Investigating the Mechanism of Prion Curing by [PSI+] No More Mutations

描述（由申请人提供）：蛋白质的折叠、定位和溶解是基本的过程，对正常的细胞功能以及生物体的生长和发育至关重要；然而，随着年龄的增长，确保这些活动忠实执行的质量控制机制越来越受到损害。不足为奇的是，当这些正常的蛋白质生物生成途径出错时，可能会造成毁灭性的后果。例如，与年龄相关的蛋白质错误折叠疾病，包括阿尔茨海默病、帕金森病和亨廷顿病，以及一系列被称为传染性海森堡脑病的独特朊病毒疾病[1,2]，都与严重的神经退行性变有关，这些退行性变是由正常宿主编码蛋白质的自我延续的错误折叠引起的。在健康个体中，这些蛋白质是可溶的并具有功能，但在患病个体中，相同的蛋白质错误折叠并形成高度结构化的聚集体，这些聚集体似乎充当正常蛋白质持续错误折叠的模板，从而促进其自身的组装。出芽酵母酿酒酵母表达几种朊病毒蛋白，这些朊病毒蛋白也在类似的自模板过程中分为可溶性和聚集形式。然而，在这些情况下，这些蛋白的朊病毒形式与疾病无关，而是作为一种表观遗传因子来指导新性状[3]的遗传。鉴于酵母的实验可追溯性和朊病毒状态的生存能力，该系统为剖析体内蛋白质构象自我复制变化的机制提供了宝贵的平台。关于高效朊病毒繁殖的顺式序列要求的详细信息已经从酵母朊病毒蛋白Sup35的体内研究中收集到。该因子的朊病毒结构域的几个突变主要干扰野生型蛋白的高效朊病毒繁殖，这表明这些变异专门针对该过程中的关键步骤。为了深入了解这些事件，我建议通过体内和体外分析的结合来阐明这些变异消除错误折叠形式的机制。总之，这些研究将确定赋予蛋白质自我复制特性的关键特性，并进一步强调哺乳动物蛋白质错误折叠疾病的治疗干预的潜在点。

DESCRIPTION (provided by applicant): The folding, localization, and solubility of proteins are basic processes, essential for proper cellular function as well as organismal growth and development; yet the quality control mechanisms that ensure faithful execution of these events become increasingly compromised with age. Not surprisingly, when these normal pathways of protein biogenesis go awry there can be devastating consequences. For example, the age related protein misfolding diseases, including Alzheimer's, Parkinson's and Huntingdon's diseases, as well as a set of unique prion diseases termed the transmissible spongiform encephalopathy's [1, 2] are each associated with severe neurodegeneration resulting from the self-perpetuated misfolding of a normal, host encoded protein. In healthy individuals, these proteins are soluble and functional, but in afflicted individuals, the same protein misfolds and forms highly structured aggregates which appear to act as templates for the continued misfolding of normal protein, thereby promoting their own assembly. The budding yeast Saccharomyces cerevisiae expresses several prion proteins that also partition between soluble and aggregated forms in an analogous, self-templated process. In these cases, however, the prion form of these proteins is not associated with disease but instead acts as an epigenetic agent to direct the inheritance of a novel trait [3]. Given the experimental tractability of yeast and the viability of the prion state, this system provides an invaluable platform with which to dissect the mechanisms underlying self-replicating changes in protein conformations in vivo. Detailed information on the cis sequence requirements for efficient prion propagation has been gleaned from a number of in vivo studies on the yeast prion protein Sup35. Several mutations in the prion domain of this factor dominantly interfere with efficient prion propagation by the wildtype protein, suggesting that these variants specifically target crucial steps in the process. To gain insight into these events, I propose to elucidate the mechanisms by which these variants eliminate the misfolded form through a combination of in vivo and in vitro analyses. Together, these studies will define the key properties that endow a protein with self-replicating character and by extension highlight potential points of therapeutic intervention for the mammalian protein misfolding diseases.