Regulation of Cdc42-Dependent Signaling Pathways Throughout the Cell Cycle

该项目的长期目标是了解真核细胞周期中控制细胞形态发生的信号转导机制。正常的形态发生对细胞分化和繁殖的保真度至关重要。本项目的目的是破译控制酵母细胞极性过程的信号转导机制。在酵母细胞周期中，响应不同信号的极化生长可以产生几种不同的形态结构，如芽、交配突起和假菌丝。首席研究员之前在酵母中描述了细胞极性装置的两种蛋白质成分；ras相关GTPase Cdc42p及其鸟嘌呤核苷酸交换因子（GEF） Cdc24p。这些成分是在细胞周期中导致细胞极性产生的信号转导途径的组成部分。这些成分在其他真核生物中有保守的对应物，表明可能存在控制细胞极性的共同信号转导机制。该项目的目标是了解Cdc42p GTPase的调控以及它如何与不同的细胞周期调控过程相连接。这些实验将验证的假设是：Cdc42p GTPase通过与不同下游效应物的相互作用调节多个细胞周期事件。Cdc42p效应域突变分析表明，Cdc42p参与有丝分裂细胞周期的多个过程，包括芽出、芽出后早期检查点、顶端各向异性开关、g2 /M形态发生检查点和细胞质分裂。然而，Cdc42p与多种下游效应物相互作用调控这些过程的分子机制尚不清楚。通过对D38E突变等位基因和新的Cdc42效应域突变体的表征，我们将研究Cdc42p在这些多细胞周期事件中的功能，重点关注芽后早期出现检查点。此外，通过功能性GFP-Cdc42融合蛋白的突变分析，将确定Cdc42p靶向极化生长位点的机制。这些问题的答案不仅关系到对细胞生物学中信号转导机制的基本理解，而且关系到对酵母和其他真核生物细胞形态发生过程的理解。

The long-term objective of this project is to understand the signal-transduction mechanisms that control cellular morphogenesis during the eukaryotic cell cycle. Normal morphogenesis is essential for the fidelity of cellular differentiation and reproduction. The goal of this project is to decipher the signal-transduction mechanisms that control the cell polarity process in the yeast Saccharomyces cerevisiae. Polarized growth in response to different signals during the yeast cell cycle can result in the generation of several different morphological structures, such as buds, mating projections, and pseudohyphae. The principal investigator has previously characterized two protein components of the cell polarity apparatus in yeast; the Ras-related GTPase Cdc42p and its guanine-nucleotide exchange factor (GEF) Cdc24p. These components are integral parts of a signal-transduction pathway that leads to the generation of cell polarity during the cell cycle. These components have conserved counterparts in other eukaryotes, suggesting that common signal-transduction mechanisms controlling cell polarity may exist. The goal of this project is directed at understanding the regulation of the Cdc42p GTPase and how it interfaces with different cell-cycle regulatory processes. The hypothesis that will be tested with these proposed experiments is: The Cdc42p GTPase regulates multiple cell-cycle events through interactions with different downstream effectors. Analysis of Cdc42p effector domain mutations has indicated that Cdc42p is involved in multiple processes during the mitotic cell cycle, including bud emergence, a early post-bud emergence checkpoint, the apical-isotropic switch, the G 2 /M morphogenetic checkpoint, and cytokinesis. However, the molecular mechanisms by which Cdc42p interacts with multiple downstream effectors to regulate these processes is still unclear. The function of Cdc42p in these multiple cell-cycle events will be examined, concentrating on the early post-bud emergence checkpoint, through the characterization of the D38E mutant allele and new Cdc42 effector-domain mutants. In addition, the mechanism by which Cdc42p is targeted to sites of polarized growth will be determined, through the mutational analysis of a functional GFP-Cdc42 fusion protein. The answers to these questions will not only be relevant to the basic understanding of signal-transduction mechanisms in cell biology, but also to the understanding of the cellular morphogenesis process in yeast and other eukaryotes.