Mechanisms of regulation of mitochondrial H+ leak and thermogenesis

项目摘要 线粒体通过将营养物质转化为质子（H+）的电化学梯度来控制细胞代谢 穿过线粒体内膜（IMM）产生ATP，细胞的货币，和热量（称为 线粒体产热）。H+在两种能量形式之间的精确平衡 生产，ATP和热量，定义了细胞的代谢稳态。棕色脂肪和米色脂肪线粒体 专门通过解偶联蛋白1（UCP 1）产生热量。然而，即使在其他组织中， 线粒体产热占线粒体总能量产生的25%，因此可以具有 对整个身体的生理产生了相当大的影响。线粒体产热作用不仅对 维持核心体温，这也是燃烧多余卡路里以防止饮食的过程， 诱发性肥胖此外，它减少了线粒体产生活性氧（ROS）， 保护细胞免受氧化损伤。此外，化学解偶联剂如2，4-二硝基苯酚（DNP）， 被认为独立于蛋白质增加H+泄漏，是迄今为止最有效的抗肥胖药物。 因此，线粒体产热是细胞代谢的有力调节剂，并且是细胞代谢的机制。 了解这一基本过程将有助于制定治疗策略， 对抗许多与线粒体功能障碍相关的病理，包括代谢综合征 和与年龄有关的疾病不幸的是，控制急性激活的精确分子机制 线粒体中的产热作用还不清楚。这种信息的缺乏主要是由于缺乏 直接测量IMM上H+电流的方法。制定一种方法， 膜片钳技术首次允许直接研究通过每个组织的IMM的H+泄漏， 线粒体转运蛋白（如UCP 1和ADP/ATP转运蛋白）的第一个生物物理特征 (AAC)，它们是这种H+泄漏的介质。这种独特的方法现在提供了前所未有的高- 分辨率直接功能分析1）线粒体离子通道和转运蛋白负责 线粒体产热和2）化学解偶联剂如DNP的作用机制。使用 新的线粒体膜片钳分析结合现代细胞和分子技术，这 研究项目将提供新的见解控制产热能力的机制， 线粒体以及如何将它们作为治疗目标。

Project Summary Mitochondria control cell metabolism by converting nutrients into an electrochemical gradient of protons (H+) across the inner mitochondrial membrane (IMM) to generate ATP, the currency of the cell, and heat (called mitochondrial thermogenesis). A precise balance in the distribution of H+ between the two forms of energy production, ATP and heat, defines the metabolic homeostasis of the cell. Brown fat and beige fat mitochondria specialize in the production of heat via the uncoupling protein 1 (UCP1). However, even in other tissues, mitochondrial thermogenesis accounts for 25% of total mitochondrial energy production and can therefore have a considerable impact on the physiology of the entire body. Mitochondrial thermogenesis is not only essential for maintaining core body temperature, it is also the process by which excess calories are burned to prevent diet- induced obesity. In addition, it reduces the production of reactive oxygen species (ROS) by the mitochondria to protect cells from oxidative damage. In addition, chemical uncouplers such as 2,4-dinitrophenol (DNP), which are believed to increase H+ leak independently of proteins, are the most effective anti-obesity drugs to date. Thus, mitochondrial thermogenesis is a powerful regulator of cellular metabolism, and a mechanistic understanding of this fundamental process will help in the development of therapeutic strategies to combat many pathologies associated with mitochondrial dysfunction, including metabolic syndrome and age-related disorders. Unfortunately, the precise molecular mechanisms that control the acute activation of thermogenesis in the mitochondria are poorly defined. This lack of information is largely due to a dearth of methods for direct measurement of H+ currents across the IMM. The development of a methodology based on the patch-clamp technique allows for the first time the direct study of H+ leak through the IMM of each tissue and the first biophysical characterization of mitochondrial transporters, such as UCP1 and the ADP/ATP transporter (AAC), which are the mediators of this H+ leak. This unique approach now provides an unprecedented high- resolution direct functional analysis of 1) the mitochondrial ion channels and transporters responsible for mitochondrial thermogenesis and 2) the mechanisms of action of chemical uncouplers such as DNP. Using the new mitochondrial patch-clamp assay combined with modern cellular and molecular techniques, this research project will provide new insights into the mechanisms that control the thermogenic capacity of the mitochondria and how they can be targeted for therapeutic purposes.