Mechanisms of Contact-Mediated Cell Polarization in the C. elegans Embryo.

胚胎细胞发育为形态发生和分化所需的专门化。对称性- 通过改变RhoGT 3信号传导，打破许多细胞的线索，这触发了不对称的皮质 PAR极性蛋白的定位。连接极性线索、RhoGT 3信号的分子链， 而PAR不对称性却知之甚少。利用C.线虫胚胎作为一个简单的模型，长期 本项目的目标是确定细胞接触信号如何调节RhoGTP酶以诱导PAR 不对称性使细胞变小。我们将联合收割机胚胎学操作与细胞生物学和 遗传工具提供了一个独特的机会，以确定和表征这些机制在活的胚胎。 鉴于细胞极性通路和调节因子的深度保守性，我们的研究将为以下方面提供新的见解： 关键接触介导的极化事件在人类的机制。这些问题包括： 在致密化过程中出现的胚胎卵裂球，为内细胞团的特化所必需 （它产生胚胎本身，是胚胎干细胞的来源）; 上皮细胞是器官发生所需的，并且是抑制肿瘤形成和侵袭所必需的。 在前一个奖项期间，我们定义了一个介导接触诱导的 通过在空间上改变RhoGTdCDC-42 - a信号蛋白的活性来极化卵裂球 在细胞极性中扮演着古老且广泛保守的角色。我们发现细胞接触会募集保守的 RhoGAP蛋白PAC-1/ARHGAP 10，在接触部位灭活CDC-42。CDC-42仍然活跃， 无接触表面，在那里它招募PAR蛋白PAR-3，PAR-6和PKC-3/aPKC，然后 每个卵裂球此外，我们获得的初步数据表明，CDC-42控制了 PAR-6和PAR-3通过不同的机制定位，CDC-42通过调节PAR-3定位， 膜运输这项计划的目标是确定细胞间的分子联系， 接触、PAC-1募集和CDC-42活性导致PAR-3不对称。我们的具体目标是 (1)以确定卵裂球之间的细胞接触如何募集PAC-1以局部抑制CDC-42;（2） 确定在非接触表面激活CDC-42的机制;以及（3）测试以下假设： CDC-42通过调节膜运输控制PAR-3不对称性。我们的实验将使我们能够 一种连接细胞-细胞接触与CDC-42活性及其极性调节的分子途径。我们 预计我们的发现将为细胞极化的基本机制提供一般性的见解， 以及细胞如何接触人类卵裂球和上皮细胞， 促进胚胎发育和抑制肿瘤发生。

Embryonic cells polarize to develop specializations needed for morphogenesis and differentiation. Symmetry- breaking cues polarize many cells by altering RhoGTPase signaling, which triggers the asymmetric cortical localization of PAR polarity proteins. The molecular links that connect polarity cues, RhoGTPase signaling, and PAR asymmetries are poorly understood. Using the C. elegans embryo as a simple model, the long-term goal of this project is to determine how cell contact cues regulate RhoGTPases to induce the PAR asymmetries that polarize cells. Our ability to combine embryological manipulations with cell biological and genetic tools provides a unique opportunity to identify and characterize these mechanisms in living embryos. Given the deep conservation of cell polarity pathways and regulators, our studies will provide new insights into mechanisms of critical contact-mediated polarization events in humans. These include the polarization of embryonic blastomeres that occurs during compaction and is required for specification of the inner cell mass (which gives rise to the embryo proper and is the source of embryonic stem cells); and the polarization of epithelial cells that is needed for organogenesis and is essential for inhibiting tumor formation and invasion. In the prior award period, we defined a molecular pathway that mediates the contact-induced polarization of blastomeres by spatially altering the activity of the RhoGTPase CDC-42 - a signaling protein with an ancient and broadly conserved role in cell polarity. We showed that cell contacts recruit the conserved RhoGAP protein PAC-1/ARHGAP10, which inactivates CDC-42 at contact sites. CDC-42 remains active at contact-free surfaces, where it recruits the PAR proteins PAR-3, PAR-6, and PKC-3/aPKC that then polarize each blastomere. In addition, we have obtained preliminary data suggesting that CDC-42 controls the localization of PAR-6 and PAR-3 through distinct mechanisms, and that CDC-42 localizes PAR-3 by regulating membrane trafficking. The goal of this proposal is to identify the molecular links between cell-cell contact, PAC-1 recruitment, and CDC-42 activity that lead to PAR-3 asymmetry. Our specific goals are (1) to determine how cell contacts between blastomeres recruit PAC-1 to locally inactivate CDC-42; (2) to identify the mechanisms that activate CDC-42 at contact-free surfaces; and (3) to test the hypothesis that CDC-42 controls PAR-3 asymmetry by regulating membrane trafficking. Our experiments will allow us to build a molecular pathway that connects cell-cell contact to CDC-42 activity and its regulation of polarity. We anticipate that our findings will provide general insights into the fundamental mechanisms of cell polarization, and specific molecular insights into how cell contacts polarize human blastomeres and epithelial cells to promote embryonic development and inhibit tumorigenesis.