C. Elegans and Mitochondrial K+ Channels

描述（由申请人提供）：急性心肌梗死是全世界死亡的主要原因，而中风是美国第三大死亡原因。与这些疾病相关的缺血再灌注损伤可导致永久性组织损伤或神经功能缺损。众所周知，短时间的非致死性缺血暴露会引发一种前适应反应，保护细胞免受随后的缺血损伤，这一过程被称为预处理。线粒体是缺血-灌注损伤发病机制的核心，被认为是预处理的主要目标。特别是，线粒体膜内运输钾的离子通道（KATP和KCa通道）被认为可以减弱预处理后的线粒体细胞死亡反应。这些通道的分子特性是有争议的。本提案的主要目标是明确识别线粒体KATP和KCa通道，第二个目标是识别在预处理响应中调节线粒体通道活性的信号过程。为实现这些目标而设计的实验将在线虫C. elegans中进行。最近有研究表明，预处理可以保护这种遗传模式生物免受缺氧损伤和死亡。此外，我们还发现秀丽隐杆线虫在其线粒体中表达功能性的KATP和KCa通道。我们建议结合两位研究者的优势，一位在线粒体生物能量学和心血管生理学方面有丰富的经验，另一位在线虫离子通道生理学方面有丰富的经验，来验证KATP和KCa通道调节是一种进化保守的机制，有助于线虫的预处理。我们将利用秀丽隐杆线虫中大量可用的遗传资源来筛选含有候选基因突变的菌株，以研究纯化线粒体中的通道活性，通过保守信号通路进行通道调节，以及它们的预调节能力。这些实验的结果将提高我们开发针对通道或上游调节因子的保护性疗法的能力，并旨在模仿哺乳动物的预处理效果。

DESCRIPTION (provided by applicant): Acute myocardial infarction is a leading cause of death throughout the world, while stroke is the third leading cause of death in the United States. Ischemia-reperfusion injury associated with these conditions can lead to permanent tissue damage or neurologic deficits. It is well recognized that non-lethal exposure to ischemia for short periods of time however, elicits a proadaptive response that protects cells from subsequent ischemic injury in a process referred to as preconditioning. Mitochondria are central to the pathogenesis of ischemia- perfusion injury and are believed to be a major target for preconditioning. In particular, ion channels in the inner mitochondrial membrane that transport potassium (KATP and KCa channels) are believed to attenuate the mitochondrial cell death response following preconditioning. The molecular identity of these channels is controversial. The major goal of this proposal is to unambiguously identify the mitochondrial KATP and KCa channels, and a second goal is to identify signaling processes that regulate channel activity in the mitochondria in response to preconditioning. The experiments designed to meet these goals will be carried out in the nematode C. elegans. It has recently been shown that preconditioning can protect this genetic model organism from hypoxic injury and death. Moreover, we have found that C. elegans express functional KATP and KCa channels in their mitochondria. We propose to combine the strengths of two investigators, one with extensive experience in mitochondrial bioenergetics and cardiovascular physiology, and the other in nematode ion channel physiology, to test the hypothesis that KATP and KCa channel regulation is an evolutionarily conserved mechanism that contributes to preconditioning in C. elegans. We will utilize the vast array of genetic resources available in C. elegans to screen strains containing mutations in candidate genes for channel activity in purified mitochondria, for channel regulation via conserved signaling pathways, and for their ability to be preconditioned. The results from these experiments will improve our ability to develop protective therapeutics targeted at channels or upstream regulators and designed to mimic the effects of preconditioning in mammals. PUBLIC HEALTH RELEVANCE: Reduced oxygen availability causes cellular damage and death, particularly in neurons (via stroke) or cardiac myocytes (via heart attack). However, brief sub-lethal exposure to low oxygen can lead to proadaptive mechanisms that protect against subsequent decreases in oxygen. This process is called "preconditioning" and acts as an evolutionarily conserved early warning system in all cell types and organisms examined so far. We propose to determine the molecular identity of membrane ion transporters that have been implicated in this proadaptive conditioning via their function in the mitochondria and to study their regulation using the genetic model organism C. elegans. The identification of these molecules will help in the development of new therapies for heart attack and stroke.