Molecular and functional properties of the TRPM2 catioin channel

描述（由申请人提供）：瞬时受体电位（TRP）离子通道家族最近出现，包含参与感觉感知、镁稳态或细胞凋亡等过程的关键通道蛋白。我们确定了TRPM 2，TRP家族的成员，作为一种钙渗透性阳离子通道门控的新的第二信使ADP-核糖（ADPR）通过通道的内在焦磷酸酶结构域。TRPM 2在多种细胞类型中表达，包括胰腺β细胞和嗜中性粒细胞。最近，在ADPR周围的代谢网络中的分子对TRPM 2的功能特性的作用已经出现。在这里，新型钙释放化合物cADPR和其他腺嘌呤二核苷酸成为TRPM 2生理学的关键因素，因此涉及细胞钙稳态的过程。基于其主要由ADPR和活性氧激活，TRPM 2可能在细胞应激中具有功能，并且仍然是参与凋亡过程的主要候选者。为了进一步了解TRPM 2在细胞生理学中的作用，我们提出了具体目标1中的实验，该实验将专注于TRPM 2在腺嘌呤核苷酸引发的事件中的功能。我们假设TRPM 2可能是利用TRPM 2过表达细胞系统和TRPM 2敲除小鼠模型作为钙内流和钙释放的双重通道。我们将探讨激动剂-受体相互作用参与产生腺嘌呤二核苷酸，即CD 38的主要候选人。我们将使用基于链脲佐菌素的1型糖尿病小鼠模型研究TRPM 2参与细胞死亡。将使用膜片钳、成像和凋亡研究的组合，在表达系统和从野生型、CD 38、PARP和TRPM 2敲除小鼠模型分离的小鼠中性粒细胞和β细胞中进行这些实验。在具体目标2中，我们将研究TRPM 2激动剂结合的分子决定因素，并解决钙调蛋白是否是TRPM 2的钙依赖性促进的介体的问题。了解TRPM 2和腺嘌呤二核苷酸在钙信号传导中的功能将完善我们对表达这种离子通道的组织中钙调节的知识，并可能为涉及TRPM 2功能的过程中的治疗干预提供新的靶点，包括糖尿病引起的胰腺β细胞细胞的细胞死亡。

DESCRIPTION (provided by applicant): The transient receptor potential (TRP) ion channel family recently emerged as harboring crucial channel proteins involved in processes such as sensory perception, magnesium homeostasis or apoptosis. We identified TRPM2, a member of the TRP family, as a calcium-permeable cation channel gated by the novel second messenger ADP-ribose (ADPR) through the channel's intrinsic pyrophosphatase domain. TRPM2 is expressed in a variety of cell types, including pancreatic beta cells and neutrophils. Recently, the role of the molecules in the metabolic network surrounding ADPR on the functional properties of TRPM2 have emerged. Here, the novel calcium-release compound cADPR and other adenine dinucleotides materialize as crucial factors in TRPM2 physiology and hence processes involving cellular calcium homeostasis. Based on its primary activation by ADPR and reactive oxygen species, TRPM2 may have a function in cellular stress and remains a prime candidate for being involved in the apoptotic process. To further our understanding of TRPM2 in cell physiology, we propose experiments in Specific Aim 1 that will focus on the function of TRPM2 in events initiated by adenine-nucleotides. We hypothesize that TRPM2 may function as dual calcium influx and calcium release channel by taking advantage of a TRPM2 overexpressing cell system and a TRPM2 knock-out mouse model. We will explore agonist-receptor interactions engaging the main candidate in generating adenine dinucleotides, namely CD38. We will investigate the involvement of TRPM2 in cell death using a streptozotocin-based mouse model for diabetes type-1. These experiments will be performed in an expression system and mouse neutrophils and beta cells isolated from wild-type, CD38, PARP and TRPM2 knock-out mice models using a combination of patch-clamp, imaging and apoptotic studies. In Specific Aim 2 we will investigate the molecular determinants of TRPM2 agonist binding and address the question whether calmodulin is the mediator of the calcium-dependent facilitation of TRPM2. Understanding TRPM2 and adenine-dinucleotide function in calcium signaling will refine our knowledge of calcium regulation in tissues expressing this ion channel and may provide new targets for therapeutic interference in processes involving TRPM2 function, including diabetes-causing cell death of pancreatic beta cells.