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

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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