Actin Myosin Interactions in Cell Motility

描述(申请人提供)：我的长期目标是了解细胞质分裂的机制。我们将使用裂殖酵母Pombe进行互补的细胞、生化和遗传策略，以解决以下特定目标：项目1.收缩环蛋白的动力学：这些对活细胞中分子的定量实验将提供破译收缩环组装途径所需的参数值。我们将测量胞质蛋白在细胞质和收缩环之间的交换速度，确定肌动蛋白细丝是否影响每个收缩环蛋白的动力学，并确定胞质分裂蛋白是否影响彼此的动力学。项目2：在前身结节的宽带中组装胞质分裂蛋白：这些实验将阐明组装收缩环前体物的生化途径。我们将确定与皮质和下游蛋白结合的苯丙素Mid1p的结构域，以及细胞质分裂蛋白如何与Mid1p以及彼此相互作用。项目3.将前体节点转变为收缩环：这项工作将提供了解收缩环组装的第二步所需的信息。我们将研究CDc12p如何控制节点上肌动蛋白细丝的组装，表征将一条宽带的节点转换为收缩环的机械过程，表征DCS蛋白Rng3p和磷酸化如何影响Myo2细丝的结构，并确定细胞质蛋白在宽带和收缩环中的组织方式。项目4.收缩环的收缩和解体：这些实验将帮助我们了解细胞如何触发收缩环的收缩，以及环如何在收缩时解体。我们将研究SIN通路如何触发收缩环的收缩，探索限制收缩环收缩速度的因素，并确定收缩环在收缩过程中如何解体。

DESCRIPTION (provided by applicant): My long range goal is to understand the mechanism of cytokinesis. We will pursue complementary cellular, biochemical and genetic strategies using the fission yeast Schizosaccharomyces pombe to address the following specific aims: Project 1. Dynamics of contractile ring proteins: these quantitative experiments on molecules in live cells-will provide the parameter values required to decipher the pathway of contractile ring assembly. We will measure the rates that cytokinesis proteins exchange between the cytoplasm and contractile rings, determine if actin filaments influence the dynamics of each contractile ring protein and determine if cytokinesis proteins influence each other's dynamics. Project 2. Assembly of cytokinesis proteins in the broad band of precursor nodes: these experiments will clarify the biochemical pathway that assembles the precursor of the contractile ring. We will determine the domains of anillin Mid1p that bind the cortex and downstream proteins and how cytokinesis proteins interact with Mid1p and with each other. Project 3. Transformation of precursor nodes into a contractile ring: this work will provide information required to understand the second step in the assembly of the contractile ring. We will investigate how Cdc12p controls the assembly of actin filaments from nodes, characterize the mechanical process that converts, a broad band of nodes into a contractile ring, characterize how DCS protein Rng3p and phosphorylation affect the structure of Myo2 filaments and determine how cytokinesis proteins are organized in the broad band and the contractile ring. Project 4. Constriction and disassembly of the contractile ring: these experiments will help us understand how cells trigger constriction of the contractile ring and how the ring disassembles as it constricts. We will investigate how the SIN pathway triggers constriction of the contractile ring, explore the factors that limit the rate of constriction of contractile rings and determine how the contractile ring disassembles during constriction.