Role of the Endosomal-Lysosomal Pathway in Mitochondrial Quality Control

项目摘要 心肌细胞线粒体功能障碍在心力衰竭发病机制中起早期作用。在内心深处， 线粒体的主要功能是满足跳动的肌细胞的高能量需求，通过提供 通过氧化磷酸化的ATP。然而，线粒体可以迅速转变为促进死亡 细胞器。作为对细胞内环境变化的反应，它们可以成为过量的 活性氧种和释放致死蛋白。毫不奇怪，细胞已经形成了防御 对抗可能对其造成伤害的异常线粒体的机制。细胞自我修复的能力和 防止不必要的死亡在有丝分裂后细胞中尤其重要，例如不能 很容易被替换。研究发现，功能失调的线粒体可以被自噬小体隔离 然后递送到溶酶体进行降解。然而，它的作用机制和调控 线粒体去除的特征不是很好，线粒体的额外机制是否 目前尚不清楚是否存在许可。我们之前已经发现E3泛素连接酶Parkin扮演着一个 应激和帕金氏导联缺失在清除心脏功能障碍线粒体中的重要作用 到心肌梗死后功能障碍的线粒体堆积。众所周知，帕金可以诱导 线粒体自噬，但我们的初步研究发现，有证据表明，帕金也可以促进 通过不依赖于自噬的机制清除线粒体。在这项建议中，我们将探讨 假设小GTP酶Rab5和内体降解途径在 清除心肌细胞中功能失调的线粒体。这一假设将以两个目的进行检验。目标1将 确定Rab5的功能重要性和内体介导的心肌细胞线粒体清除 体外和体内。我们还将研究内体途径和 心脏中的传统/替代自噬途径。在目标2中，我们将描述Beclin1在 启动内体途径以应对线粒体损伤。我们将检查Beclin1是否 通过形成一种特异性的形式来调节内体降解途径的激活以响应细胞应激 与Rab5和Vps34的前内体复合体。用Unique进行体内外功能丧失的研究 将利用心脏特异性诱导型Beclin1缺陷小鼠来确认Beclin1的功能重要性 启动早期内小体的形成，以响应线粒体损伤和应激。这些研究将 为功能障碍和潜在危险的线粒体是如何清除的提供了重要的新见解 心脏。这些研究还将为这一途径中新的潜在治疗靶点提供洞察力。

Project Summary Mitochondrial dysfunction in cardiac myocytes occurs early in the pathogenesis of heart failure. In the heart, the primary function of mitochondria is to meet the high energy demand of the beating myocytes by providing ATP through oxidative phosphorylation. However, mitochondria can quickly change into death–promoting organelles. In response to changes in the intracellular environment, they can become excessive producers of reactive oxygen species and release pro-death proteins. Not surprisingly, cells have developed defense mechanisms against aberrant mitochondria that can cause harm to it. The ability of a cell to repair itself and prevent unnecessary death is particularly important in a post-mitotic cell such as a myocyte that cannot be easily replaced. Studies have found that dysfunctional mitochondria can be sequestered by autophagosomes and subsequently delivered to lysosomes for degradation. However, the mechanism and regulation of mitochondrial removal are not well characterized and whether additional mechanisms of mitochondrial clearance exist is currently unclear. We have previously found that the E3 ubiquitin ligase Parkin plays an important role in clearing dysfunctional mitochondria in the heart in response to stress and lack of Parkin leads to accumulation of dysfunctional mitochondria after a myocardial infarction. Parkin is known to induce autophagy of mitochondria but our preliminary studies have uncovered evidence that Parkin can also promote clearance of mitochondria via an autophagy-independent mechanism. In this proposal, we will explore the hypothesis that the small GTPase Rab5 and the endosomal degradation pathway play an important role in clearing dysfunctional mitochondria in myocytes. This hypotheses will be tested with two aims. Aim 1 will define the functional importance of Rab5 and endosomal-mediated mitochondrial clearance in myocytes in vitro and in vivo. We will also examine the relationships between the endosomal pathway and traditional/alternative autophagy pathways in the heart. In Aim 2, we will delineate the role of Beclin1 in initiating the endosomal pathway in response to mitochondrial damage. We will examine whether Beclin1 regulates activation of the endosomal degradation pathway in response to cellular stress by forming a specific pro-endosomal complex with Rab5 and Vps34. Loss-of-function studies in vitro and in vivo using unique cardiac specific inducible Beclin1 deficient mice will be utilized to confirm the functional importance of Beclin1 initiating formation of early endosomes in response to mitochondrial damage and stress. These studies will provide important novel insight into how dysfunctional and potentially dangerous mitochondria are cleared in the heart. These studies will also provide insights into new potential therapeutic targets in this pathway.