Hsp110 protein chaperone function in yeast

Hsp110 蛋白伴侣在酵母中的功能

基本信息

批准号：
8788364
负责人：
KEVIN ANTHONY MORANO
金额：
$ 29.91万
依托单位：
UNIVERSITY OF TEXAS HLTH SCI CTR HOUSTON
依托单位国家：
美国
项目类别：
财政年份：
2006
资助国家：
美国
起止时间：
2006-04-01 至 2016-12-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8788364
关键词：
ATP Hydrolysis ATP phosphohydrolase Affinity Age Alleles Alzheimer&apos s Disease Ataxia Binding Binding Sites Biochemical Biogenesis Biological Cell physiology Cells Complex Conflict (Psychology)Cytosol Data Defect Disease Dissociation Endoplasmic Reticulum Escherichia coli Eukaryota Eukaryotic Cell Family Funding Genetic Genomic approach Goals Guanine Nucleotide Exchange Factors Heat-Shock Proteins 70 Homeostasis Homologous Gene Human Human Resources Huntington Disease In Vitro Investigation Lead Locales Malignant Neoplasms Mass Spectrum Analysis Mediating Membrane Modeling Modification Molecular Chaperones Mutation Nerve Degeneration Neurodegenerative Disorders Nucleotides Outcome Parkinson Disease Physiological Play Process Protein Family Proteins Publishing Quality Control Recruitment Activity Regulation Relative (related person)Reperfusion Injury Research Ribosomes Role Saccharomyces cerevisiae Saccharomycetales Specificity Stress Testing Therapeutic Intervention Translations Work Yeasts base biochemical tools biological systems chemical genetics cofactor design experience genetic analysis genome-wide heat-shock proteins 110 human disease in vivo mutant novel polypeptide protein folding protein misfolding repaired research study therapeutic target

项目摘要

DESCRIPTION (provided by applicant): In humans, defects in protein folding can lead to neurodegenerative disorders such as Alzheimer's, Parkinson's, and Huntington's disease and debilitating forms of ataxia. The 70 kD heat-shock proteins (Hsp70) are a ubiquitous family of protein chaperones involved in all aspects of protein homeostasis including protein biogenesis, repair and degradation. Hsp70 activity is governed by two types of cofactors; J domain-containing proteins activate the ATP hydrolysis step leading to high affinity substrate binding, while nucleotide exchange factors (NEFs) promote ADP dissociation and polypeptide release. Eukaryotes from yeast to humans express three recently described families of structurally diverse cytosolic NEFs (Hsp110 (Sse1/2 in yeast), HspBP1 (Fes1), and Bag-domain (Snl1)) that perform essentially the same biochemical function. The distinct contributions these proteins make to Hsp70-dependent activities are unknown in any biological system. The primary goal of this proposal is to determine how these NEFs partner with cytosolic Hsp70 chaperones to mediate proteostasis in eukaryotic cells. We hypothesize that while the NEFs share a common ability to stimulate Hsp70 activity, they differentially interact with the Ssa and Ssb families of cytosolic Hsp70 to promote protein folding and repair. In addition, we hypothesize that the Bag homolog Snl1 simultaneously recruits Hsp70 and the ribosome to promote protein biogenesis at the endoplasmic reticulum (ER). Three lines of investigation are proposed to test these hypotheses. In the first aim, we will determine how the apparently principal NEF Hsp110 is integrated into the cytosolic NEF network to promote protein biogenesis and repair. A major component of this aim will be genetic and cell biological experiments to assess the contribution of the Hsp110 substrate binding domain to Hsp70- dependent protein folding in vivo. In contrast to published results that Fes1 binds Ssa and Ssb in vitro, we have obtained preliminary evidence that Fes1 interacts solely with Ssa in vivo. In Aim 2 we will resolve these conflicting findings and determine the mechanism of Fes1-Hsp70 specificity. Lastly, we have discovered that the ER membrane-associated Snl1 binds the ribosome in addition to Hsp70. Aim 3 will be focused on determining the physiological significance of membrane recruitment of the translation machinery by an Hsp70 NEF. These studies represent the first comprehensive analysis of cytosolic Hsp70 NEFs by genetic, biochemical, and genomic approaches. Due to the high degree of conservation of these and other components of the Hsp70 chaperone network in eukaryotes, the results will be directly applicable to understanding the roles of Hsp70 NEFs in protein biogenesis and quality control in human cells.

描述（由申请人提供）：在人类中，蛋白质折叠的缺陷会导致神经退行性疾病，例如阿尔茨海默氏症，帕金森氏症，帕金森氏病，亨廷顿氏病以及共济失调的衰弱形式。 70 kD的热休克蛋白（HSP70）是无处不在的蛋白质伴侣家族，涉及蛋白质稳态的各个方面，包括蛋白质生物发生，修复和降解。 HSP70活性由两种类型的辅助因子控制；含J域的蛋白激活ATP水解步骤，导致高亲和力底物结合，而核苷酸交换因子（NEF）促进ADP解离和多肽释放。从酵母到人类的真核生物表达了三个最近描述的结构多样的胞质NEF（HSP110（酵母中的SSE1/2），HSPBP1（FES1）和Bag-Domain（SNL1））的家族，这些家族基本上具有同一生物化学功能。这些蛋白质对HSP70依赖性活性的独特贡献在任何生物系统中均不清楚。该提案的主要目标是确定这些NEF与胞质HSP70伴侣伴侣如何介导真核细胞中的蛋白质抑制剂。我们假设，尽管NEF具有刺激HSP70活性的共同能力，但它们与胞质HSP70的SSA和SSB家族差异相互作用，以促进蛋白质折叠和修复。此外，我们假设Bag同源性SNL1同时募集HSP70和核糖体以促进内质网（ER）的蛋白质生物发生。提出了三种研究线以检验这些假设。在第一个目标中，我们将确定如何将看似主的NEF HSP110整合到胞质NEF网络中以促进蛋白质的生物发生和修复。该目标的一个主要组成部分是遗传和细胞生物学实验，以评估HSP110底物结合结构域对体内HSP70依赖性蛋白折叠的贡献。与FES1在体外结合SSA和SSB的公开结果相反，我们获得了FES1仅与体内SSA相互作用的初步证据。在AIM 2中，我们将解决这些矛盾的发现，并确定FES1-HSP70特异性的机制。最后，我们发现ER膜相关的SNL1除HSP70外还结合了核糖体。 AIM 3将集中于确定由HSP70 NEF捕获翻译机械的膜募集的生理意义。这些研究代表了通过遗传，生化和基因组方法对胞质HSP70 NEF进行的首次综合分析。由于真核生物中HSP70伴侣网络的这些和其他组件的高度保存，结果将直接适用于理解Hsp70 Nefs在人类细胞中蛋白质生物发生和质量控制中的作用。