Functional Analysis of Rad23 Protein

描述（由申请人提供）：尽管在了解核苷酸切除修复（NER）的生化基础方面取得了重大进展，但对于患有色素性干皮病、科凯恩综合征、范可尼贫血和其他DNA修复缺陷的个体尚无治愈方法。目前的治疗重点是缓解症状和提高生活质量。细胞间蛋白降解的泛素/蛋白酶体系统（UPS）已被广泛研究，涉及细胞周期控制、应激反应、信号转导和DNA修复。我们确定了NER蛋白Rad4是高度不稳定的，并且被蛋白酶体迅速降解。值得注意的是，许多额外的DNA修复蛋白具有蛋白质降解功能，包括Rad6、Rad7、Rad16和FANCL。在过去的15年里，我的实验室一直在研究蛋白质降解在DNA修复中的作用。我们确定，当Rad4与DNA修复因子Rad23形成复合体时，它是稳定的。Rad23也可以结合蛋白酶体并传递泛素化底物。我们最近发现，蛋白酶体在细胞核外周的定位是DNA修复所必需的。本文提出的研究将采用遗传、生化和细胞生物学的方法来研究UPS在核苷酸切除修复中的作用。我们将描述运输蛋白酶体的蛋白质，并确定包括Rad4在内的核底物如何靶向蛋白酶体。这些机制研究预计将产生广泛的影响，因为它们将表明蛋白酶体运输代表了细胞内蛋白质降解中一个以前未知的调节过程。

DESCRIPTION (provided by applicant): Despite significant advances in understanding the biochemical basis of nucleotide excision repair (NER), there is no cure for individuals with Xeroderma pigmentosum, Cockayne Syndrome, Fanconi Anemia, and other DNA repair defects. Current therapy is focused on relieving symptoms and improving quality of life. The ubiquitin/proteasome system (UPS) of interacellular protein degradation has been extensively studied, and is implicated in cell cycle control, stress response, signal transduction, and DNA repair. We determined that the NER protein Rad4 is highly unstable, and is rapidly degraded by the proteasome. Remarkably, a number of additional DNA repair proteins have protein degradation functions, including Rad6, Rad7, Rad16 and FANCL. My laboratory has, for the past 15 years, investigated the role of protein degradation in DNA repair. We determined that Rad4 is stabilized when it forms a complex with the DNA repair factor, Rad23. Rad23 can also bind the proteasome and deliver ubiquitinated substrates. We recently discovered that the localization of proteasomes at the nuclear periphery is required for DNA repair. The studies proposed here will use genetic, biochemical and cell biological approaches to investigate the role of the UPS in nucleotide excision repair. We will characterize the proteins that traffic proteasomes, and determine how nuclear substrates, including Rad4, are targeted to the proteasome. These mechanistic studies are expected to have a broad impact, as they will show that proteasome trafficking represents a previously unknown regulatory process in intracellular protein degradation.