PROTEIN COMPOSITION OF ACTIN TAILS ASSEMBLED IN YEAST EXTRACTS

这个子项目是利用资源的许多研究子项目之一。 由NIH/NCRR资助的中心拨款提供。对子项目的主要支持 子项目的首席调查员可能是由其他来源提供的， 包括美国国立卫生研究院的其他来源。为子项目列出的总成本可能 表示该子项目使用的中心基础设施的估计数量， 不是由NCRR赠款提供给次级项目或次级项目工作人员的直接资金。 笼蛋白介导的内吞作用是活细胞中的一个基本过程，它允许活细胞有效地从细胞外环境中捕获大分子，并将它们包装成胞浆中的囊泡。许多疾病可以追溯到导致内吞的一系列分子反应的缺陷。在萌芽中的酿酒酵母中，内吞作用定位于皮质肌动蛋白斑块，在皮质肌动蛋白斑块中，肌动蛋白细丝的致密网络在笼罩着网状蛋白的凹坑周围聚合。这一网络由一组复杂的蛋白质调节，这些蛋白质相互协作以产生膜内陷。 我们最近在实验室开发了一种酵母提取物系统，用于在聚苯乙烯微珠周围组装基于肌动蛋白的结构。我们认为，这些结构的蛋白质组成可能类似于在体内发现的皮质肌动蛋白斑块中的蛋白质组成。我们分离了这些珠子及其相关的肌动蛋白结构，并希望通过质谱学来确定蛋白质的组成。这些结果应该使我们能够进一步分析不同成分在内吞作用中的作用。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Clathrin-mediated endocytosis is a fundamental process in living cells, allowing them to capture macromolecules efficiently from the extracellular environment and package them into vesicles in the cytosol. Numerous diseases can be traced to a defect in the succession of molecular reactions leading to endocytosis. In the budding yeast Saccharomyces cerevisiae, endocytosis is localized to cortical actin patches, where a dense network of actin filaments is polymerized around clathrin-coated pits. This network is regulated by a complex set of proteins which cooperate in order to generate membrane invagination. We have recently developed in our laboratory a system in yeast extracts for the assembly of actin based structures around polystyrene microbeads. We believe that the protein composition of these structures could be similar to the protein composition found in vivo in cortical actin patches. We isolated the beads with their associated actin structures, and we would like to determine the protein composition by using mass spectroscopy. These results should allow us to further analyze the role of the different components during endocytosis.