Degradation mechanisms for inhibitor of apoptosis proteins and their antagonists

凋亡蛋白抑制剂及其拮抗剂的降解机制

基本信息

批准号：
9274332
负责人：
Shawn B Bratton
金额：
$ 32.43万
依托单位：
UNIVERSITY OF TX MD ANDERSON CAN CTR
依托单位国家：
美国
项目类别：
财政年份：
2015
资助国家：
美国
起止时间：
2015-09-01 至 2019-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9274332
关键词：
Adaptor Signaling Protein Address Apoptosis Apoptosis Inhibitor Apoptosis Regulation Gene Applications Grants BIRC4 gene Baculoviridae Baculoviruses Binding Biochemical Biochemical Genetics Biological CASP5 gene Caspase Caspase Inhibitor Cell Death Cells Complex Copper Cysteine Proteinase Inhibitors Data Degenerative Disorder Development Disease Drosophila genus Drosophila inhibitor of apoptosis 1 Excision Exposure to Feedback Genetic Models Goals Growth Factor Hepatolenticular Degeneration Human In Vitro Inflammation Injury Link Lysine Malignant Neoplasms Mammals Mediating Mediator of activation protein Modeling Molecular Mutation N-terminal Nerve Degeneration Neurodegenerative Disorders Neuronal Injury Neurons Organism Pathologic Pathway interactions Patients Play Post-Translational Protein Processing Predisposition Process Protease Inhibitor Proteins Radio Regulation Resistance Role Site Source Therapeutic Ubiquitination Wilson disease protein Withdrawal Work biochemical model cancer cell cancer therapy chemotherapy feeding fly gain of function genetic approach human disease in vivo inhibitor-of-apoptosis protein insight loss of function multicatalytic endopeptidase complex novel overexpression protein function public health relevance tumor ubiquitin-protein ligase virtual

项目摘要

DESCRIPTION (provided by applicant): Apoptosis, or programmed cell death, is broadly conserved in multicellular organisms from flies to humans and plays critical roles in everything from cancer to inflammation and neurodegeneration. Inhibitor of apoptosis (IAP) proteins, including Drosophila IAP1 (DIAP1) and X-linked IAP (XIAP), are E3 ubiquitin ligases that play major roles in the regulation of apoptosis, at least in part through direct inhibition and/or ubiquitination of caspases. IAP antagonists, such as Reaper, Hid, Grim, and Smac, are thought to induce cell death by displacing active caspases from IAPs, thereby leading to increased caspase activity and cell death, but are themselves targets of ubiquitination. Indeed, we have recently discovered that the IAP antagonist Grim is ubiquitinated by DIAP1; however, the lysine targeted for ubiquitination is also subject to removal by caspases, thereby enhancing Grim's stability and initiating a feed-forward caspase amplification loop that results in greater cell deah. As a result of this work and additional preliminary data, we have begun to appreciate and hypothesize that IAPs, IAP antagonists, and caspases reciprocally regulate one another through highly novel mechanisms that impact the function and turnover of IAPs and their antagonists in cells. A major goal of this grant application is to characterize in molecular detail the biochemica mechanisms through which IAPs and IAP antagonists are selectively degraded in order to develop strategies to inhibit or promote cell death in a given pathological context. Recent studies in flies have demonstrated that caspase cleavage of DIAP1 at its N-terminus renders DIAP1 susceptible to degradation through the so-called N-end rule pathway. We have uncovered evidence that caspase cleavage of DIAP1 and XIAP, at a second site, generates IAP fragments that can serve as novel carrier molecules, targeting IAP antagonists for transubiquitination by Ubr-type E3 ligases. In aim #1, we will evaluate the role of the N-end rule pathway in mediating the degradation of IAP antagonists, utilizing a variety of biochemical and genetic approaches. In additional preliminary data, we have discovered that exposure to excess copper modifies DIAP1 and XIAP, rendering them susceptible to caspase cleavage, autoubiquitination, and turnover by the proteasome. In aim #2, we will elucidate the biochemical and structural effects of copper on IAP function and will determine if copper induces cell death through degradation of DIAP1 in a fly model of Wilson's Disease. Finally, we have discovered that the highly unusual initiator caspase Strica is a primary mediator of DIAP1 cleavage. As virtually nothing is known about this caspase, in Aim #3, we will characterize it in biochemical and structural detail and will assess its role in mediating copper-induced degradation and apoptosis. Overall, the proposed studies will provide significant insight into the complex relationships that exist between IAPs, IAP antagonists, and caspases and will inform our efforts at targeting these proteins for the treatment of cancer and various neurodegenerative diseases.

描述（由申请人提供）：细胞凋亡或程序性细胞死亡在从果蝇到人类的多细胞生物中广泛存在，并且在从癌症到炎症和神经变性（IAP）蛋白（包括果蝇 IAP1 (DIAP1)）的各个方面发挥着关键作用。 ) 和 X-linked IAP (XIAP) 是 E3 泛素连接酶，至少部分通过直接抑制在细胞凋亡调节中发挥重要作用IAP 拮抗剂（例如 Reaper、Hid、Grim 和 Smac）被认为通过取代 IAP 中的活性 caspase 来诱导细胞死亡，从而导致 caspase 活性增加和细胞死亡，但它们本身也是泛素化的目标。事实上，我们最近发现 IAP 拮抗剂 Grim 被 DIAP1 泛素化，然而，赖氨酸是泛素化的目标；也可被半胱天冬酶去除，从而增强 Grim 的稳定性并启动前馈半胱天冬酶扩增循环，从而导致更大的细胞死亡。根据这项工作和其他初步数据，我们已经开始认识到 IAP、IAP。拮抗剂和半胱天冬酶通过影响 IAP 及其拮抗剂在细胞中的功能和周转的高度新颖的机制相互调节。这项资助申请的一个主要目标是表征分子细节。最近对果蝇的研究表明，IAP 和 IAP 拮抗剂选择性降解的生化机制，以制定抑制或促进细胞死亡的策略。通过所谓的 N 端规则途径，我们发现了半胱天冬酶在第二个位点切割 DIAP1 和 XIAP 产生 IAP 的证据。可以作为新型载体分子的片段，通过 Ubr 型 E3 连接酶靶向 IAP 拮抗剂进行转泛素化在目标#1 中，我们将利用多种方法评估 N 端规则途径在介导 IAP 拮抗剂降解中的作用。在其他初步数据中，我们发现暴露于过量的铜会改变 DIAP1 和 XIAP，使它们容易受到半胱天冬酶裂解、自动泛素化和半胱天冬酶转换的影响。在目标 2 中，我们将阐明铜对 IAP 功能的生化和结构影响，并确定铜是否通过威尔逊氏病果蝇模型中 DIAP1 的降解诱导细胞死亡。 caspase Strica 是 DIAP1 裂解的主要介体，由于对该 caspase 几乎一无所知，因此在目标 #3 中，我们将在生化和结构细节方面对其进行表征。总体而言，拟议的研究将为 IAP、IAP 拮抗剂和半胱天冬酶之间存在的复杂关系提供重要见解，并将为我们针对这些蛋白质治疗癌症和癌症的努力提供信息。各种神经退行性疾病。