Signaling mechanisms in cell polarity in yeast

描述（由申请人提供）：细胞极性对发育和分化至关重要，它在细胞迁移、上皮间营养转运和神经元传递等基本过程中起着至关重要的作用。细胞极性缺陷通常与癌症等严重的人类疾病有关。我们的长期目标是利用遗传上易于处理的真核生物酿酒酵母来阐明细胞极化的原理。在这篇文章中，我们将探讨Cdc42p（一个进化上保守的小GTPase）在极性发展中的核心作用。在Aim1中，我们将确定Cdc42p的活性如何受到其gap、Rga1p和Bem2p的时空调节，从而形成一个单一的极化域，这是所有极化系统中的一个共同问题。已知有四种gap （Rga1p, Rga2， Bem3p和Bem2p）作用于Cdc42p。之前，我们和其他人已经证明了三个gap （Rga1p， Rga2和Bem3p）在细胞周期开始时在分隔素环组装中起着共同的作用。最近，我们发现Rga1p在细胞分裂后细胞的旧细胞分裂位点中是唯一需要防止极化的。在本提案中，我们将测试我们的新想法，即Rga1p和Bem2p在芽颈作为“守门人”冗余地将活跃的Cdc42p和Rho1p限制在一个单一的极化域，即芽皮质。迄今为止，我们的研究表明，不同的gap可以单独或共同调节涉及Cdc42p的特定细胞过程，并帮助建立了研究哺乳动物Rho gtpase（人类基因组中~17个）通过其众多gap（人类基因组中~68个）的复杂调控的范例。在Aim2中，我们将提出实验来测试我们的Cdc42p在极化肌动蛋白和septin组织中的作用的综合模型，它们共同决定了细胞的整体形状，并决定了它们在极化胞外分泌和定向细胞迁移等基本过程中的不同功能。我们的模型表明，Cdc42p通过两种遗传上可分离的，但生物化学上的交叉对话途径控制极化肌动蛋白和间隔蛋白的组织，一种涉及进化上保守的“极化体”，以形成蛋白Bni1p、支架蛋白Spa2p和Rab GAPs Msb3p和Msb4p为中心，另一种涉及酵母特异性Cdc42p效应物Gic1p和Gic2p。在本提案中，我们将确定极化体途径如何通过肌动蛋白细胞骨架影响间隔蛋白的组织，Gic途径如何通过间隔蛋白影响肌动蛋白的组织，以及这两种途径如何在细胞应激（如370C）下通过基于spa2p的相互作用进行串接。Cdc42p的同源物参与许多细胞功能，如细胞极性、细胞迁移和细胞生长控制。哺乳动物体内Cdc42p活性的减弱与癌症等严重的人类疾病有关。因此，研究酵母中Cdc42p的信号机制将在基础生物学和临床科学中具有深远的意义。

DESCRIPTION (provided by applicant): Cell polarity is essential for development and differentiation, and it plays vital roles in fundamental processes such as cell migration, nutrient transport across epithelia, and neuronal transmission. Defects in cell polarity are often associated with serious human diseases such as cancer. Our long-term objective is to use the genetically tractable eukaryote Saccharomyces cerevisiae to elucidate the principles of cell polarization. In this proposal, we will address two important questions regarding the central role of Cdc42p, an evolutionarily conserved small GTPase, in polarity development. In Aim1, we will determine how Cdc42p activity is spatiotemporally regulated by its GAPs, Rga1p and Bem2p, in the formation of a single polarization domain, a common issue among all polarization systems. There are four GAPs (Rga1p, Rga2, Bem3p, and Bem2p) known to act on Cdc42p. Previously, we and others have shown that three GAPs (Rga1p, Rga2, and Bem3p) share a role in septin ring assembly at the beginning of the cell cycle. Recently, we discovered that Rga1p is uniquely required for preventing polarization within old cell division sites in post-cytokinesis cells. In this proposal, we will test our novel idea that Rga1p and Bem2p function redundantly at the bud neck as "gatekeepers" to restrict active Cdc42p and Rho1p to a single polarization domain, the bud cortex. To date, our studies have indicated that different GAPs can act alone or together to regulate specific cellular processes involving Cdc42p, and have helped establish a paradigm for studying complex regulation of mammalian Rho GTPases (~17 in human genome) by their numerous GAPs (~68 Rho GAPs in human genome). In Aim2, we will propose experiments to test our integrative model for the role of Cdc42p in polarized actin and septin organization, which together determine the overall cell shape and dictate their diverse functions in fundamental processes such as polarized exocytosis and directed cell migration. Our model states that Cdc42p controls polarized actin and septin organization via two genetically separable, but biochemically cross-talking pathways, one involving the evolutionarily conserved "polarisome" that is centered on the formin Bni1p, scaffold protein Spa2p, and the Rab GAPs Msb3p and Msb4p, and the other involving the yeast-specific Cdc42p effectors Gic1p and Gic2p. In this proposal, we will determine how the polarisome pathway affects septin organization via the actin cytoskeleton, how the Gic pathway affects actin organization via the septins, and how these two pathways crosstalk under cellular stresses such as 370C via Spa2p-based interactions. Homologues of Cdc42p are involved in numerous cellular functions such as cell polarity, cell migration, and cell growth control. Deregulation of Cdc42p activity in mammals is associated with serious human diseases, such as cancer. Thus, studying the signaling mechanisms of Cdc42p in yeast will have profound implication in basic biology and clinical sciences. PUBLIC HEALTH RELEVANCE: Cdc42p, an evolutionarily conserved small GTPase, plays essential roles in diverse cellular processes such as cell polarity, cell migration, and cell growth control. Deregulation of Cdc42p activity in mammals is associated with serious human diseases, such as cancer. Thus, studying the signaling mechanisms of Cdc42p in yeast will have profound implication in basic biology and clinical sciences.