Characterization of the Mitotic Exit Network

描述(申请人提供)：真核细胞必须在时间上协调不同的细胞过程，以确保细胞增殖的保真度。细胞周期过程的执行或时间协调的缺陷会增加遗传的不稳定性，并促进癌症的发生。因此，解决保守的细胞周期调控机制是非常重要的。酿酒酵母有丝分裂退出网络(MAN)在细胞有丝分裂退出过程中协调几个细胞周期过程，包括细胞周期蛋白依赖性激酶(CDK)失活、胞质分裂、有丝分裂纺锤体的分解和G1基因表达的激活，是必不可少的。Mob1-Dbf2激酶复合体是人类的一个关键成分，与人的Mob1a-Lats激酶复合体同源，后者具有哺乳动物肿瘤抑制因子的功能。酵母Mob1-Dbf2参与了胞质分裂、CDC14磷酸酶的激活(通过抑制CDK磷酸化使有丝分裂退出)和有丝分裂纺锤体功能的调节。然而，酵母和哺乳动物Mob1-Dbf2激酶的作用机制和底物尚不清楚。这项拟议工作的目标是确定Mob1-Dbf2激酶复合体如何在时间上协调胞质分裂与有丝分裂退出和染色体分离。我们提出了三个目标。第一个目的是确定Mob1-Dbf2在胞质分裂中的作用。我们将通过细胞生物学、生化和遗传学方法来验证这一假设，即男性通过调节靶向膜沉积和激活RAM(调节ACE2转录因子和极化形态发生)信号网络来控制细胞质分裂，RAM信号网络控制细胞质分裂、基因表达和其他功能。第二个目的是确定Mob1-Dbf2激酶复合体在有丝分裂退出和胞质分裂过程中CDC14磷酸酶激活中的作用。我们将检验Mob1-Dbf2激酶直接诱导CDC14磷酸酶从核仁释放的假设，并将定义Mob1-Dbf2和CDclp与胞质分裂的上位关系。第三个目的是确定Mob1-Dbf2在有丝分裂检查点信号中的作用。我们将测试这一假设，即Mob1-Dbf2是极光依赖的检查点信号所必需的。鉴于Mob1-Dbf2的保守性，拟议的工作将有助于解决所有与男性相关的信号网络的功能，包括人类hMob1A-LATS1肿瘤抑制通路，并将有助于阐明癌症发生的潜在机制。

DESCRIPTION (provided by applicant): Eukaryotic cells must temporally coordinate diverse cellular processes to ensure the fidelity of cell proliferation. Defects in the execution or temporal coordination of cell cycle processes increase genetic instability and promote cancer. Thus, it is important to resolve the mechanisms of conserved cell cycle regulations. The Saccharomyces cerevisiae Mitotic Exit Network (MEN) is essential for coordinating several cell cycle processes during mitotic exit, including cyclin dependent kinase (CDK) inactivation, cytokinesis, mitotic spindle disassembly and activation of G1 gene expression. The Mob1-Dbf2 kinase complex is a key component of MEN and is orthologous to the human Mob1A-LATS kinase complex, which functions as mammalian tumor suppressor. Yeast Mob1-Dbf2 has been implicated in cytokinesis, activation of Cdc14 phosphatase (which enables mitotic exit by inactivating CDK phosphorylations) and mitotic spindle function via regulation of Aurora kinase. Nevertheless, the functional mechanisms and substrates of yeast and mammalian Mob1-Dbf2 kinases are not known. The goal of the proposed work is to determine how the Mob1-Dbf2 kinase complex temporally coordinates cytokinesis with mitotic exit and chromosome segregation. We propose three aims. The first aim is to determine the role of Mob1-Dbf2 in cytokinesis. We will conduct cell biological, biochemical and genetic approaches to test the hypothesis that MEN controls cytokinesis by regulating targeted membrane deposition and by activating the RAM (Regulation of Ace2 transcription factor and polarized Morphogenesis) signaling network, which controls cytokinesis, gene expression and other functions. The second aim is to determine the role of the Mob1-Dbf2 kinase complex in Cdc14 phosphatase activation during mitotic exit and cytokinesis. We will test the hypothesis that Mob1- Dbf2 kinase directly induces Cdc14 phosphatase release from the nucleolus and will define the epistatic relationship of Mob1-Dbf2 and Cdclp with respect to cytokinesis. The third aim is to determine the role of Mob1-Dbf2 in mitotic checkpoint signaling. We will test the hypothesis that Mob1-Dbf2 is required for Aurora-dependent checkpoint signaling. Given the conservation of Mob1-Dbf2, the proposed work will help resolve the function of all MEN-related signaling networks, including the human hMob1A-LATS1 tumor suppressor pathway, and will help elucidate the underlying mechanisms of cancer development.