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、定位和溶解性是基本过程，对于适当的细胞功能以及生物体生长和发育至关重要;然而，确保忠实执行这些事件的质量控制机制随着年龄的增长而日益受到损害。毫不奇怪，当这些正常的蛋白质生物合成途径出错时，可能会产生毁灭性的后果。例如，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，以及一组称为传染性海绵状脑病的独特朊病毒疾病[1，2]，每种都与由正常宿主编码蛋白质的自我永久错误折叠引起的严重神经变性相关。在健康的个体中，这些蛋白质是可溶的和功能性的，但在患病的个体中，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继续错误折叠的模板，从而促进它们自身的组装。芽殖酵母酿酒酵母（Saccharomyces cerevisiae）表达几种朊病毒蛋白，这些朊病毒蛋白也以类似的自模板过程在可溶性和聚集形式之间分配。然而，在这些情况下，这些蛋白质的朊病毒形式与疾病无关，而是作为表观遗传因子指导新性状的遗传[3]。鉴于酵母的实验易处理性和朊病毒状态的可行性，该系统提供了一个宝贵的平台，解剖的机制，在体内蛋白质构象的自我复制的变化。从对酵母朊病毒蛋白Sup 35的大量体内研究中已经收集到关于朊病毒有效繁殖所需顺式序列的详细信息。该因子的朊病毒结构域中的几个突变主要干扰野生型蛋白的朊病毒有效繁殖，表明这些变体特异性靶向该过程中的关键步骤。为了深入了解这些事件，我建议阐明的机制，这些变体消除错误折叠的形式，通过结合体内和体外分析。总之，这些研究将定义赋予蛋白质自我复制特性的关键特性，并通过扩展突出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.