营养物中枢感知器溶酶体v-ATPase的动态乙酰化和功能研究

营养物的动态变化能被机体感应并相应地调控代谢通路以维持代谢稳态。我们之前发现了溶酶体上的v-ATPase是感知葡萄糖调控AMPK的关键分子，加之它还能感知氨基酸等调节mTORC1，因此是营养物的感知中心。限制营养物质的摄入，如卡路里限制能引起长寿等益处，其机制涉及AMPK、mTORC1等，但其“长寿信号”如何传导到此尚不明确。我们的前期工作发现二甲双胍和白藜芦醇等“类卡路里限制”依赖v-ATPase的活性改变而激活AMPK，该过程不涉及能量状态或葡萄糖水平，而是直接改变v-ATPase的乙酰化水平，引发其调控的溶酶体途径的活化。我们进一步通过蛋白质质谱分析，发现v-ATPase上发生了一系列乙酰化修饰改变，并鉴定出了关键位点。本项目拟深入探索卡路里限制相关的“信号”的本质，以及该信号如何通过改变v-ATPase的乙酰化而实现其生理意义，有望为预防和治疗代谢性疾病以及延缓衰老提供线索和依据。

Changes in availability of nutrients are sensed by the cells and organisms, leading to reprogramming of metabolic programs to maintain metabolic homeostasis. We have delineated a sensing pathway for glucose, in which low glucose are sensed by the lysosomal v-ATPase, leading to the inhibition of such proton pump and the activation of AMPK, in the absence of energy change. We named it as the “lysosomal pathway”. Combining the previous findings that v-ATPase is also a hub for the regulation of mTORC1 in response to changes in amino acids, glucose and cholesterol, it is clear that v-ATPase plays a pivotal role in sensing the availability of nutrients. Restricted food intake, such as calorie restriction, is well established to have a positive impact on healthy and life span, and involves the regulation of AMPK and mTORC1. However, how v-ATPase senses the changes in food availability remains relatively ill-defined. In addition to our earlier work with metformin, our preliminary data have also shown that resveratrol, a “calorie restriction mimetic”, activates AMPK via causing the inhibition of v-ATPase, and the subsequent activation of the lysosomal pathway. We have also found that in sensing resveratrol, as well as calorie restriction, v-ATPase undergoes massive changes in acetylation. Through extensive mass spectrometry analyses, we have identified multiple candidates of acetylated v-ATPase subunits, and have characterized their acetylation sites that undergo dynamic acetylation and deacetylation in response to resveratrol treatment and calorie restriction. We propose to continue to thoroughly study the mechanisms of the subunits of v-ATPase that are acetylated/deacetylated in light of nutrient change. We will also use animal models to study the biological consequences of the acetylation/deacetylation of v-ATPase. Our work will help provide new targets for therapeutic intervention of metabolic diseases and of longevity.