Roles of Cdc42 and its signaling partners in cell growth and differentiation

描述(由申请人提供)：Ras相关的GTPase CDC42在广泛的细胞过程中发挥重要作用，包括确定细胞极性和控制细胞生长和迁移。对CDC42偶联信号活性的适当调控对其细胞功能至关重要，癌基因鸟嘌呤核苷酸交换因子(GEF)或突变导致GDP加速交换为GTP，导致细胞转化和裸鼠肿瘤形成，证明了这一点。在过去的资助期间，我们确定了CDC42及其调控蛋白Cool-1(用于从文库中克隆出来)是如何通过帮助维持适当的EGF受体(EGFR)内环境平衡来调节细胞生长的。我们还发现Cool-1的磷酸化-去磷酸化循环影响细胞迁移，增加了CDC42及其信号伙伴可能协调细胞生长控制与细胞运动调节的有趣可能性。此外，我们对不同细胞和遗传模型系统的研究突出了CDC42在细胞分化和细胞命运决定中的作用。在接下来的资助期间，我们将继续将基于生化和结构生物学的研究与遗传学方法相结合，以扩展这些发现，并更好地确定CDC42及其调节蛋白如何影响三个基本的重要细胞过程，即细胞生长、迁移和分化。这些研究将构成三条具体的调查路线。1)确定CDC42及其信号伙伴如何协同工作以维持适当的EGFR动态平衡。在这些研究中，我们将特别感兴趣的是确定调控信号，这些信号设定了CDC42介导的信号的时间，从而在EGFR偶联的有丝分裂信号与受体下调和降解之间建立了适当的平衡。2.)研究Cool-1及其结合伙伴是如何调控细胞迁移的。在这里，我们将跟踪最近的线索，表明Cool-1的磷酸化刺激其与Cat(针对Cool相关酪氨酸激酶底物)的解离，并帮助触发焦点复合体的解体。我们还想看看Cool-1的CDC42-Egf活性是否由其磷酸化触发，对细胞迁移具有重要的调节作用。3.)研究CDC42及其信号伙伴在细胞分化中的作用。我们将探索CDC42在确保神经元分化所需的信号活动的适当寿命方面的可能参与。此外，我们将建立在我们最近对小鼠胚胎(P19)细胞的研究基础上，这些研究表明，我们最近发现的CDC42和一个双重功能的CDC42-环境基金/靶-效应器，在神经发生中发挥重要作用。我们预计，这些研究将对CDC42如何能够调节正常生物结果所必需的多种细胞反应，并在解除调控时，导致包括癌症和神经退行性疾病在内的各种疾病状态产生新的见解。

DESCRIPTION (provided by applicant): The Ras-related GTPase Cdc42 plays important roles in a wide range of cellular processes including the establishment of cell polarity and the control of cell growth and migration. The proper regulation of Cdc42- coupled signaling activities is crucial for its cellular functions, as evidenced by the hyper-activation of Cdc42 by oncogenic guanine nucleotide exchange factors (GEFs), or mutations that give rise to an accelerated exchange of GDP for GTP, resulting in cellular transformation and tumor formation in nude mice. During the past funding period, we established how Cdc42 and its regulatory protein Cool-1 (for Cloned-out of library), which functions both as a GEF and a target/effector for Cdc42, regulate cell growth by helping to maintain proper EGF receptor (EGFR) homeostasis. We also discovered that the phosphorylation-dephosphorylation cycle of Cool-1 influences cell migration, raising the interesting possibility that Cdc42 and its signaling partners may coordinate cell growth control with the regulation of cell motility. Moreover, our studies with different cell and genetic model systems have highlighted a role for Cdc42 in cellular differentiation and cell fate determination. In the coming funding period, we will continue to combine biochemical and structural biology-based studies with genetic approaches to extend these findings and better establish how Cdc42 and its regulatory proteins impact three fundamentally important cellular processes, namely cell growth, migration, and differentiation. These studies will constitute 3 specific lines of investigation. 1) Determine how Cdc42 and its signaling partners work together to maintain proper EGFR homeostasis. In these studies, we will be particularly interested in determining the regulatory cues that set the timing for the Cdc42- mediated signals that establish the proper balance between EGFR-coupled mitogenic signaling versus receptor down-regulation and degradation. 2.) Examine how Cool-1 and its binding partners regulate cell migration. Here, we will follow-up recent clues suggesting that the phosphorylation of Cool-1 stimulates its dissociation from Cat (for Cool-associated tyrosine kinase substrate) and helps trigger the disassembly of focal complexes. We also will want to see whether the Cdc42-GEF activity of Cool-1, which is triggered by its phosphorylation, confers important regulatory effects on cell migration. 3.) Examine the roles of Cdc42 and its signaling partners in cellular differentiation. We will explore the possible involvement of Cdc42 in ensuring the proper lifetime for signaling activities necessary for neuronal differentiation. In addition, we will build on our recent studies in mouse embryonic (P19) cells that suggest Cdc42 and a dual function Cdc42- GEF/target-effector that we recently discovered, play important roles in neurogenesis. We expect that these studies will yield new insights into how Cdc42 is able to mediate a diversity of cellular responses that are necessary for normal biological outcomes and, when de-regulated, gives rise to a variety of disease states including cancer and neurodegenerative disorders.