Mcl-1 as an essential regulator of cardiac mitochondrial function

描述(由申请人提供)： 线粒体在心肌细胞的生死过程中起着至关重要的作用。它们是重要的能量生成器，通过氧化磷酸化提供ATP。然而，线粒体也监测来自环境和细胞内环境的复杂信息，包括生长因子、氧、活性氧和DNA损伤的存在或不存在。因此，线粒体功能障碍与心血管疾病之间存在密切联系也就不足为奇了。Bcl2家族蛋白调控线粒体外膜通透性，在调控线粒体凋亡途径中发挥关键作用。MCL-1是一种抗凋亡蛋白，其在心肌中的表达水平高于其他抗凋亡蛋白，如Bcl2和Bclxl。令人惊讶的是，关于Mcl-1如何调节心肌细胞的存活，人们知之甚少。因此，我们产生了心脏特异的可诱导的Mcl-1基因敲除，并发现Mcl-1在心肌细胞中的缺失导致迅速的线粒体功能障碍和细胞死亡。令人惊讶的是，Mcl-1缺陷的心肌细胞显示出坏死性细胞死亡的迹象，而不是凋亡性细胞死亡的迹象，这表明除了抗凋亡作用外，Mcl-1在维持心肌细胞线粒体功能方面具有重要但尚未确定的作用。为了更好地了解Mcl-1在心脏中的生理功能(S)，我们计划在目标1中寻找与Mcl-1相互作用的新蛋白，并阐明这种相互作用的功能意义。线粒体是高度动态的细胞器，不断经历分裂和融合，这些过程在线粒体的正常周转中发挥重要作用。这些过程中的缺陷会影响线粒体功能和细胞生存。我们发现Mcl-1可以通过诱导分裂来影响线粒体的动力学。因此，在目标2中，我们将检查Mcl-1是否通过招募线粒体分裂机制的组件来调节线粒体动力学，以及这一过程是否对正常的生物能量功能是必要的。通过自噬去除功能障碍的线粒体是一个基本的过程，心脏中这一途径的缺陷会导致功能障碍的线粒体积聚和心力衰竭的发生。超微结构分析显示，在缺乏Mcl-1的心肌细胞中缺乏线粒体自噬，这表明Mcl-1缺陷的心肌细胞不会将受损的线粒体运送到自噬小体。在目标3中，我们将研究Mcl-1在调控线粒体自噬中的作用。线粒体的周转率随着年龄的增长而下降，导致功能障碍的线粒体在细胞内积累。因此，在目标4中，我们将使用野生型和Mcl-1转基因小鼠来研究增强的Mcl-1水平是否可以保护心肌细胞免受心肌病的攻击，并延长衰老的心肌细胞的存活时间。该项目将为心肌细胞线粒体功能以及线粒体功能障碍如何促进心血管疾病的发展提供重要的新见解。(摘要结束)

DESCRIPTION (provided by applicant): Mitochondria play critical roles in both the life and death of cardiac myocytes. They are important generators of energy, providing ATP through oxidative phosphorylation. However, mitochondria also monitor complex information from the environment and intracellular milieu, including the presence or absence of growth factors, oxygen, reactive oxygen species, and DNA damage. Thus, it is not surprising that there is a strong link between mitochondrial dysfunction and cardiovascular disease. The Bcl-2 family proteins control mitochondrial outer membrane permeabilization and play a key role in regulating the mitochondrial apoptotic pathway. Mcl-1 is an anti-apoptotic Bcl-2 protein which is expressed at higher levels in the myocardium compared to other anti-apoptotic proteins such as Bcl-2 and Bcl-XL. Surprisingly, little is known about how Mcl-1 regulates cell survival in myocardial cells. We therefore generated a heart specific inducible knockout of Mcl-1 and discovered that loss of Mcl-1 in cardiac myocytes led to rapid mitochondrial dysfunction and cell death. Surprisingly, Mcl-1 deficient myocytes displayed signs of necrotic cell death instead of apoptotic cell death, suggesting that besides its anti-apoptotic role, Mcl-1 has an essential but yet unidentified role in maintaining mitochondrial function in cardiac myocytes. To better understand the physiological function(s) of Mcl-1 in the heart, we plan to identify new proteins that interact with Mcl-1 and elucidate the functional significance of this interaction in Aim 1. Mitochondria are highly dynamic organelles that are constantly undergoing fission and fusion, and these processes play important roles in the normal turnover of mitochondria. Defects in these processes can affect mitochondrial function and cell survival. We found that Mcl-1 can influence mitochondrial dynamics by inducing fission. Thus, in Aim 2, we will examine if Mcl-1 regulates mitochondrial dynamics by recruiting components of the mitochondrial fission machinery and whether this process is essential for normal bioenergetic function. Removal of dysfunctional mitochondria by autophagy is an essential process, and defects in this pathway in the heart lead to accumulation of dysfunctional mitochondria and development of heart failure. Ultrastructural analysis revealed a lack of mitochondrial autophagy in myocytes lacking Mcl-1, suggesting that Mcl-1 deficient myocytes are not delivering damaged mitochondria to autophagosomes. In Aim 3, we will investigate the role of Mcl-1 in regulating mitochondrial autophagy. Mitochondrial turnover decreases with age resulting in accumulation of dysfunctional mitochondria in the cell. Therefore, in Aim 4, we will investigate if enhanced levels of Mcl-1 will protect against cardiomyopathic challenge and as well as prolong survival of cardiac myocytes in aging using wild type and Mcl-1 transgenic mice. This project will provide important new insights into mitochondrial function in cardiac myocytes and how mitochondrial dysfunction contributes to development of cardiovascular disease. (End of Abstract)