Role of the Actin Cytoskeleton in Yeast Prion Propagation

描述（由申请人提供）：正常蛋白质的错误折叠已成为哺乳动物疾病以及低等真核生物中某些表型的表观遗传传递的重要机制。与这些现象有关的蛋白质，统称为淀粉样变性，有一个关键的共同特征：自我复制异常折叠的能力。构象自我复制在体内是一个多步骤的过程，包括三个高度调控的步骤：转换，维持和繁殖。虽然这个过程的某些方面可以在体外的简化系统中建模，但对自繁殖构象的生理后果的充分理解需要了解体内的这种途径，其中基本的细胞过程可以与蛋白质动力学相交并调节蛋白质动力学。一个特别有趣但很大程度上未被表征的相互作用是淀粉样蛋白形成蛋白和细胞骨架之间的关系，这在许多系统中被越来越多的研究反复联系在一起。这里提出的工作的总体目标是了解这两个系统之间的功能相互作用。为此，我的工作将集中在朊病毒在低等真核生物中的繁殖，这为研究体内蛋白质自我繁殖的调节提供了一个实验上易于处理的模型。对S. cerevisiae是一种蛋白质，其正常功能由朊病毒周期可逆地调节，将用于研究肌动蛋白动力学如何影响错误折叠的蛋白质自我复制。正如许多哺乳动物淀粉样蛋白已被证明可以促进肌动蛋白捆绑的情况一样，以前对Sup 35朊病毒的研究已经确定了与肌动蛋白细胞骨架相关的蛋白质的多种相互作用。通过生物化学、细胞生物学和遗传学方法的结合，我将剖析肌动蛋白细胞骨架在Sup 35朊病毒（[PS/"*]）表型的多步骤、自我繁殖周期中的作用。由于酵母朊病毒模型已被证明是直接适用于高等真核生物的疾病，包括一系列的人类疾病，在这里获得的见解将提供肌动蛋白在分子水平上的哺乳动物蛋白质折叠疾病的永久化的作用机制的线索。

DESCRIPTION (provided by applicant): The misfolding of normal proteins has emerged as an important mechanism in both mammalian disease as well as the epigenetic transmission of some phenotypes in lower eukaryotes. The proteins implicated in these phenomena, collectively known as amyloidoses, share a key common trait: the capacity to self-replicate the aberrant fold. Conformational self-replication is a multistep process in vivo, consisting of three highly regulated steps: conversion, maintenance and propagation. While some aspects of this process can be modeled in a simplified system in vitro, a full understanding of the physiological consequences of self-propagating conformations requires an understanding of this pathway in vivo, where basic cellular processes can intersect with and regulate protein dynamics. One particularly intriguing but largely uncharacterized interaction is the relationship between amyloid-forming proteins and the cellular cytoskeleton, which are repeatedly linked together in many systems by a growing series of studies. The overall goal of the work proposed here is to understand the functional interplay between these two systems. Toward this end, my work will focus on prion propagation in lower eukaryotes, which provides an experimentally tractable model for studying the regulation of protein self-propagation in vivo. The Sup35 protein of S. cerevisiae, a protein whose normal function is reversibly modulated by a prion cycle, will be used to examine how actin dynamics influence misfolded protein self-replication. As is the case for a number of mammalian amyloidogenic proteins that have been shown to promote actin bundling, previous studies on the Sup35 prion have identified multiple interactions with proteins associated with the actin cytoskeleton. Through a combination of biochemical, cell biological, and genetic approaches, I will dissect the role of the actin cytoskeleton in the multistep, self-propagation cycle of the Sup35 prion ([PS/"*]) phenotype. As the yeast prion model has been shown to be directly applicable to disorders in higher eukaryotes including a range of human diseases, the insights gained here will provide mechanistic clues into the role of actin in the perpetuation of mammalian protein folding disorders on a molecular level.