New Ubiquitin-Proteasome System Components that Protect against Proteotoxicity

描述（由申请人提供）：细胞生物学中的一个中心问题涉及细胞如何破坏自身蛋白质。蛋白质错误折叠代表了对细胞的普遍威胁，并且有许多原因，包括热，翻译错误，DNA损伤，重金属和类金属，遗传突变，氧化和衰老。蛋白质错误折叠也与大多数神经退行性疾病和某些癌症类型有关。泛素-蛋白酶体系统，凭借其破坏蛋白质的能力，对蛋白质错误折叠的威胁做出反应，有时称为蛋白毒性。我们已经确定了这种应激反应途径的一个新的分支，它特异性地保护细胞免受三价类金属（如砷）诱导的错误折叠蛋白质的伤害，而不是蛋白质错误折叠的其他原因。这一反应途径的关键介质是Cuz 1，以及相关但很大程度上未表征的蛋白质Tmc 1。我们发现Cuz 1作为一种新型的锌依赖性泛素结合蛋白，与蛋白酶体和多功能伴侣Cdc 48/p97相互作用。这些数据表明，Cuz 1通过识别错误折叠的蛋白质并将其传递给蛋白酶体进行破坏来保护细胞免受三价金属的侵害。我工作的长期目标是更好地了解细胞内蛋白质降解是如何发生的。在这个应用程序中，我使用生物化学，遗传学，细胞生物学，蛋白质组学和结构方法的组合来了解Cuz 1和Tmc 1介导的应激反应途径。首先，我将描述Cuz 1的泛素结合功能。我的数据表明一个潜在的新的进化保守的泛素识别基序内Cuz 1。如果得到证实，Cuz 1可能代表了一类新的泛素结合蛋白的创始成员。其次，我将利用蛋白质组学的方法来确定特定的底物Cuz 1。由于Cuz 1在保护细胞免受类金属诱导的蛋白毒性方面具有显著的特异性，因此这些研究可能会深入了解蛋白质降解中最重要的问题之一，这与特异性如何产生和维持有关。第三，我们将进行Tmc 1的首次重要表征，Tmc 1是一种与Cuz 1具有序列和表型相似性的锌指蛋白。我将确定Cuz 1和Tmc 1如何协调它们的保护性应激反应。总之，这些目标将提供对细胞生物学的一个基本方面的见解，调节蛋白质降解，并在这样做将告知与错误折叠蛋白质相关的许多疾病。

DESCRIPTION (provided by applicant): A central question in cell biology concerns how cells destroy their own proteins. Protein misfolding represents a universal threat to cells, and has numerous causes including heat, errors in translation, DNA damage, heavy metals and metalloids, inherited mutation, oxidation, and aging. Protein misfolding is also associated with most neurodegenerative disease and some cancer types. The ubiquitin-proteasome system, by virtue of its ability to destroy proteins, responds to the threat of protein misfolding, sometimes termed proteotoxicity. We have identified a novel arm of this stress response pathway that specifically protects cells from misfolded proteins induced by trivalent metalloids like arsenic, bt not other causes of protein misfolding. The key mediators of this response pathway are Cuz1, and the related but largely uncharacterized protein, Tmc1. We showed that Cuz1 functions as a novel zinc-dependent ubiquitin binding protein that interacts with the proteasome and the multifunctional chaperone Cdc48/p97. These data suggest a model in which Cuz1 protects cells from trivalent metalloids by recognizing misfolded proteins and delivering them to the proteasome for destruction. The long-term goal of my work is to provide a better understanding of how intracellular protein degradation occurs. In this application, I use a combination of biochemical, genetic, cell biologic, proteomic, and structural approaches to understand the stress response pathway mediated by Cuz1 and Tmc1. First, I will characterize the ubiquitin binding function of Cuz1. My data indicate a potentially novel evolutionarily conserved ubiquitin recognition motif within Cuz1. If verified, Cuz1 might represent the founding member of a new class of ubiquitin binding proteins. Second, I will utilize proteomics approaches to identify specific substrates of Cuz1. Because Cuz1 is remarkably specific in protecting cells from metalloid-induced proteotoxicity, these studies may provide insight into one of the most important questions in protein degradation, which relates to how specificity is generated and maintained. Third, we will undertake the first significant characterization of Tmc1, a zinc finger protein with sequence and phenotypic similarity to Cuz1. I will determine how Cuz1 and Tmc1 function in coordinating their protective stress response. Together, these aims will provide insight into a fundamental aspect of cell biology, regulated protein degradation, and in so doing will inform the many diseases associated with misfolded proteins.