YEAST MEMBRANE CYTOSKELETON

酵母膜细胞骨架

• 批准号: 6564220
• 负责人: DAVID DRUBIN
• 金额: $ 14.33万
• 依托单位: UNIVERSITY OF CALIF-LAWRENC BERKELEY LAB
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-01-01 至 2002-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6564220
• 关键词: Mammalia; actin binding protein; actins; cell osmotic pressure; cytoskeletal proteins; genetic library; laboratory rabbit; mechanical pressure; membrane permeability; membrane proteins; membrane structure; molecular biology; polymerase chain reaction; protein structure function; yeasts

The membrane cytoskeletons of mammalian cells and budding yeast have so far been studied intensively but by almost completely distinct methodologies. We propose hare to initiate a comparison of the molecular components and physical properties of the membrane cytoskeletons of yeast and mammals. We hope that general principles may be distilled when common features are found. (1) Physical properties of yeast with mutations in membrane cytoskeleton proteins. Mutants isolated in a wide variety of cytoskeletal proteins in budding yeast show osmosensitivity and apparent membrane fragility. These observations suggest that the yeast membrane actin cytoskeletal proteins in budding yeast show osmosensitivity and apparent membrane fragility. These observations suggest that the yeast membrane actin cytoskeleton, like the erythrocyte membrane cytoskeleton, plays a critical role in providing structural integrity to the plasma membrane. We will use the ~sensitive force technique: and the ~micropipette technique~ to test the physical properties of the yeast cell cortex in cells with and without cell walls. These studies will provide a deeper understanding of how physical al properties of the cell cortex are determined. (2) Identification of mammalian homologues of yeast membrane cytoskeleton proteins. Genetic analysis of the yeast membrane cytoskeleton has identified both homologues of actin-binding proteins found in other cells ( capping protein, cofilin and fimbrin) and novel proteins (Sla2p, Abp1p, Rvs167, Srv2 and S1a1p). Finding mammalian homologues of the novel proteins will provide an opportunity to apply knowledge from genetic analysis in yeast to mammalian cells. The prospects for finding such homologues are good. A mammalian Srv2 homologue has already been found, an Sla2 homologue exists in C. Elegans, and Sla1 homologue exists in S. pombe, and an Abp1 homolog exists in S. exiguus. (3). Erythrocyte cytoskeleton homologues in yeast. Band 4.1, spectrin and ankyrin have been extensively studies in erythrocytes using biochemistry and molecular-genetics. Finding proteins in yeast with either extensive sequence homology, or homology to a functional domain, would allow classical genetic analysis of their functions. We will query the yeast genome database (presently, 70% accessible, 90% complete) using particularly conserved domains of these cytoskeletal proteins. We will disrupt the genes for any homologues identified in yeast for analysis as described in Aim (1), and will raise antisera to immunolocalize the proteins in vivo.

哺乳动物细胞和芽殖酵母的膜细胞骨架具有如此 迄今为止已被深入研究，但几乎完全不同 方法论。 我们建议野兔发起比较 膜的分子成分和物理性质 酵母和哺乳动物的细胞骨架。 我们希望一般原则能够 当发现共同特征时进行蒸馏。 (1) 膜细胞骨架突变酵母的物理特性 蛋白质。 在多种细胞骨架蛋白中分离出突变体 芽殖酵母表现出渗透敏感性和明显的膜脆性。 这些观察表明酵母膜肌动蛋白细胞骨架 芽殖酵母中的蛋白质表现出渗透敏感性和明显的膜 脆弱性。 这些观察表明酵母膜肌动蛋白 细胞骨架与红细胞膜细胞骨架一样，起着 在提供质膜结构完整性方面发挥着关键作用。 我们将使用〜敏感力技术：和〜微量移液器 技术〜测试酵母细胞皮层的物理特性 有细胞壁和无细胞壁的细胞。 这些研究将提供更深入的 了解细胞皮层的物理特性 决定。 (2)酵母膜哺乳动物同源物的鉴定 细胞骨架蛋白。 酵母膜的遗传分析 细胞骨架已鉴定出肌动蛋白结合蛋白的两种同源物 在其他细胞中发现（加帽蛋白、丝切蛋白和纤维蛋白）并且新颖 蛋白（Sla2p、Abp1p、Rvs167、Srv2 和 S1a1p）。 寻找哺乳动物 新型蛋白质的同源物将提供应用的机会 从酵母到哺乳动物细胞的遗传分析知识。 这 寻找此类同系物的前景良好。 哺乳动物 Srv2 已经发现了同源物，C 中存在 Sla2 同源物。 线虫中存在 Sla1 同源物，而 Abp1 同源物存在于粟酒裂殖酵母中 存在于 S. exiguus 中。 （3）。酵母中的红细胞细胞骨架同源物。 波段 4.1，血影蛋白 和锚蛋白已在红细胞中进行了广泛的研究 生物化学和分子遗传学。 寻找酵母中的蛋白质 广泛的序列同源性，或与功能域的同源性， 将允许对其功能进行经典的遗传分析。 我们将 查询酵母基因组数据库（目前，70%可访问，90% 完整）使用这些细胞骨架的特别保守的结构域 蛋白质。 我们将破坏基因中发现的任何同源物 酵母用于目标 (1) 中所述的分析，并将抗血清提高到 使蛋白质在体内免疫定位。