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Development of a new screening system for drugs that modify the signaling pathways regulating the actin cytoskeleton

开发一种新的筛选系统，用于修饰调节肌动蛋白细胞骨架的信号通路的药物

基本信息

批准号：
11557008
负责人：
TANAKA Kazuma
金额：
$ 7.04万
依托单位：
HOKKAIDO UNIVERSITY
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B).
财政年份：
1999
资助国家：
日本
起止时间：
1999 至 2000
项目状态：
已结题

项目摘要

Abnormal regulation of the actin cytoskeleton is implicated in the invasion and metastasis of the cancer cells. The actin cytoskeleton system is conserved in the yeast Saccharomyces cerevisiae, suggesting that S.cerevisiae can be used to search for a new drug which prevents the invasion and metastasis of the cancer cells. However, there are still many things that remain to be studied. In this study, we have investigated the regulation of the actin cytoskeleton and attempted to establish such a system. (1) Isolation of the MTI1 gene as a regulator of the type I myosins. The yeast type I myosins, Myo3/5p, are involved in the reorganization of the actin cytoskeleton. The SH3 domain of Myo5p has been shown to interact with Las17p [a homolog mammalian Wiskott-Aldrich syndrome protein (WASP)] and Vrp1p [a homolog of WASP-interacting protein (WIP)] to regulate polymerization of actin. Vrp1p is required for the localization of Myo5p to cortical patch-like structures and for the ATP-independent … More interaction of the Myo5p tail region with actin filaments. We found that a new adaptor protein, Mti1p (Myosin tail region-interacting protein), interacts with the SH3 domains of Myo3/5p. Mti1p is coimmunoprecipitated with Myo5p and Mti1p-EGFP is colocalized with cortical actin patches. Although the mti1 null mutation did not cause any obvious phenotype by itself, it suppressed the phenotypes of the vrp1 mutants, including temperature-sensitive growth, abnormally large morphology, and endocytosis deficiency. Our results indicate that Mti1p and Vrp1p antagonistically regulate the functions of type I myosins. (2) Development of the screening system. Mammalian WASP, WIP, or type I myosins are being expressed in yeast. We are looking for a mammlian homolog of Mti1p. We will generate mutant genes that dominantly inhibit cell growth by affecting the reorganization of the actin cytoskeleton. Drugs that rescue growth inhibition of the yeast cells may interfere with the function of the mutant mammalian proteins. Less

肌动蛋白细胞骨架的异常调节与癌细胞的侵袭和转移有关。肌动蛋白的细胞骨架系统在酿酒酵母中是保守的，这表明酿酒酵母可以用来寻找防止癌细胞侵袭和转移的新药。然而，仍有许多事情有待研究。在这项研究中，我们研究了肌动蛋白细胞骨架的调节，并试图建立这样的系统。(1)分离作为I型肌球蛋白调节因子的MTI1基因。酵母I型肌球蛋白Myo3/5p参与肌动蛋白细胞骨架的重组。Myo5p的SH3结构域与Las17p[哺乳动物Wiskott-Aldrich综合征蛋白(WASP)的同源蛋白]和Vrp1p[WASP-相互作用蛋白(WIP)的同源蛋白]相互作用，调节肌动蛋白的聚合。Vrp1p是Myo5p定位为皮质斑片状结构和不依赖三磷酸腺苷的…所必需的Myo5p尾部区域与肌动蛋白细丝的相互作用更多。我们发现一种新的接头蛋白Mti1p(肌球蛋白尾部相互作用蛋白)与Myo3/5p的SH3结构域相互作用。Mti1p与Myo5p免疫共沉淀，Mti1p-EGFP与皮质肌动蛋白贴片共定位。虽然mti1缺失突变本身没有引起任何明显的表型，但它抑制了vrp1突变体的表型，包括对温度敏感的生长，异常大的形态和内吞缺陷。我们的结果表明，Mti1p和Vrp1p拮抗地调节I型肌球蛋白的功能。(2)筛选系统的开发。哺乳动物的WASP、WIP或I型肌球蛋白正在酵母中表达。我们正在寻找Mti1p的哺乳动物同源物。我们将产生突变基因，通过影响肌动蛋白细胞骨架的重组来主要抑制细胞生长。挽救酵母细胞生长抑制的药物可能会干扰突变的哺乳动物蛋白质的功能。较少