Molecular Mechanisms of Mitochondrial Uncoupling and Thermogenesis

项目摘要 通过底物氧化，线粒体产生跨越线粒体内膜的高电位 （IMM）。这种电位的能量通常由线粒体ATP合酶转化为ATP，但线粒体ATP合酶的作用是通过线粒体ATP合成酶的作用来实现的。 由于存在穿过IMM的H+泄漏，部分作为热耗散。这种线粒体H+泄漏是 介导的特殊蛋白质的IMM，如解偶联蛋白1（UCP 1），它具有重要的 生理功能。它控制着身体的新陈代谢效率，帮助支撑核心身体 温度，并减少线粒体产生的活性氧，以防止氧化 损害线粒体H+泄漏被认为是预防肥胖的重要机制， 糖尿病和与年龄相关的退行性疾病以及针对病理状况， 线粒体氧化应激，如缺血-再灌注。尽管它的生理和病理生理 重要的是，线粒体H+泄漏仍然知之甚少，主要是由于缺乏直接的方法 我们最近开发了一种方法，可以消除这种技术障碍， 直接膜片钳记录来自整个IMM的H+漏电流。这种方法帮助我们解决了长期- 关于UCP 1依赖的产热H+泄漏穿过IMM的机制的长期问题， 棕色脂肪在本申请中，我们建议使用膜片钳技术来进一步表征 线粒体H+泄漏在几种组织中起重要作用的产热和能量代谢。 该提案的具体目标是：1）鉴定负责线粒体H+泄漏的蛋白质， 非脂肪组织; 2）表征脂肪酸激活的线粒体H+泄漏的机制， 腺嘌呤核苷酸转运子（ANT）; 3）表征线粒体H+泄漏的机制， 质子载体DNP和FCCP。这些具体目标的实现将有助于我们阐明 调节代谢效率和产热的机制，这些知识将有助于 开发治疗干预措施，以控制肥胖、糖尿病和与年龄相关的退行性疾病。

Project Summary Through substrate oxidation, mitochondria generate a high potential across the inner mitochondrial membrane (IMM). The energy of this potential is usually converted into ATP by the mitochondrial ATP synthase, but a fraction is dissipated as heat due to the presence of a H+ leak across the IMM. This mitochondrial H+ leak is mediated by specialized proteins of the IMM, such as uncoupling protein 1 (UCP1), and it has important physiological functions. It controls the metabolic efficiency of the body, helps to support the core body temperature, and reduces mitochondrial production of reactive oxygen species to protect against oxidative damage. The mitochondrial H+ leak is considered to be important in protective mechanisms against obesity, diabetes, and age-related degenerative disorders as well as against pathological conditions involving mitochondrial oxidative stress such as ischemia-reperfusion. Despite its physiological and pathophysiological significance, the mitochondrial H+ leak remains poorly understood, primarily due to the lack of direct methods to study it. We recently developed a method that removes this technical barrier and for the first time allows direct patch-clamp recording of H+ leak currents from the whole IMM. This method helped us resolve long- standing questions about the mechanism of the UCP1-dependent thermogenic H+ leak across the IMM of brown fat. In this application, we propose to use the patch-clamp technique to further characterize the mitochondrial H+ leak in several tissues that play important roles in thermogenesis and energy metabolism. The specific aims of this proposal are to: 1) identify the protein(s) responsible for the mitochondrial H+ leak in non-adipose tissues; 2) characterize the mechanism of the fatty acid-activated mitochondrial H+ leak via the adenine nucleotide translocator (ANT); 3) characterize the mechanism of the mitochondrial H+ leak induced by protonophores DNP and FCCP. Accomplishment of these specific aims will help us elucidate the principal mechanism that regulates metabolic efficiency and thermogenesis, and such knowledge will aid the development of therapeutic interventions to control obesity, diabetes, and age-related degenerative disorders.