ARC REGULATES MITOCHONDRIAL DEATH SIGNALING IN HEART

描述（由申请人提供）：心肌细胞损失是与许多心血管疾病相关的重要病理特征，包括心肌梗死、缺血/再灌注、慢性缺血和心力衰竭。ARC（凋亡抑制因子与CARD/Caspase募集结构域）是最近发现的肌肉特异性细胞死亡抑制因子。它最初被确定为一种与死亡受体（TNFa, Fas）信号传导相关的启动物caspase 8相互作用并调节其活性的蛋白质。我们发表的和初步的数据表明，ARC的心脏保护靶点涉及caspase依赖性和非依赖性途径。这些靶点是两种主要细胞内细胞器的内在死亡信号通路，线粒体和内质网/高尔基复合体。ARC存在于含有这些细胞器的亚细胞部分中，染色显示ARC与新生儿心肌细胞的内质网/发育肌浆网（SR）有关，并在成人心肌纤维中呈横纹线分布，与SR广泛重叠。我们假设ARC与内质网/SR的结构关联及其与线粒体的密切功能关联是其抑制多种内在代谢的能力的原因死亡刺激，通常在死亡信号的早期阶段进行干预。在线粒体中，我们假设ARC干扰BAX的易位/激活，而在ER/SR中，它作为内源性ER应激反应程序保护臂的一个组成部分。为了验证这些假设，我们提出以下具体目标：具体目标#1将定义ARC抑制BAX诱导和BAX相关细胞死亡的分子机制，以及其与线粒体融合/裂变装置关键调节因子相关的功能意义。特异性目标#2将确定ARC是否是内质网应激反应（即未折叠蛋白反应（UPR））的一部分，它在内质网预处理中起什么作用，以及它是否抑制内质网特异性启动物caspase 12。具体的Aim #3将定义ARC在调节ER/SR和线粒体死亡途径之间的凋亡串串中的作用，通过其对缺氧反应中钙动力学和引发剂caspase激活的影响。提出的研究将确定ARC在心脏细胞损伤时线粒体和内质网/高尔基体应激反应管理中的关键作用。

DESCRIPTION (provided by applicant): Cardiac cell loss is a prominent pathological feature associated with a number of cardiovascular disorders, including myocardial infarction, ischemia/reperfusion, chronic ischemia, and heart failure. ARC (Apoptosis Repressor with CARD/Caspase Recruitment Domain) is a recently identified muscle-specific repressor of cell death. It was originally identified as a protein that interacts with and regulates the activity of initiator caspase 8 which is associated with death receptor (TNFa, Fas) signaling. Our published and preliminary data indicate that there are additional targets for cardioprotection by ARC involving both caspase-dependent and -independent pathways. These targets are the intrinsic death signaling pathways of two major intracellular organelles, the mitochondria and the endoplasmic reticulum/Golgi complex. ARC is present in both subcellular fractions containing these organelles and staining reveals an association of ARC with the ER/developing sarcoplasmic reticulum (SR) of neonatal cardiomyocytes and a striated transverse distribution in adult cardiac muscle fibers that overlaps extensively with the SR. We hypothesize that ARCs structural association with the ER/SR and its close functional association with mitochondria are what is responsible for its ability to suppress a variety of intrinsic death stimuli, often intervening at the earliest stages in death signaling. In the mitochondria, we hypothesize that ARC interferes with BAX translocation/activation, while in the ER/SR it acts as an integral component of the protective arm of the endogenous ER stress response program. To test these hypotheses, we propose the following specific aims: Specific Aim #1 will define the molecular mechanism by which ARC inhibits BAX-induced and BAX associatedcell death as well as the functional significance of its association with a critical regulator of the mitochondrial fusion/fission apparatus. Specific Aim #2 will establish whether ARC is part of the ER stress response known as the unfolded protein response (UPR), what role it plays in ER preconditioning, and whether it suppresses the ER-specific initiator caspase 12. Specific Aim #3 will define ARC's role in regulating the apoptotic crosstalk between the ER/SR and mitochondrial death pathways through its effects on calcium dynamics and initiator caspase activation in response to hypoxia. The studies proposed will define the critical role of ARC in the management of the mitochondrial and ER/Golgi stress responses to cellular injury in the heart.