Cell Cycle Regulation of Membrane Trafficking

膜运输的细胞周期调控

• 批准号: 10607265
• 负责人: Joshua Nathaniel Bembenek
• 金额: $ 23.9万
• 依托单位: WAYNE STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-03-15 至 2025-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10607265
• 关键词: Cell; Cycle; Regulation; Membrane; Trafficking

Project Summary Cell division requires a carefully coordinated series of essential events that must be precisely regulated. A central feature of cell division is the accurate segregation of chromosomes into daughter cells. Other compartments of the cell must also be carefully packaged into daughter cells, and coordinated with chromosome segregation. Membrane trafficking pathways are essential for the completion of cytokinesis at the end of cell division. How cells trigger membrane trafficking during cytokinesis is not well understood. The large protease separase is a central player in chromosome segregation due to its role in cohesin cleavage, which allows chromosome separation at the onset of anaphase. After chromosome segregation, separase promotes several events during anaphase. This proposal aims to understand a novel role of separase in the exocytosis of RAB-11 vesicles required for cytokinesis. Separase also regulates exocytosis of large cortical granules during anaphase of meiosis I to block polyspermy, which is an ideal context to analyze this regulatory pathway. This proposal describes research aimed at understanding how separase promotes exocytosis in collaboration with other closely related cell cycle regulators that have well characterized roles in chromosome segregation. The dynamic localization of separase is regulated during cell division and only localizes to vesicles during anaphase. Proposed research will investigate how chromosome segregation regulators control separase activity and localization to vesicles. Overexpression of non-degradable securin will be used to determine how this inhibitory chaperone controls the exocytic function of separase. Mutations of the PPH-5 phosphatase and its activator HSP-90 were identified as suppressors of separation of function alleles of separase. The functions of PPH-5 and HSP-90 will be characterized to determine whether they directly regulate separase during the meiotic divisions. Separase phosphorylation sites will be mapped and phosphorylation mutants will be studied to determine how they affect separase function. PPH-5 will be tested to determine if it directly dephosphorylates separase in vitro. Finally, biochemical and genetic approaches will be employed to identify substrates or targets of separase on vesicles to define the mechanism by which it promotes exocytosis. These studies will be performed using the genetically tractable C. elegans embryo. This work will provide new insight into how cells coordinate the essential process of chromosome segregation with exocytosis during cytokinesis, which is relevant to understanding normal development and diseases such as infertility and cancer.

项目摘要 细胞分裂需要精心协调的一系列基本事件，必须精确调节。 细胞分裂的一个主要特征是将染色体的准确分离为子细胞。其他 细胞的隔室也必须小心地包装到子细胞中，并与 染色体分离。膜运输途径对于完成细胞因子至关重要 细胞分裂的末端。细胞如何触发细胞因子期间的膜运输尚不清楚。大 蛋白酶分离酶是染色体隔离的核心参与者，因为它在粘蛋白裂解中的作用 允许在后期开始时染色体分离。染色体分离后，分离酶促进 后期期间的几个事件。该建议旨在了解分离酶在胞吐作用中的新作用 rab-11囊泡的细胞因子。分离酶还调节大型皮质颗粒的胞吐作用 在减数分裂后期，I阻断多植物，这是分析该调节途径的理想背景。 该建议描述了旨在了解分离酶如何促进胞吐作用的研究 与其他密切相关的细胞周期调节剂在染色体分离中具有良好表征的作用。 分离酶的动态定位受到细胞分裂的调节，仅在囊泡中定位于囊泡 后期。拟议的研究将研究染色体分离调节剂如何控制分离酶 活动和囊泡的定位。非依赖性证券的过表达将用于确定如何 该抑制性伴侣控制分离酶的外囊肿功能。 PPH-5磷酸酶和 其激活剂HSP-90被鉴定为分离酶功能等位基因分离的抑制剂。功能 PPH-5和HSP-90的特征是确定它们是否在 减数分裂分裂。将映射分离酶磷酸化位点，并研究磷酸化突变体 确定它们如何影响分离酶功能。 PPH-5将进行测试以确定它是否直接 去磷酸化的分离酶体外。最后，将采用生化和遗传方法来识别 囊泡上分离酶的底物或靶标，以定义促进胞吐作用的机制。这些 研究将使用遗传性的秀丽隐杆线虫胚胎进行。这项工作将提供新的见解 细胞如何协调染色体分离的基本过程与细胞因子期间的胞吐作用， 这与了解正常发育和疾病（例如不育和癌症）有关。