MICROTUBULE/ACTIN INTERACTIONS IN CELL MOTILITY

细胞运动中的微管/肌动蛋白相互作用

• 批准号: 6181997
• 负责人: Clare Michal Waterman
• 金额: $ 29.11万
• 依托单位: SCRIPPS RESEARCH INSTITUTE, THE
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-09-01 至 2005-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6181997
• 关键词: Xenopus oocyte; actins; cell motility; digital imaging; fluorescence microscopy; fluorescent dye /probe; guanine nucleotide binding protein; guanine nucleotide exchange factors; intermolecular interaction; microtubules; tubulin

Cell locomotion is critical to development, wound healing and tissue remodeling, and a central problem during metastasis in cancer. Thus, it is important to gain an understanding of cell motility at the basic level. The cytoskeletal polymers, microtubules and filamentous actin (f-actin) are required for cell motility. Polarized f-actin assembly and movement is thought to drive cell motility, while microtubules are believed to spatially direct sites of f-actin assembly. There is evidence that microtubules may achieve this by activation of cellular signaling via small GTPases of the Rho family. However, the organization and dynamics of microtubules and how they are affected by actin is not thoroughly characterized, and the molecules involved in mediating their interactions during cell motility are unknown. I approach these problems at 3 levels: First, by examining microtubules and f-actin dynamics in living cells using multimode microscopy, by exploring with both microscopy and biochemistry microtubule/f-actin interactions in a cellular extract model systems, and finally by examining the biochemical interactions of purified components of the motile machinery in vitro. The combination of unique microscopy methods and molecular biochemistry is a novel means of carefully characterizing cellular phenomenon and then going directly to the test tube test hypotheses about the molecular mechanisms underlying the phenomena. In this proposal, I describe three specific aims to shed light on our understanding of microtubule/f- actin interactions in cell motility. 1. To develop new microscopy methods for the simultaneous visualization and analysis of f-actin and microtubule dynamics and to carefully analyze microtubule/actin interactions in living migrating cells. 2. To biochemically and microscopically probe the molecular mechanism dynamic microtubule/actin interactions in Xenopus egg extracts. 3. To biochemically analyze microtubule mediation of Rho family signaling and study the mechanism by which growing microtubules activate Rac1 during cell motility. The results of these studies will greatly enhance our understanding of the mechanisms and mechanics of cell locomotion and will provide new methods for analyzing protein dynamics in living cells.

细胞运动对于癌症转移期间的转移期间的发育，伤口愈合和组织重塑至关重要。 因此，重要的是要了解基本水平的细胞运动。 细胞骨架聚合物，微管和丝状肌动蛋白（F-肌动蛋白）是细胞运动所必需的。 偏振-ACTIN的组装和运动被认为可以驱动细胞运动，而微管被认为可以在空间上引导F-肌动蛋白组件的位点。 有证据表明，微管可以通过Rho家族的小GTPase激活细胞信号传导来实现这一目标。 但是，微管的组织和动力学及其如何受肌动蛋白的影响没有彻底表征，并且在细胞运动过程中介导其相互作用的分子尚不清楚。 我在3个级别上解决这些问题：首先，通过使用多模显微镜检查活细胞中的微管和F-肌动蛋白动力学，通过在细胞提取物模型中探索显微镜和生物化学显微管/F-肌动蛋白的相互作用，最后通过检查Motile Motile Motile Macninery In Vetro的生物化学组合的生物化学相互作用。 独特的显微镜方法和分子生物化学的组合是一种仔细表征细胞现象，然后直接进行测试管测试的新颖手段，假设有关该现象背后的分子机制。 在此提案中，我描述了三个特定的目的，以阐明我们对细胞运动中微管/F-肌动蛋白相互作用的理解。 1。开发新的显微镜方法，用于同时可视化和分析F-肌动蛋白和微管动力学，并仔细分析活细胞中的微管/肌动蛋白相互作用。 2。在生化和微观上探测分子机制动态微管/肌动蛋白相互作用中的Xenopus卵提取物中的相互作用。 3。生化分析Rho家族信号传导的微管介导，并研究生长的微管在细胞运动过程中激活Rac1的机制。 这些研究的结果将大大增强我们对细胞运动机制和力学的理解，并将为分析活细胞中的蛋白质动力学提供新的方法。