Regulation of Energy Metabolism by PDP1 and PDP2

描述(申请人提供)：本项目的长期目标是了解线粒体丙酮酸脱氢酶复合体(PDC)调控的分子机制。由PDC催化的反应在一般代谢中起着核心作用，为柠檬酸循环、脂肪生成和胆固醇生物合成等主要能量生成和生物合成途径提供碳源。哺乳动物PDC的活性是通过丙酮酸脱氢酶激酶和丙酮酸脱氢酶磷酸酶催化的可逆的磷酸化/去磷酸化循环来调节的。越来越多的证据有力地表明，控制PDC的磷酸化状态在饥饿、代谢性酸中毒、缺血和糖尿病中变得至关重要。这一应用的主要目的是了解PDC在磷酸酶反应中重新激活的分子机制。以前，我们已经证明了PDC的去磷酸化是由两种密切相关的磷酸酶(PDP1和PDP2)催化的，它们在组织分布、酶活性和调节方面都有明显的不同。我们还发现，PDP2蛋白在糖尿病中下调，从而导致PDC不必要的失活，从而阻止碳水化合物燃料的有氧氧化。最近，我们首次获得证据表明，在线粒体中，PDP1和PDP2与不同的附属或靶向亚基相互作用。基于这些初步数据，我们假设体内磷酸酶活性的调节主要是通过特定的蛋白质-蛋白质相互作用发生的。在目前的应用中，这一主要的工作假说将通过以下具体目标进行研究：1)分离和鉴定PDP1和PDP2的相互作用伙伴；2)建立PDP1的靶向和调控的分子机制；3)建立PDP2的靶向和调控的分子机制；以及4)阐明PDP2在糖尿病中长期调控的分子机制。丙酮酸脱氢酶复合体的过度磷酸化使其失活，在代谢性酸中毒、缺血和糖尿病中具有不利影响。揭示丙酮酸脱氢酶磷酸酶1和2重新激活丙酮酸脱氢酶复合体的分子机制对开发新的治疗药物至关重要。

DESCRIPTION (provided by applicant): The long-term goal of this project is to understand the molecular mechanisms responsible for the regulation of mitochondrial pyruvate dehydrogenase complex (PDC). The reaction catalyzed by PDC plays a central role in general metabolism providing a source of carbon for the major energy-generating and biosynthetic pathways such as the Citric Acid Cycle, lipogenesis, and the biosynthesis of cholesterol. The activity of mammalian PDC is regulated through a reversible phosphorylation/dephosphorylation cycle catalyzed by pyruvate dehydrogenase kinase and pyruvate dehydrogenase phosphatase. Growing evidence strongly suggests that control over the phosphorylation state of PDC becomes critically important in starvation, metabolic acidosis, ischemia, and diabetes. The major thrust of this application is to understand the molecular mechanisms responsible for the re-activation of PDC in phosphatase reaction. Previously, we have demonstrated that dephosphorylation of PDC is catalyzed by two closely related phosphatases (PDP1 and PDP2) that are markedly different with respect to their tissue distribution, enzymatic activities and regulation. We have also found that PDP2 protein is down-regulated in diabetes thereby contributing to the unwanted inactivation of PDC, which prevents the aerobic oxidation of carbohydrate fuels. Recently, we obtained the first evidence indicating that, in mitochondria, PDP1 and PDP2 interact with different accessory or targeting subunits. Based on these preliminary data, we hypothesize that the regulation of phosphatase activity in vivo occurs largely through specific protein-protein interactions. In the present application, this major working hypothesis will be investigated through the following specific aims: 1) to isolate and identify the interacting partners of PDP1 and PDP2; 2) to establish the molecular mechanisms of targeting and regulation of PDP1; 3) to establish the molecular mechanisms of targeting and regulation of PDP2; and 4) to elucidate the molecular mechanism of long-term regulation of PDP2 in diabetes. Inactivation of pyruvate dehydrogenase complex due to its hyperphosphorylation has detrimental effect in metabolic acidosis, ischemia, and diabetes. Uncovering the molecular mechanisms responsible for the reactivation of pyruvate dehydrogenase complex carried out by pyruvate dehydrogenase phosphatases 1 and 2 is critical for development of new therapeuticals.