The Role of Apoptosis Repressor with Caspase Recruitment Domain (ARC) in Amyloid-Induced Beta-cell Apoptosis

 描述（由申请方提供）：胰岛淀粉样蛋白形成是2型糖尿病的标志，与β细胞凋亡和β细胞质量损失相关。先前的研究已经确定c-Jun N-末端激酶（JNK）、Bcl-2相关死亡启动子（Bad）和Bcl-2相关X蛋白（Bax）在淀粉样蛋白诱导的β细胞凋亡中具有重要的促凋亡作用。相反，最近在β-细胞中鉴定的新型抗凋亡蛋白，具有半胱天冬酶募集结构域（ARC）的凋亡阻遏物的作用还没有很好地理解。因此，我建议研究ARC在淀粉样蛋白诱导的β细胞凋亡中的作用。这项工作的目标是获得ARC的知识，这将导致新的策略来预防β细胞损失，这是2型糖尿病发展的关键组成部分。我提出了一个模型，ARC拮抗β细胞中三个众所周知的促凋亡蛋白，即JNK，Bad和Bax的作用。证实在正常情况下，ARC抑制JNK活化并隔离Bad和Bax。在淀粉样蛋白形成的情况下，ARC不再抑制JNK活化，并且不能隔离Bad和Bax，导致β细胞凋亡。在本申请中，我提出了三个具体目标来解决这个假设。具体目的1：确定ARC在β细胞中的抗凋亡作用是否部分通过抑制JNK途径活化介导。具体目的2：确定Bad和Bax的ARC隔离是否有助于其在β细胞中的抗凋亡作用。具体目标3：确定ARC过表达是否改善胰岛移植后hIAPP转基因胰岛移植物存活。我将使用来自胰岛淀粉样蛋白转基因小鼠模型的胰岛、野生型小鼠胰岛和永生化β细胞系来进行所提议的研究。这一三年计划是一项全面的培训建议，侧重于科学和专业发展目标。研究将扩大目前对β细胞凋亡的理解，同时促进获得新的技能，如胰岛的腺病毒转导、免疫组织化学、定量显微镜、邻位连接测定和胰岛移植。卡恩博士的科学 方向，实验室的能力，并承诺卓越的指导将有助于这个项目的成功完成，并对我的职业发展。参加研讨会，实验室会议，期刊俱乐部，区域和国家会议，以及由华盛顿糖尿病研究中心（DRC）大学赞助的研究活动将有助于全面的培训和职业发展经验。总之，这些活动将使我能够成功地竞争职业发展（K）奖，并朝着我成为胰岛生物学独立研究者的目标前进。

 DESCRIPTION (provided by applicant): Islet amyloid formation is a hallmark of type 2 diabetes that is associated with β-cell apoptosis and loss of β-cell mass. Prior studies have determined that c-Jun N-terminal kinase (JNK), Bcl-2-associated death promoter (Bad), and Bcl-2-associated X protein (Bax) have important pro-apoptotic roles in amyloid-induced β- cell apoptosis. Conversely, the role of a novel anti-apoptotic protein that was recently identified in the β-cell, apoptosis repressor with caspase recruitment domain (ARC), is not well understood. Therefore, I propose to study ARC's role in amyloid-induced β-cell apoptosis. The goal of this work is to gain knowledge of ARC which will lead to novel strategies to prevent β-cell loss, a critical component of the development of type 2 diabetes. I propose a model whereby ARC antagonizes the actions of three well-known pro-apoptotic proteins in the β-cell, namely JNK, Bad and Bax. I posit that under normal conditions, ARC represses JNK activation and sequesters Bad and Bax. In the setting of amyloid formation, ARC no longer represses JNK activation, and is unable to sequester Bad and Bax, leading to β-cell apoptosis. In this application I propose three specific aims to address this hypothesis. Specific Aim 1: Determine whether the anti-apoptotic effect of ARC in the β-cell is mediated in part by inhibition of JNK pathway activation. Specific Aim 2: Determine whether ARC sequestration of Bad and Bax contributes to its anti-apoptotic effect in the β-cell. Specific Aim 3: Determine whether ARC overexpression improves hIAPP transgenic islet graft survival following islet transplantation. I will use islets from a transgenic mouse model of islet amyloid, wild type mouse islets, and an immortalized β-cell line to perform the proposed studies. This three-year plan is a comprehensive training proposal focused on scientific and professional development objectives. Studies will expand the current understanding of β-cell apoptosis, while facilitating the acquisition of new skills such as adenoviral transduction of islets, immunohistochemistry, quantitative microscopy, proximity ligation assay, and islet transplantation. Dr. Kahn's scientific direction, laboratory capabilities, and commitment to exceptional mentoring will contribute to the successful completion of this project, and to my career development. Participation in seminars, lab meetings, journal club, regional and national meetings, and research events sponsored by the University of Washington Diabetes Research Center (DRC) will contribute to a comprehensive training and career development experience. Together, these activities will enable me to successfully compete for a Career Development (K) Award and progress towards my goal of becoming an independent investigator of islet biology.