Planarian anterior–posterior axis is established by interaction of voltage-dependent Ca2+ influx and β-catenin signaling at the early stage of regeneration

Abstract

Concurrent session 3: Mechanisms of developmental patterning Program/Abstract # 18 Dynamic interpretation of the Hedgehog gradient drives wing disc patterning in Drosophila Marcos Nahmad, Angela Stathopoulos Department of CDS, Caltech, Pasadena, CA, USA Div. of Biol., Caltech, Pasadena, CA, USA Morphogens are classically defined as molecules that control patterning by acting at a distance to regulate gene expression in a concentration-dependent manner. In the Drosophila wing imaginal disc, secreted Hedgehog (Hh) forms an extracellular gradient that organizes patterning along the anterior–posterior axis by defining at least three different domains of gene expression. However, a direct spatial correspondence between the boundaries of these patterns and Hh-concentration thresholds has not been demonstrated. We have acquired evidence that the interpretation of Hh signaling depends on the history of exposure to Hh and that a single concentration threshold is sufficient to support multiple outputs. Mathematical modeling and computer simulations suggest that a spatial ‘overshoot’ of the Hh gradient occurs, that is, a transient gradient in which the Hh-concentration profile is expanded compared to the steady-state gradient. This overshoot results from the gene network architecture, in particular, it depends on Hh-dependent up-regulation of the Hh receptor, Patched (Ptc). Through a temporal examination of Hh-target gene expression in vivo, we observe that particular borders of expression shift posteriorly, and subsequently refine. Furthermore, when the network structure is altered such that Ptc transcription is no longer activated in response to Hh signaling, we find that the patterns of gene expression, which have distinct borders in wild-type discs, now in fact overlap. Taken together, our results demonstrate that the overshoot of the Hh gradient is necessary to establish different patterns of gene expression. doi:10.1016/j.ydbio.2009.05.025 Program/Abstract # 19 Planarian anterior–posterior axis is established by interaction of voltage-dependent Ca influx and β-catenin signaling at the early stage of regeneration Taisaku Nogi, Dan Zhang, John D. Chan, Jonathan S. Marchant Department of Pharmacology and The Stem Cell Institute, University of Minnesota Medical School, MN, USA Since the classic experiments of Marsh and Beams in the 1950s, it has been suggested that endogenous voltage gradients play an important role in planarian regeneration. Here, we show voltagedependent Ca influx is a key regulator of anterior–posterior (AP) axis specification during planarian regeneration. Using a pharmacological screen of Ca signaling modulators in the planarian D. japonica, we identified that praziquantel (PZQ) – a voltage-operated Ca channel (VOCC) activator – had a robust effect to yield a bipolar (two-headed) phenotype from regenerating trunk fragments. PZQtreated fragments displayed a mirror-imaged pharynx and posterior, rather than ectopic, head. This effect was Ca-dependent and increased by a variety of depolarizing agents known to couple to VOCCs. Time-window experiments and in situ hybridization of marker gene expression demonstrated PZQ treatment impacted the early stages of regeneration (<18 h post amputation) and produced a complete anteriorization. Chemical genetic analysis identified that Ltype voltage-operated Cachannel (VOCC) β subunits were essential for PZQ action as in vivo RNAi for βsubunits counteracted PZQ-evoked bipolarity. Finally, β-catenin RNAi and PZQ co-treatment synergistically caused the bipolar phenotype, suggesting PZQ-evoked Ca signals inhibit Wnt/β-catenin signaling. Taken together, these data suggest voltage-dependent Ca influx is an upstream factor to Wnt/ β-catenin signals that establish AP axis polarity during planarian regeneration. doi:10.1016/j.ydbio.2009.05.026 Program/Abstract # 20 The diverse functions of transcriptional factor AP2 (TFAP2) targets in neural crest development Ting Thalia Luo, Yan Xu, Janaki Rangarajan, Trevor Hoffman, Yoo-Seok Hwang, Thomas Schilling, Thomas Sargent Lab. of Mol. Genet., NICHD/NIH, Bethesda, MD 20892, USA Department of Dev. and Cell. Biol. Univ. of California, Irvine, CA 92697, USA Neural crest (NC) cells are a unique feature of vertebrate embryos, arising from the border of neural and non-neural ectoderm as the result of inductive signals including BMP and Wnt, among others. NC cells migrate extensively to give rise to a wide variety of cell lineages, i.e., the craniofacial bones, pigment cells, neurons and glia of the peripheral nervous system. We showed that the transcription factor AP2 (TFAP2) plays an important role in regulating gene expression in NC cells, and is positively regulated by BMP and Wnt. By the microarray approach, we have identified three predominantly NC expressed genes as TFAP2 targets: PCNS, Myosin X (MyoX) and Inca, were intensively studied in the Xenopus and zebrafish by in vivo and in vitro. PCNS is a novel protocadherin which appears to regulate cadherin mediated cell–cell adhesion. MyoX is a non-muscle myosin known to play important roles in the formation of filopodia and cell Developmental Biology 331 (2009) 390–391