CHIP: A link between the chaperone and proteasome system

描述（由申请人提供）：70-和90-kDa热休克蛋白（Hsps）最为人所知的是其作为分子伴侣的作用，参与新生蛋白质的折叠、成熟和适当的亚细胞靶向，或者相反地，恢复已被错误折叠或以其他方式被细胞应激源损伤的蛋白质的功能。然而，热休克蛋白参与各种过程-细胞内信号事件，增殖，分化和蛋白质的靶向降解-似乎与蛋白质折叠仅间接相关，或根本不相关。与此同时，越来越明显的是，分子伴侣并不代表细胞应激反应的总和，并且需要蛋白质重折叠以外的一系列事件来维持活力并在不利情况下保护细胞内环境。1999年，我们在筛选应激反应基因时鉴定了CHIP（Hsc 70相互作用蛋白的羧基末端）。从那时起，我们发现CHIP与伴侣Hsc 70，Hsp 70和Hsp 90相互作用，对它们的功能具有复杂和协调的影响。此外，CHIP具有泛素连接酶活性，并在调节细胞质内的蛋白质质量控制中起关键作用。最后，我们发现使用体外和体内方法，CHIP是细胞和有机体应激反应的中央调节器。令人惊讶的是，这些作用中至少有一些似乎与CHIP对转录、信号传导和代谢反应的特定作用有关。这项资助的目的是作为我们最初筛选参与分子伴侣事件的蛋白质的逻辑延伸。为了实现我们的目标，我们已经制定了一个新的和高度集成的方法，使用在体外和体内测定。本提案的目的是：（1）。使用联合转录组/蛋白质组分析检查CHIP对细胞应激背景下mRNA和蛋白质表达的总体变化的影响;（2）表征CHIP对细胞信号传导和代谢反应的作用;（3）评估CHIP对体内病理生理应激的综合作用。本提案的范围旨在使用最先进的分子生物学技术解决相关的生物学和生理学问题。从这个建议中获得的知识应该提供关于这些不同的途径如何调节在泛素/蛋白酶体系统控制下的生理和病理过程的关键信息，并且可以为治疗涉及该系统的疾病提供潜在的治疗靶点。 公共卫生相关性：正确的蛋白质折叠对于最佳蛋白质性能和正常细胞功能至关重要。在新蛋白质的合成和变性蛋白质的重折叠过程中，细胞的分子伴侣和其降解机制之间必须发生合作，因为一些蛋白质不能自发地获得其正确的三级构象。相互排斥的折叠和降解途径构成了细胞的蛋白质质量控制系统。伴侣蛋白和泛素-蛋白酶体系统在调节健康和疾病中的应激反应信号和细胞保护机制中起关键作用。我们的总体目标是确定CHIP（一种分子伴侣）的分子和生理功能如何协调以协调其应激反应能力。这些研究将有助于我们建立细胞质质量控制机制与细胞应激反应之间关系的新模型。

DESCRIPTION (provided by applicant): 70- and 90-kDa heat shock proteins (Hsps) are best known by their roles as molecular chaperones, participating in the folding, maturation, and proper subcellular targeting of nascent proteins or, conversely, the recovery of function of proteins that have been misfolded or otherwise damaged by cellular stressors. However, Hsps participate in a variety of processes- intracellular signaling events, proliferation, differentiation, and targeted degradation of proteins- that seem related to protein folding only indirectly, or not at all. At the same time, it is becoming increasingly apparent that molecular chaperones do not represent the sum total of the cell stress response, and that a range of events other than protein refolding are required to maintain viability and protect the intracellular environment in adverse circumstances. We identified CHIP (carboxyl-terminus of Hsc70-interacting protein) in a screen for stress-responsive genes in 1999. Since then, we have found that CHIP interacts with the chaperones Hsc70, Hsp70, and Hsp90 and has complex and coordinated effects on their functions. In addition, CHIP has ubiquitin ligase activity and plays a critical role in regulating protein quality control within the cytoplasm. Finally, we have found using both in vitro and in vivo approaches that CHIP is a central regulator of cellular and organismal stress responses. Surprisingly, at least some of these effects seem to be related to specific effects of CHIP on transcriptional, signaling, and metabolic responses. The aims of this grant are intended as a logical extension of our initial screen for proteins participating in molecular chaperone events. To accomplish our aims, we have formulated a novel and highly integrated approach using both in vitro and in vivo assays. The aims of this proposal are to: (1). Examine the consequences of CHIP on global changes in mRNA and protein expression in the context of cellular stress using a combined transcriptome/proteome analysis; (2) Characterize the role of CHIP on cell signaling and metabolic responses and (3) Evaluate the integrated effects of CHIP on pathophysiologic stress in vivo. The scope of this proposal is intended to address relevant biological and physiological questions using state of the art molecular biology techniques. Knowledge gained from this proposal should provide crucial information about how these various pathways regulate physiologic and pathologic processes that are under control of the ubiquitin/proteasome system and may provide potential therapeutic targets for treatment of diseases where this system is involved. PUBLIC HEALTH RELEVANCE: Proper protein folding is essential for optimum protein performance and normal cellular function. During synthesis of new proteins and refolding of denatured proteins, cooperation between the cell's molecular chaperones and its degradation machinery must occur because some proteins cannot attain their correct tertiary conformation spontaneously. The mutually exclusive pathways of folding and degradation constitute the cell's protein quality control system. The chaperone and ubiquitin-proteasome systems play critical roles in regulating stress-responsive signaling and cellular protective mechanisms in health and disease. Our overall goal is to determine how the molecular and physiologic functions of CHIP, a molecular chaperone, are coordinated to orchestrate its stress response capabilities. These studies will help us develop new models about the relationship between cytoplasmic quality control mechanisms and the cellular stress response.