Prion Cycle Regulation In Vivo

描述(申请人提供)：从哺乳动物神经退行性疾病的出现、进展和扩散到真菌独特特征的非孟德尔遗传，Prion假说为一系列以前无法解释的现象提供了解释。根据这一想法，当普恩蛋白采用另一种物理状态时，与普里恩相关的表型就会出现，当这种形式自我复制时，它就会持续存在。这种自我复制是通过将交替折叠的Prion蛋白组装成聚集体来调节的，聚集体以其他形式的蛋白质转换为类似状态的模板。Prion蛋白质利用其构象灵活性的固有能力是建立不同表型的中心事件，但在活细胞的背景下，将这一过程扩展到实践中是一个多步骤的努力。蛋白质质量控制途径、Prion蛋白质的生物发生和细胞生物学都在体内改变了Prion蛋白质的错误折叠，从而在生理上产生了可传递的变化。对这些力量如何相交的分子理解是当前知识中的一个明显缺口，限制了我们在体外将蛋白质错误折叠机制与体内疾病机制联系起来的能力。由于与Prion相关的表型的出现、扩散和逆转必然涉及蛋白质状态的变化，这些力量必须聚集在调节Prion形式之间转换的事件上。这项研究的长期目标是通过了解Pron蛋白动力学与其细胞环境之间的相互作用，阐明Pron蛋白作为疾病和遗传性元素的分子机制。这项应用的总体目标是确定Pron蛋白的物理特性如何调节Pron聚合动态，以创建不同的表型。中心假说是，Pron蛋白中的特定序列元素影响其识别和/或分子伴侣的处理，创建了一个动态系统的连续体，允许不同的表型出现和持续，但也偶尔进行相互转换。在利用酿酒酵母实验上易驯化的Sup35/[Psi+]Prion的强大初步数据的指导下，这一假说将通过三个具体目标进行检验：1)确定Sup35序列变异改变Prion繁殖的分子机制；2)确定过量Hsp104导致Prion丢失的分子机制；3)确定Sup35/[PSI+]Prion表型的分子基础。这些拟议的研究是创新的，因为他们使用了独特的实验和数学分析的组合来在时间上将Pron蛋白质的物理和功能状态的转变联系起来。这项拟议的研究具有重要意义，因为它将为探索蛋白质错误折叠与其生理后果之间的关系提供一个新的动态框架。鉴于从酵母到人类的Pron序列和错误折叠途径的惊人相似性，这些观察结果将广泛适用于由这些迷人的蛋白质调控的广泛的生物事件。

DESCRIPTION (provided by applicant): The prion hypothesis provides an explanation for a diverse collection of previously inexplicable phenomena, ranging from the appearance, progression and spread of neurodegenerative disease in mammals to the non- Mendelian inheritance of unique traits in fungi. According to this idea, prion-associated phenotypes arise when a prion protein adopts an alternative physical state and persist when that form self-replicates. This self- replication is mediated by the assembly of alternatively folded prion protein into aggregates, which template the conversion of other forms of the protein to a like state. The inherent ability of prion proteins to harness their conformational flexibility is a central event in establishing distinct phenotypes, but the extension of this process to practice becomes a multistep endeavor within the context of a living cell. Protein quality control pathways, prion protein biogenesis, and cell biology all modify prion protein misfolding in vivo to create transmissible changes in physiology. A molecular understanding of how these forces intersect is a clear gap in current knowledge, limiting our ability to correlate protein misfolding mechanisms in vitro and disease mechanisms in vivo. As the appearance, spread and reversal of prion-associated phenotypes necessarily involve changes in protein state, these forces must converge on events that regulate transitions between prion forms. The long-term goal of this research is to elucidate the molecular mechanisms allowing prion proteins to act as elements of disease and heritability by developing an understanding of the interplay between prion protein dynamics and its cellular context. The overall objective of this application is to determine how the physical characteristics of prion proteins modulate prion aggregate dynamics to create distinct phenotypes. The central hypothesis is that specific sequence elements within prion proteins impact their recognition and/or processing by molecular chaperones, creating a continuum of dynamic systems that allow distinct phenotypes to appear and persist but also to occasionally interconvert. Guided by strong preliminary data using the experimentally tractable Sup35/[PSI+] prion of S. cerevisiae, this hypothesis will be tested through three specific aims: 1) Deter- mine the molecular mechanism by which sequence variants of Sup35 alter prion propagation, 2) Determine the molecular mechanism by which excess Hsp104 leads to prion loss, and 3) Determine the molecular basis of the Sup35/[PSI+] prion phenotype. These proposed studies are innovative because they use a unique combination of experimental and mathematical analyses to temporally link transitions in prion protein physical and functional state. The proposed research is significant because it will provide a new and dynamic framework for exploring the relationship between prion protein misfolding and its physiological consequences. Given the remarkable similarity of prion sequences and misfolding pathways from yeast to man, these observations will be broadly applicable to the wide range of biological events regulated by these fascinating proteins.