Mechanism of prion species barrier at short phylogenetic distances in a yeast model

酵母模型中短系统发育距离的朊病毒物种屏障机制

• 批准号: 0614772
• 负责人: Yury Chernoff
• 金额: $ 56.82万
• 依托单位: Georgia Tech Research Corporation
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-09-01 至 2011-08-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=0614772&HistoricalAwards=false
• 关键词: Mechanism; prion; species; barrier; short

Amyloids are highly ordered self-seeded fibrous protein aggregates possibly representing an ancient protein fold and serving as a molecular tool for propagation of prions, which are infectious proteins. Prion infection is thought to spread via nucleated polymerization, so that polymeric prion "seeds" immobilize a normal monomeric protein of the same amino acid sequence and convert it into a prion. Transmission of the prion state from a pre-existing amyloid polymer to the newly mobilized protein molecules is a strictly sequence-specific process. The so-called "species barrier" prevents transmission of the prion state even between closely related proteins. If the barrier is overcome, this results in a cross-species prion transmission. The overall goal of this project is to uncover the molecular mechanisms that control the prion species barrier at low levels of protein sequence divergence. Cross-species prion transmission involves immobilization of a divergent protein into heteropolymers (co-aggregation), and propagation of the newly generated amyloid via its fragmentation, producing new "seeds". The species barrier between highly divergent prion domains is due to their inability to co-aggregate. It is hypothesized that at low levels of sequence divergence, a heterologous prion domain can be incorporated into the heteropolymers, but is unable to acquire the conformational state which would allow for efficient fragmentation and production of new prion "seeds" in the heterologous host. The project is designed to test this hypothesis. Specific Research Objectives are as follows: 1) to study cross-seeding capabilities of the divergent prion domains in vitro; 2) to identify the sequence differences that are responsible for the species barrier and defective propagation of the heterologous prions; 3) to investigate the role of cellular environment in the propagation of heterologous prions and the species barrier. Prion proteins of the closely related yeast species will be studied. Level of variation among these proteins corresponds to the range of divergence observed in the species barrier and cross-species prion transmission phenomena in the mammalian systems. A combination of both in vitro and in vivo approaches will be employed, followed by a direct comparison of the results. Prions can be found in mammals, yeast, and other fungi. They control inheritance of non-Mendelian traits in yeast and other fungi, and provide a mechanism for the "structural inheritance," possibly playing an important role in evolution. Results of this research will contribute to development of the strategies for prevention of cross-species prion transmission among closely related animals, and may have far-reaching implications for understanding the mechanisms controlling the specificity of protein-protein interactions involved in structural inheritance. The research project relies heavily on the participation of graduate and undergraduate students, and is being carried out by a diverse team of males and females. The project will be integrated courses for undergraduares and graduate students, who will contribute to it through their doctoral projects. The PI's laboratory participates in the multi-institutional Center for Fundamental and Applied Molecular Evolution (FAME), which promotes interdisciplinary research and education on various topics related to chemical and biological evolution. New genetically marked strains of the Saccharomyces species other than S. cerevisiae, constructed in the course of this project, will help to advance further studies of these organisms.

淀粉样蛋白是高度有序的自接种纤维状蛋白质聚集体，可能代表古老的蛋白质折叠，并作为感染性蛋白质朊病毒繁殖的分子工具。朊病毒感染被认为是通过有核聚合来传播的，因此聚合朊病毒“种子”覆盖具有相同氨基酸序列的正常单体蛋白并将其转化为朊病毒。朊病毒状态从预先存在的淀粉样蛋白聚合物到新动员的蛋白质分子的传递是严格的序列特异性过程。所谓的“物种屏障”阻止了朊病毒状态的传播，即使是在密切相关的蛋白质之间。如果障碍被克服，这将导致跨物种朊病毒传播。这个项目的总体目标是揭示在低水平的蛋白质序列差异下控制朊病毒物种屏障的分子机制。跨物种朊病毒传播涉及将趋异蛋白固定成异聚体（共聚集），以及通过其片段化使新产生的淀粉样蛋白增殖，从而产生新的“种子”。高度分化的朊病毒结构域之间的物种屏障是由于它们不能共聚集。假设在低水平的序列差异下，异源朊病毒结构域可以被掺入到异源聚合物中，但不能获得允许在异源宿主中有效片段化和产生新朊病毒“种子”的构象状态。该项目旨在验证这一假设。具体的研究目标如下：1）研究不同朊病毒结构域在体外的交叉接种能力; 2）鉴定导致异源朊病毒种属屏障和缺陷性繁殖的序列差异; 3）研究细胞环境在异源朊病毒繁殖和种属屏障中的作用。将研究密切相关的酵母物种的朊病毒蛋白。这些蛋白质之间的变化水平对应于在哺乳动物系统中的物种屏障和跨物种朊病毒传播现象中观察到的分歧范围。将采用体外和体内方法的组合，然后直接比较结果。朊病毒可以在哺乳动物、酵母和其他真菌中找到。 它们控制着酵母和其他真菌的非孟德尔性状的遗传，并为“结构遗传”提供了一种机制，可能在进化中发挥着重要作用。本研究的结果将有助于发展的战略，以防止跨物种的朊病毒在密切相关的动物之间的传播，并可能有深远的影响，了解机制控制的特异性蛋白质-蛋白质相互作用的结构遗传。该研究项目在很大程度上依赖于研究生和本科生的参与，并由一个由男性和女性组成的多元化团队进行。该项目将为本科生和研究生提供综合课程，他们将通过博士项目为该项目做出贡献。PI的实验室参与了多机构基础和应用分子进化中心（FAME），该中心促进了与化学和生物进化相关的各种主题的跨学科研究和教育。新的遗传标记菌株的酵母属物种以外的S。在该项目过程中建造的酿酒酵母将有助于推动对这些生物的进一步研究。