Regulation of myocardial growth and death by autophagy

描述（由申请人提供）：自噬是长寿命蛋白质和胞内细胞器降解的主要机制。自噬在能量饥饿（如心肌缺血）条件下发挥适应性作用，介导细胞存活，而在一些病理条件下，如再灌注损伤，自噬与细胞程序性死亡有关。因此，有必要阐明自噬在各种病理生理条件下的功能，并确定自噬在心脏中的调节方式。在永久性冠状动脉结扎（PCL）诱导的心肌梗死（MI）小鼠模型中，尽管自噬过度激活会增加急性期的死亡率，但自噬下调会导致慢性期的心功能障碍。哺乳动物不育20样激酶1 （Mst1）是一种有效的细胞凋亡和心力衰竭刺激物，可强烈抑制自噬，而FoxO1则可通过营养饥饿和心脏卸液激活，刺激自噬。本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，以及自噬如何受到心脏应激反应信号机制的调节。我们假设：A) Beclin 1在心肌梗死急性期强烈诱导自噬是有害的，而FoxO1在心肌梗死慢性期诱导自噬是适应性的。B) Mst1通过与Beclin1的直接蛋白-蛋白相互作用作为内源性自噬抑制剂，从而通过泛素相互作用蛋白p62引起蛋白聚集体的积累。C) FoxOs在饥饿和心脏卸载过程中去乙酰化或上调，在介导适应性自噬中起重要作用。这些假设将被验证，使用(1)已建立的实验方法来评估体外和体内的自噬体形成和自噬通量，(2)独特的基因改变小鼠模型，包括心脏特异性和诱导Beclin1敲低，at7ko， FoxO1 KO小鼠和全身p62 KO小鼠，(3)PCL和主动脉脱带小鼠模型，(4)shrna介导的敲低和蛋白质组学。我们的研究将阐明自噬在应激下介导心脏生理和病理功能的作用，以及调节自噬的潜在信号机制。

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.