of [3H]cytochalasin B binding reveals a complex curvilinear binding pattern. This pattern can be resolved into three components: (1) a high-affinity (ca. 10(-8) M) cytochalasin E sensitive site unrelated to glucose uptake, (2) a glucose-sensitive site, and (3) a low-affinity site. When 5 microM cytochalasin E is employed to mask the high-affinity site, glucose displaces 40-60% of the remaining [3H]cytochalasin B binding. Analysis of this glucose-sensitive cytochalasin B binding according to Scatchard reveals a Kd = 1.7 microM, indistinguishable from the concentration of cytochalasin B which half-maximally inhibits hepatic glucose uptake. These data identify a glucose-sensitive cytochalasin B binding site in liver plasma membranes which corresponds to the glucose carrier in the intact hepatocyte. The Ki of 1.9 microM for inhibition of hepatic glucose uptake by cytochalasin B and the Kd of 1.7 microM for [3H]cytochalasin B binding to liver plasma membranes are values 1 order of magnitude higher than values for the same parameters determined in all previous studies of facilitated hexose diffusion systems. The hepatic hexose carrier is therefore unique, and this uniqueness may be of regulatory significance with regard to glucose homeostasis.

    View details for Web of Science ID A1983QM57500025

    View details for PubMedID 6683102


    View details for Web of Science ID A1981LD10900019

    View details for PubMedID 7007367