The Role of Competitive Forces in Prion Propagation and Appearance

描述（由申请人提供）：根据朊病毒假说，正常宿主编码蛋白质的替代构象（称为朊病毒）可作为体内可传播表型状态的表观遗传决定因素。一旦被认为是非典型的，这一过程已被链接到一个扩大范围的以前神秘的生物学现象，如传染性海绵状脑病（TSE）的病因学和非孟德尔遗传的一组亚稳态性状的真菌，这表明它是一个广泛的过程，通过它的生物体可以访问和延续平行的表型状态。虽然这些表型的蛋白质基础是有据可查的，但令人惊讶的是，我们对它们之间的转换如何发生的了解有限，尽管它们对正常细胞生理学具有重要意义。我们的长期目标是揭示体内朊病毒转变的途径。为了获得这一认识，我们将利用S.啤酒。朊病毒在该系统中的繁殖，也可能在哺乳动物中，通过蛋白质生物合成的多步途径发生，该途径受朊病毒的固有特性和细胞环境的影响。[PSI繁殖通常是有效的，但许多条件，与哺乳动物中的相似之处，诱导表型转变。该提议的总体目标是将经验研究和计算研究联合收割机结合起来，以确定体内朊病毒传播途径的哪些步骤被这些操作改变，以及这些变化引起开关的细胞途径。到目前为止，大多数分析都是从朊病毒的角度来考虑这些影响，但我们提出的研究是在整个系统的背景下考虑它们的作用。虽然诱导朊病毒相关表型之间转换的条件将朊病毒的不同构象或序列变体引入细胞中，但竞争力在朊病毒转换中的作用从未被探索过。我们假设朊病毒传播途径中的关键步骤应被视为受竞争力影响的酶限制过程，这可以有效地引起表型转变。为了直接验证这一假设，我们将确定：1）显性阴性突变体治愈朊病毒的途径，2）朊病毒变体建立显性的途径，3）第二种朊病毒从头出现时需要现有朊病毒的分子基础。通过这种独特的视角，我们将开始揭示朊病毒相关表型之间转换的细胞途径，这是朊病毒生物学的一个重要但知之甚少的方面。 公共卫生相关性：拟议的研究是相关的公共卫生，因为外观的替代，自我复制的蛋白质构象与一系列广泛的家族性，散发性和传染性神经退行性疾病的man.Our目前无法解释，预测或利用正常和疾病状态之间的转换是一个关键的障碍，在这些疾病的治疗进展。我们提出的研究是相关的NIH的使命，因为这里获得的知识将提供洞察力约束这些状态之间的切换和系统的鲁棒性外部扰动。

DESCRIPTION (provided by applicant): According to the prion hypothesis, an alternative conformation of a normal, host-encoded protein, known as a prion, can function as an epigenetic determinant of transmissible phenotypic states in vivo. Once considered atypical, this process has been linked to an expanding range of previously enigmatic biological phenomena, such as the etiology of the transmissible spongiform encephalopathies (TSEs) and the non-Mendelian inheritance of a group of metastable traits in fungi, suggesting it is a widespread process through which organisms can access and perpetuate parallel phenotypic states. While the protein-only basis of these phenotypes is well documented, we have surprisingly limited insight into how transitions between them occur, despite their significance for normal cellular physiology. Our long-term goal is to reveal the pathways through which prion transitions are evoked in vivo. To gain this insight, we will exploit the manipulability of the Sup35/[PSI+] prion of S. cerevisiae. Prion propagation in this system, and also likely in mammals, occurs through a multi-step pathway of protein biogenesis that is influenced by both the inherent properties of the prion and by aspects of the cellular environment. [PSI+] propagation is normally efficient, but many conditions, with parallels in mammals, induce phenotypic transitions. The overall objective of this proposal is to combine empirical and computational studies to determine which step(s) of the in vivo prion propagation pathway are altered by these manipulations and the cellular pathways through which these changes evoke the switch. To date, most analyses have considered these effects from the perspective of the prion alone, but our proposed studies consider their action in the context of the entire system. While conditions that induce transitions between prion-associated phenotypes introduce different conformational or sequence variants of the prion into cells, the role of competitive forces in prion transitions has never been explored. We hypothesize that the crucial steps in the prion propagation pathway should be considered as enzyme-limited processes that are subject to competitive forces, which can effectively evoke phenotypic transitions. To directly test this hypothesis, we will determine: 1) the pathways by which dominant-negative mutants cure prions, 2) the pathways by which prion variants establish dominance, and 3) the molecular basis of the requirement for an existing prion in the de novo appearance of a second prion. Through this unique perspective, we will begin to reveal the cellular pathways underlying transitions between prion-associated phenotypes, a crucial yet poorly understood aspect of prion biology. PUBLIC HEALTH RELEVANCE: The proposed research is relevant to public health because the appearance of alternative, self-replicating protein conformations are associated with a wide array of familial, sporadic and transmissible neurodegenerative diseases in man. Our current inability to explain, predict or exploit transitions between normal and disease states is a critical barrier to progress in the development of treatments for these disorders. Our proposed studies are relevant to the mission of the NIH because the knowledge gained here will provide insight into the forces constraining switches between these states and the robustness of the system to external perturbation.