Characterization of Mitochondrial GTP as an Intramitochondrial Metabolic Signal

描述（由申请人提供）： 2型糖尿病（T2 DM）是一种目前在世界范围内流行的疾病，具有巨大的社会和医疗成本。β-细胞不能充分感知葡萄糖并释放胰岛素是导致糖尿病的关键事件。细胞的线粒体代谢感知血糖水平，因此理解葡萄糖刺激的胰岛素分泌的一个重要问题是如何“感知”线粒体燃料代谢。因此，线粒体“感知”代谢流的分子机制对于理解2型糖尿病的病理生理学至关重要。 根据对高胰岛素血症性低血糖儿科疾病的观察，我假设由TCA循环酶琥珀酰辅酶A合成酶（SCS）产生的线粒体GTP（mtGTP）是将线粒体代谢与胰岛素分泌偶联的重要分子信号。一个关键的见解是，与ATP不同，mtGTP是由TCA循环的每一轮化学计量产生的线粒体内信号。由于其紧耦合速率的TCA循环氧化，mtGTP可能作为一个分子的TCA循环的转速计。通过基因沉默改变mtGTP合成速率，观察到mtGTP和胰岛素分泌之间的直接关系。令人惊讶的是，mtGTP极大地改变了线粒体的功能，从ATP合成到利用代谢能量将钙泵入和泵出线粒体的细胞器。此外，mtGTP信号通过依赖于胞浆钙增加的非经典KATP-独立机制引起胰岛素分泌。由于其孤立的位置，mtGTP可能是一个非常早期的信号依赖于其他介质，以传递其信号的分泌囊泡在胞质溶胶。我在此提议： 1)通过SCS亚型的过表达评估mtGTP在代谢和胰岛素分泌中的作用。 2)评估mtGTP对线粒体回补的调节作用。 3)评估mtGTP如何将线粒体从ATP合成转化为钙和磷酸盐的转运。 本提案中概述的项目不仅旨在回答有关糖尿病的基本问题，而且还确保我作为一名全面的临床研究者的发展，该奖项的支持将提供一个密集的，监督的研究经验，这是成为一名独立研究人员所必需的，在R 01水平上具有竞争力。由于燃料感知是导致胰岛素分泌的基本机制，因此该信号将对理解和治疗T2 DM具有深远的意义。

DESCRIPTION (provided by applicant): Type-2 diabetes (T2DM), a disease that now of epidemic proportions across the world with enormous social and medical costs. Failure of (-cells to adequately sense glucose and release insulin is the critical event leading to diabetes. Mitochondrial metabolism of (-cells senses blood glucose levels and thus a seminal question to understanding glucose stimulated insulin secretion is how mitochondrial fuel metabolism is "sensed." Thus the molecular mechanisms by which mitochondria "sense" metabolic flux are essential to understand the pathophysiology of T2DM. From observations in a pediatric disorder of hyperinsulinemic hypoglycemia I hypothesized that mitochondrial GTP (mtGTP) produced by the TCA cycle enzyme Succinyl CoA Synthetase (SCS) was an essential molecular signal that coupled mitochondrial metabolism to insulin secretion. A key insight was that mtGTP, unlike ATP, is an intramitochondrial signal produced stoichiometrically by each turn of the TCA cycle. By virtue of its tight coupling rates of TCA cycle oxidation, mtGTP might serve as a molecular tachometer for the TCA cycle. By altering mtGTP synthesis rates via gene silencing, a direct relationship between mtGTP and insulin secretion was observed. Surprisingly, mtGTP dramatically altered the function of mitochondria from ATP-synthesis to organelles that harness the energy of metabolism to pump calcium in and out of the mitochondria. Furthermore, the mtGTP signal provoked insulin secretion via a non-canonical KATP-independent mechanism by was dependent upon cytosolic calcium increases. Because of its isolated location, mtGTP is likely to be a very early signal dependent upon other mediators to transmit its signal to the secretory vesicles in the cytosol. Here I propose to: 1) To assess the role of mtGTP in metabolism and insulin secretion by over-expression of SCS isoforms. 2) To assess mtGTP regulation of mitochondrial anaplerosis. 3) To assess how mtGTP converts mitochondria from ATP synthesis to transporting calcium and phosphate. The projects outlined in this proposal are designed to answer not only essential questions about diabetes, but also to ensure my development as a well rounded clinical investigator and support from this award will provide an intensive, supervised research experience that is necessary to be an independent researcher and competitive at the R01 level. As fuel sensing is the essential mechanism that leads to insulin secretion, this signal will have profound implications for understanding and treatment of T2DM.