Reversible protein acetylation and sirtuin function

蛋白质-赖氨酸乙酰化是一种主要的翻译后修饰，其中乙酰化蛋白质组学研究 编目了数千个乙酰化位点，代表了不同细胞途径中的蛋白质。机械论 研究揭示了特定蛋白质乙酰化的不同后果，但这样的功能研究 已经落后于蛋白质编目。脱乙酰酶和乙酰辅酶A(AcCoA)依赖的转移酶是 涉及到控制靶蛋白的乙酰化状态，通过动力学和机制 导致细胞室特异性乙酰化的机制尚不清楚。而有证据表明酶催化的 乙酰化存在于细胞质、细胞核和内质网中，大多数线粒体蛋白的乙酰化被认为是 不催化的。我们最近描述了细胞蛋白质对生长因子的快速乙酰化反应。 刺激，其中显著包括线粒体蛋白质的一个子集，表明酶催化 乙酰化。因此，在AcCoA通量/水平的推动下，非酶乙酰化的贡献和 酶催化的乙酰化反应仍然知之甚少。在战略上利用培养的细胞系和 小鼠模型，这项提案的主要部分是研究控制特定和 在急性细胞反应和慢性能量耗竭条件下的蛋白质组范围的乙酰化。 除了促进乙酰化的细胞机制外，对脱乙酰酶的调节还会推动 在各种细胞器中的功能后果。依赖NAD+的蛋白脱乙酰酶(SIRT1-7)是一种 在不同的亚细胞室中发现的酶的主要家族。线粒体SIRT3与核 SIRT6和SIRT7是本提案的主题。SIRT3脱乙酰化并提高催化效率 参与氧化新陈代谢的酶。SIRT6和SIRT7是染色质结合蛋白，可以 去除组蛋白上特定的赖氨酸乙酰化。SIRT6和SIRT7催化 核小体的脱乙酰化是完全未知的，然而这些脱乙酰化在遗传学和生物学上的重要性 蛋白质正在迅速发展。SIRT6功能丧失(去乙酰化)突变导致癌症和围产儿 杀伤力。我们最近关于SIRT6的数据为SIRT6的分子功能提供了前所未有的洞察力 并揭示了小分子激活的治疗潜力。我们关于SIRT7的初步数据表明 与核小体结合相关的新活性。 为了解决我们对sirtuin生物学和机制的理解中的这些主要差距 以特定间隔的方式驱动功能相关的蛋白质乙酰化，目标如下 目的1，确定SIRT6和SIRT7对核小体脱乙酰基的作用机制；目的2， 阐明急性刺激和慢性刺激下隔室特异性蛋白乙酰化的机制 以及目标3，揭示了蛋白质乙酰化在途径水平和位置特异性的功能作用。 能量代谢和线粒体蛋白平衡。

Protein-lysine acetylation is a major post-translational modification, where acetyl-proteomic studies have catalogued thousands of acetylation sites, representing proteins in diverse cellular pathways. Mechanistic studies have revealed diverse consequences of specific protein acetylation, but such functional studies have lagged behind protein cataloguing. Deacetylases and acetyl-CoA (AcCoA) dependent transferases are implicated in controlling the acetylation state of target proteins, though the dynamics and the mechanisms that lead to cellular compartment-specific acetylation is unclear. While evidence of enzyme-catalyzed acetylation exits in the cytoplasm, nucleus and ER, most mitochondrial protein acetylation is thought to be uncatalyzed. We recently described rapid cellular protein acetylation in response to growth factor stimulation, which remarkably includes a subset of mitochondrial proteins, indicative of enzyme-catalyzed acetylation. Thus, the contribution of non-enzymatic acetylation, driven by AcCoA flux/levels, and of enzyme-catalyzed acetylation remains poorly understood. With strategic use of cultured cell lines and mouse models, a major portion of this proposal investigates the mechanisms that control specific and proteome-wide acetylation in both acute cellular responses and chronic energy-depleted conditions. In addition to cellular mechanisms that promote acetylation, regulation of deacetylases drive functional consequences in various organelles. The NAD+-dependent protein deacetylases (SIRT1-7) are a major family of enzymes found in diverse sub-cellular compartments. Mitochondrial SIRT3 and nuclear SIRT6 and SIRT7 are the subject of this proposal. SIRT3 deacetylates and increases the catalytic efficiency of enzymes involved in oxidative metabolism. SIRT6 and SIRT7 are chromatin bound proteins that can remove specific lysine acetylations on histones. The mechanisms by which SIRT6 and SIRT7 catalyze the deacetylation of nucleosomes are completely unknown, yet the genetic and biological importance of these proteins is advancing rapidly. Loss-of-function (deacetylation) mutants of SIRT6 cause cancer and perinatal lethality. Our recent data on SIRT6 provides unprecedented insight into the molecular functions of SIRT6 and reveal the therapeutic potential of small-molecule activation. Our preliminary data on SIRT7 indicates novel activities related to nucleosome binding. To address these major gaps in our understanding of sirtuin biology and of the mechanisms that drive functionally relevant protein acetylation in a compartment-specific manner, the following aims are proposed: Aim 1, Determine the mechanisms of nucleosomal deacetylation by SIRT6 and SIRT7; Aim 2, Elucidate the mechanisms of compartment-specific protein acetylation under acute stimulation and chronic metabolic stress; and Aim 3, Reveal pathway-level and site-specific functional roles of protein acetylation in energy metabolism and mitochondrial proteostasis.