Mitochondrial Protein Acetylation and Energy Metabolism in Muscle

 描述(申请人提供)：这个项目研究了蛋白质乙酰化在营养诱导的线粒体功能障碍中的作用。赖氨酸乙酰化(ACK)是一种重要的可逆翻译后修饰。线粒体的生化环境特别有利于乙酰化，最近的研究发现在大量线粒体蛋白上有多个ACK位点，包括几种代谢酶和参与氧化磷酸化的蛋白质。在衰老和肥胖的背景下，线粒体ack水平的增加与代谢功能障碍相关。越来越多的证据表明，线粒体蛋白质上特定赖氨酸残基的乙酰化会损害代谢调节，导致糖耐量异常，并损害骨骼肌在运动中的表现。线粒体内负责脱乙酰基的蛋白质sirtuin3(SIRT3)已经被确定。然而，促进线粒体ACK的分子事件仍然知之甚少。乙酰辅酶A(CoA)是葡萄糖和脂肪分解代谢的中间产物，是可逆乙酰化所需的底物。体外研究 提示由于线粒体乙酰辅酶A的产生和/或积累增加，线粒体ACK可以非酶的方式发生。此外，最近对线粒体基质酶肉碱乙酰转移酶(CrAT)的研究强调了乙酰辅酶A水平升高、线粒体ACK和代谢调节之间的重要联系。CRAT将乙酰辅酶A转化为其膜渗透性肉碱结合物乙酰肉碱。CrAT反应允许线粒体外流过量的乙酰基，从而有可能影响由质量作用驱动的ACK事件。这一预测得到了来自实验室的初步研究的支持，这些研究表明，小鼠缺乏CrAT会加剧线粒体ACK，以应对营养过剩。此外，骨骼肌中的CrAT消融会导致代谢功能障碍和运动不耐受。这项建议的目的是确定线粒体蛋白质的赖氨酸乙酰化是否与CrAT缺乏的代谢后果有关。拟议的研究将集中在CrAT缺陷细胞和小鼠的分子和生理特征上。首先，我们将检验一种假设，即减少线粒体乙酰化的刺激可以恢复CrAT缺乏的心肌细胞的代谢功能。其次，我们将使用肌肉特异性缺失CrAT的小鼠来确定线粒体ack状态的正常化是否可以在缺失肌肉CrAT的情况下恢复葡萄糖和/或运动耐量。这些研究的结果将扩大我们对线粒体ACK功能相关性的理解，并可能导致确定治疗与年龄和肥胖相关的骨骼肌代谢障碍和运动耐量不足的治疗靶点。

 DESCRIPTION (provided by applicant): This project examines the role of protein acetylation in mediating nutrient-induced mitochondrial dysfunction. Lysine acetylation (AcK) has emerged as an important reversible post-translational modification. The biochemical environment of the mitochondrial compartment is particularly favorable for acetylation and recent studies have identified multiple AcK sites on a large number of mitochondrial proteins, including several metabolic enzymes and proteins involved in oxidative phosphorylation. Increased mitochondrial AcK levels correlate with metabolic dysfunction in the context of aging and obesity. A growing body of evidence suggests that acetylation of specific lysine residues on mitochondrial proteins impairs metabolic regulation leading to glucose intolerance, and compromises skeletal muscle performance during exercise. The protein responsible for deacetylation within the mitochondria, sirtuin 3 (SIRT3), has been identified. However the molecular events that promote mitochondrial AcK remain poorly understood. Acetyl-coenzyme A (CoA), a metabolic intermediate of glucose and fat catabolism, is the proposed substrate required for reversible acetylation. In vitro studies suggest mitochondrial AcK can occur non-enzymatically as a result of elevated mitochondrial acetyl-CoA production and/or accumulation. In addition, recent studies of the mitochondrial matrix enzyme, carnitine acetyltransferase (CrAT) underscore an important link between elevated acetyl-CoA levels, mitochondrial AcK and metabolic regulation. CrAT converts acetyl-CoA to its membrane permeate carnitine conjugate, acetylcarnitine. The CrAT reaction permits the mitochondrial efflux of excess acetyl groups and thereby has the potential to influence AcK events that are driven by mass action. This prediction is supported by preliminary studies from the laboratory showing that CrAT deficiency in mice exacerbates mitochondrial AcK in response to nutrient excess. Furthermore, CrAT ablation in the skeletal muscle results in metabolic dysfunction and exercise intolerance. The objective of this proposal is to determine if lysine acetylation of mitochondrial proteins contributes to the metabolic consequences of CrAT deficiency. The proposed studies will focus upon the molecular and physiological characterization of CrAT deficient cells and mice. First, we will test the hypothesis that stimuli that decrease mitochondrial acetylation can restore metabolic function in CrAT deficient myocytes. Second, we will use mice with a muscle-specific deletion of CrAT to determine whether normalization of mitochondrial AcK status restores glucose and/or exercise tolerance in the absence muscle CrAT. The results from these studies will expand our understanding of the functional relevance of mitochondrial AcK and could lead to the identification of therapeutic targets for treating age- and obesity- related skeletal muscle metabolic dysfunction and exercise intolerance.