CENTROSOME MATURATION IN VITRO

中心体参与细胞内细胞结构的组织 从酵母到人类。作为微管的组织者(微管组织 中心体)、中心体参与了许多基本细胞 包括细胞迁移和细胞分裂的过程。因此，和 对中心体组成和生化调控的认识 将对理解癌症、转移、趋化性和 早期发展的各个方面。已经开发了一些方法来采取 文蛤卵母细胞的独特特性的优势使 制备和纯化足够量的中心体 分析生物化学。这项提案中概述的实验将 确定中心体的蛋白质组成，并确定 组成变化是减数分裂细胞周期的一个函数。功能性 体外检测结合视频和电子显微镜将被用于 量化中心粒含量和纯化的中心体的能力 直接微管聚合。将会产生单抗 用于鉴定和纯化分子(MTOC)，这些分子直接 微管组织。最后，将进行重点实验，以 确定中心体是否可以在体外被诱导成熟，如 它们形成和复制中心粒的能力或它们的能力的变化 来组织微管。这些研究将为经典的研究开辟道路 生化方法被设计用来纯化调节 细胞周期中的中心体。

Centrosome are involved in the organization of cytoarchitecture in cells from yeast to man. As organizers of microtubules (microtubule organizing centers, MTOCs), centrosomes are involved in numerous fundamental cell processes including cell migration and cell division. Thus, and understanding of the composition and biochemical regulation of centrosomes will have an impact on understanding cancer, metastasis, chemotaxis, and aspects of early development. Methods have been developed to take advantage of the unique characteristics of clam oocytes which allow the purification of sufficient quantities of centrosomes for preparative and analytical biochemistry. The experiments outlined in this proposal will define the protein composition of centrosomes and determine if compositional changes are a function of the meiotic cell cycle. Functional in vitro assays coupled with video- and electron-microscopy will be used to quantitate centriole content and the ability of purified centrosomes to direct microtubule polymerization. Monoclonal antibodies will be generated for the identification and purification of molecules (MTOCs) which direct microtubule organization. Finally, key experiments will be carried out to determine if centrosomes can be induced to mature in vitro, as indicated by changes in their ability to form and replicate centrioles or their ability to organize microtubules. These studies will open the way for classic biochemical approaches designed to purify molecules which regulate centrosomes during the cell cycle.