Investigation of the Establishment of the Prion State

朊病毒国家的建立调查

• 批准号: 7267687
• 负责人: IRINA L DERKATCH
• 金额: $ 26.26万
• 依托单位: NEW YORK UNIVERSITY SCHOOL OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-08-01 至 2011-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7267687
• 关键词: Algorithms; Amyloid fibers; Amyloidosis; Appearance; Atomic Force Microscopy; Attention; Benign; Biogenesis; Biological; Bovine Spongiform Encephalopathy; Cattle; Cell physiology; Cells; Chronic Wasting Disease; Deer; Development; Disease; Elements; Endopeptidases; Epigenetic Process; Evolution; Goals; Homologous Gene; Homologous Protein; Human; In Vitro; Investigation; Knowledge; Lead; Libraries; Light; Macromolecular Complexes; Messenger RNA; Modeling; Molecular; Molecular Conformation; Monitor; Mutagenesis; Mutation; Names; Neurodegenerative Disorders; Normal Cell; Peptide Hydrolases; Peptide Library; Peptides; Phenotype; Physiological; Play; Prion Diseases; Prions; Process; Protein Conformation; Protein Structure Initiative; Proteins; RNA Splicing; Regulation; Research; Research Personnel; Resistance; Role; Role playing therapy; Saccharomyces; Screening procedure; Seeds; Signal Transduction; Spinal Muscular Atrophy; Structure; System; Testing; Variant; Work; Yeasts; base; comparative; computer data analysis; computerized data processing; deletion analysis; design; fungus; genetic analysis; human disease; in vivo; link protein; mRNA Stability; macromolecular assembly; mutant; novel; prevent; prion biogenesis; prospective; protein aggregate; protein protein interaction; research study; transmission process

DESCRIPTION (provided by applicant): Prions cause tremendous concern as the causative agents of lethal neurodegenerative disorders such as CJD and BSE. Comparative analysis of prions and other proteins linked to human amyloidosis, and the identification of prions among benign epigenetic modifiers of cellular functions in fungi suggest the existence of molecular mechanisms governing the formation and elimination of self-perpetuating protein conformations. Revealing these mechanisms is expected to facilitate the development of new strategies for preventing the onset and transmission of prion diseases. The long-term goal of PI's research is to understand the biogenesis and biological role of prions, whereas the objective of the current proposal is to elucidate the mechanism of initial establishment of prion state. Genetic analysis in yeast is the major approach. The first indication that the de novo formation of prions may not be truly spontaneous came from the finding that the [PSI+] prion appears only in the presence of a prion named [PIN+] (for [PSI+] inducibility). The demonstrations that various prions can substitute for [PIN+] during [PSI+] induction and that formation of prions other than [PSI+] can be similarly facilitated by heterologous prions suggest that there is a general mechanism through which pre-existing prion aggregates seed novel prions. Aim 1 is to elucidate this mechanism. The working hypothesis postulates that the formation of [PSI+] is initiated by the formation of a short "polar zipper" between Q/N-rich sequences in the prion domains of [PSI+] and [PIN+]-composing proteins, Sup35 and Rnq1. The initial deletion analysis of Rnq1 will determine, which of multiple Q/N-rich regions of Rnq1 are involved in the facilitation of the [PSI*] formation by [PIN*]. Then mutagenesis will be directed to these region(s). Screening for compensatory Sup35 mutations that restore disrupted seeding will be used validate the hypothesis. The conclusions of in vivo studies will be verified in vitro with bacterially expressed proteins, and atomic force microscopy will be used to monitor the seeding process. Also, the efficiency of interconversion of prions encompassing Sup35 prion domains from different Saccharomycetes species will be analyzed to test if the seeding model is applicable to prion transmission between species. Aim 2 is to identify new prions. Strategy (A) will test whether the proteins that satisfy several previously established criteria for presumptive prions can indeed take on self-propagating conformations. Among them, Lsm4 and Yck1 play roles in mRNA processing and signaling, respectively, and their ability to become prions is expected to advance our understanding of splicing, mRNA stability and signal transduction. Lsm4 studies may also shed light on the mechanism of spinal muscular atrophy. Strategy (B) involves designing several libraries of peptides. Screening these peptides for the ability to form prions or promote prion formation, and subsequent computer analysis of data will allow finding an algorithm for aggregation-prone proteins. Such proteins may not cause detectable prion phenotypes themselves but may induce dangerous prions.

描述(由申请人提供)：作为CJD和BSE等致命性神经退行性疾病的病原体，Prion引起了极大的关注。对Prion和其他与人类淀粉样变性相关的蛋白质的比较分析，以及在真菌细胞功能的良性表观遗传修饰物中对Prion的鉴定表明，存在着控制自我永久化蛋白质构象的形成和消除的分子机制。这些机制的揭示预计将有助于制定新的预防普恩病毒疾病发病和传播的战略。PI研究的长期目标是了解Pron的生物发生和生物学作用，而目前的建议的目的是阐明Prion状态的初始建立机制。酵母菌的遗传分析是主要的手段。Prion从头形成的第一个迹象可能不是真正自发的，这是因为发现[PSI+]Prion只有在名为[PIN+]的Prion存在时才出现(为了[PSI+]诱导性)。在[PSI+]诱导过程中，不同的Prion可以替代[PIN+]，而[PSI+]以外的Prion的形成也可以类似地被异源Prion促进，这表明存在一种通用的机制，通过这种机制，先前存在的Prion聚集了新的Prion。目的1是阐明这一机制。工作假说假设，[PSI+]的形成是由[PSI+]和[PIN+]-组成蛋白质Sup35和Rnq1的Pron结构域中富含Q/N的序列之间形成一条短的“极性拉链”启动的。Rnq1的初步缺失分析将确定Rnq1的多个富Q/N区中的哪一个参与了[PIN*]促进[PSI*]的形成。然后将突变定向到这些区域(S)。对恢复中断播种的补偿性Sup35突变的筛选将用于验证这一假说。体内研究的结论将在体外用细菌表达的蛋白质来验证，原子力显微镜将被用来监测播种过程。此外，还将分析来自不同酵母菌物种的包含Sup35蛋白结构域的Prion之间的相互转换效率，以测试种子模型是否适用于Pron在物种之间的传递。目标2是识别新的普恩病毒。策略(A)将测试满足先前建立的几个推定普恩标准的蛋白质是否真的具有自我繁殖的构象。其中，Lsm4和Yock1分别在mRNA的加工和信号转导中发挥作用，它们成为普恩的能力有望促进我们对剪接、mRNA稳定性和信号转导的理解。LSM4研究也可能揭示脊髓性肌萎缩症的机制。策略(B)包括设计几个多肽文库。筛选这些多肽形成Prion或促进Prion形成的能力，以及随后对数据的计算机分析将使找到一种易于聚集的蛋白质的算法。这类蛋白质本身可能不会引起可检测到的普恩表型，但可能会引发危险的普恩。