Regulation of myocardial growth and death by autophagy

描述(申请人提供)：自噬是长寿命蛋白质和细胞内细胞器降解的主要机制。自噬在能量缺乏时发挥适应性作用，如心肌缺血，从而介导细胞存活，而在某些病理条件下，自噬与细胞程序性死亡有关，如再灌注损伤。因此，有必要阐明自噬在各种病理生理条件下的功能，并确定自噬是如何在心脏中调节的。在永久性冠状动脉结扎(PCL)诱导的心肌梗死(MI)小鼠模型中，尽管在急性期过度激活自噬会增加死亡率，但在慢性期自噬下调会导致心功能障碍。哺乳动物不育蛋白20样激酶1(Mst1)是一种强有力的细胞凋亡和心力衰竭的刺激因子，它强烈地抑制自噬，而FoxO1被营养饥饿和心脏负荷所激活，刺激自噬。这个项目的总体目标是阐明应激下心脏自噬的生理和病理功能，以及自噬是如何由心脏中的应激反应信号机制调节的。我们假设：a)心肌梗死急性期Beclin 1诱导的自噬是有害的，而在MI慢性期由FoxO1诱导的自噬是适应的。B)Mst1通过与Beclin1直接的蛋白质-蛋白质相互作用，作为自噬的内源性抑制因子，从而通过泛素相互作用蛋白p62导致蛋白质聚集体的积累。C)Foxos要么去乙酰化，要么因饥饿和心脏负荷而上调，在介导适应性自噬中起着重要作用。这些假说将被验证，使用(1)建立的实验方法来评估体内外自噬小体的形成和自噬通量，(2)独特的转基因小鼠模型，包括心脏特异的和可诱导的Beclin1基因敲除小鼠，ATG7 KO和FoxO1 KO小鼠和系统性p62 KO小鼠，(3)PCL和主动脉剥离的小鼠模型，(4)shRNA介导的击倒和蛋白质组学。我们的研究将阐明自噬在调节应激状态下的生理和病理功能中的作用，以及调节心脏自噬的潜在信号机制。 公共卫生相关性：自噬是通过溶酶体降解蛋白质的一种重要机制，在各种病理生理条件下发挥着适应性作用。我们的研究将阐明自噬在调节应激状态下的生理和病理功能中的作用，以及调节心脏自噬的潜在信号机制。从这项研究中获得的知识可能有助于更好地了解心肌损伤和心力衰竭的机制，并开发治疗心肌梗死和心肌病患者的新策略。

DESCRIPTION (provided by applicant): Autophagy is a major mechanism of degradation for long-lived proteins and intracellular organelles. Autophagy plays an adaptive role under energy starvation, such as myocardial ischemia, thereby mediating cell survival, whereas autophagy associates with programmed cell death under some pathological conditions, such as reperfusion injury. Thus, it is essential to elucidate the function of autophagy in various pathophysiological conditions and to determine how autophagy is regulated in the heart. In a mouse model of myocardial infarction (MI), induced by permanent coronary ligation (PCL), although excessive activation of autophagy increases the mortality at an acute phase, downregulation of autophagy leads to cardiac dysfunction at a chronic phase. Mammalian sterile 20 like kinase 1 (Mst1), a potent stimulator of apoptosis and heart failure, strongly inhibits autophagy whereas FoxO1, which is activated by nutrient starvation and cardiac unloading, stimulates autophagy. The overall goal of this project is to elucidate both physiological and pathological functions of autophagy in the heart under stress and how autophagy is regulated by stress responsive signaling mechanisms in the heart. We hypothesize that: A) Strong induction of autophagy by Beclin 1 at an acute phase of MI is detrimental, whereas autophagy induced by FoxO1 at a chronic phase of MI is adaptive. B) Mst1 acts as an endogenous inhibitor of autophagy through direct protein-protein interaction with Beclin1, thereby causing an accumulation of protein aggregates through p62, an ubiquitin interacting protein. C) FoxOs are either deacetylated or upregulated by starvation and cardiac unloading and plays an essential role in mediating adaptive autophagy. These hypotheses will be tested, using (1) established experimental methods to evaluate autophagosome formation and autophagic flux in vitro and in vivo, (2) unique genetically altered mouse models, including cardiac specific and inducible Beclin1 knock down, atg7 KO, and FoxO1 KO mice and systemic p62 KO mice, (3) the mouse models of PCL and aortic debanding, (4) shRNA-mediated knock- down and proteomics. Our study will elucidate the role of autophagy in mediating both physiological and pathological functions under stresses and underlying signaling mechanisms regulating autophagy in the heart. PUBLIC HEALTH RELEVANCE: Autophagy, an important mechanism of protein degradation through lysosomes, plays an adaptive role in various pathophysiological conditions. Our study will elucidate the role of autophagy in mediating both physiological and pathological functions under stresses and underlying signaling mechanisms regulating autophagy in the heart. The knowledge obtained from this study may lead to better understanding of the mechanism of myocardial injury and heart failure and the development of novel strategies to treat patients with myocardial infarction and cardiomyopathy.