Secretion of mitochondria as a cellular quality control mechanism

项目总结 在心脏中，线粒体的主要功能是满足心脏跳动的高能量需求 通过氧化磷酸化提供三磷酸腺苷。然而，线粒体可以迅速转变为 促进死亡的细胞器。毫不奇怪，细胞已经发展出多种防御机制来对抗 会对细胞造成伤害的异常线粒体。消除功能障碍的线粒体和 对于不易替代的有丝分裂后的心肌细胞，防止不必要的死亡尤为重要。它是 已经证实，功能失调的线粒体会被自噬小体迅速隔离，随后 输送到溶酶体进行降解。最近的研究发现，细胞也可以消除线粒体 通过依赖Rab9的替代自噬途径或依赖Rab5的早期内体途径。 线粒体也可以被溶酶体直接摄取。显然，线粒体消除的多个途径 存在于细胞中，以防止它们积累并确保生存。然而，这些退化途径都是 在溶酶体水平上汇聚，目前尚不清楚替代线粒体质量控制 当溶酶体功能受损时，存在多种途径。我们发现功能失调的线粒体 当内部降解途径不堪重负或受损时，也可以从细胞分泌。我们的 初步数据显示，已经损害溶酶体降解的Rab7-/-和LAMP2-/-细胞仍然 能够有效地消除去极化的线粒体。我们还发现循环细胞外的水平增加 心肌梗死后血浆和心脏组织中含有线粒体的囊泡(EV) 基础水平的Rab7和LAMP2缺陷小鼠。EVS的蛋白质组学分析与蛋白质印迹分析相结合 提示线粒体存在于源自内体途径的囊泡中。在这项提案中，我们 将调查EVS中功能障碍的线粒体是从细胞中分泌出来的假设，以及这一假设 代表着心脏中重要的质量控制途径，当溶酶体被 不堪重负或妥协。这一假设将通过两个具体目标进行检验。具体目标1将确定 含线粒体的胞外小泡的起源和去向及其病理生理决定 诱导它们从心肌细胞中释放的条件。《特殊目标2》将剖析分子机制。 调节EVS中线粒体的分泌。总体而言，这些研究将为 心肌线粒体消除的新替代机制。这些研究还将提供 洞察这些EV是否可以潜在地作为心脏应激的早期诊断生物标志物 疾病的发展。

Project summary 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. Not surprisingly, cells have developed multiple defense mechanisms against aberrant mitochondria that can cause harm to the cell. The ability to eliminate dysfunctional mitochondria and prevent unnecessary death is particularly important in post-mitotic myocytes that cannot be easily replaced. It is well established that dysfunctional mitochondria are rapidly sequestered by autophagosomes and subsequently delivered to lysosomes for degradation. Recent studies have identified cells that can also eliminate mitochondria via a Rab9-dependent alternative autophagy pathway or via a Rab5-dependent-early endosomal pathway. Mitochondria can also be directly taken up by lysosomes. Clearly, multiple pathways of mitochondrial elimination exist in cells to prevent their accumulation and ensure survival. However, these degradation pathways all converge at the level of the lysosomes and it is not clear whether alternative mitochondrial quality control pathways exists when lysosomal function is compromised. We have discovered that dysfunctional mitochondria can also be secreted from cells when internal degradation pathways are overwhelmed or impaired. Our preliminary data demonstrate that Rab7-/- and LAMP2-/- cells that have impaired lysosomal degradation are still able to efficiently eliminate depolarized mitochondria. We also found increased levels of circulating extracellular vesicles (EVs) containing mitochondria in plasma after a myocardial infarction, as well as in cardiac specific Rab7- and LAMP2-deficient mice at baseline. Proteomics analysis of EVs combined with Western blot analysis suggest that the mitochondria are in vesicles that originate from the endosomal pathway. In this proposal, we will investigate the hypothesis that dysfunctional mitochondria are secreted from cells in EVs and that this represent an important quality control pathway in the heart that can compensate when lysosomes are overwhelmed or compromised. This hypothesis will be tested with two specific aims. Specific aim 1 will identify the origin and fate of the extracellular vesicles containing mitochondria and determine the pathophysiological conditions that induce their release from myocytes. Specific aim 2 will dissect the molecular mechanisms regulating secretion of mitochondria in EVs. Overall, these studies will provide important new insights into a novel alternative mechanism of mitochondrial elimination in the myocardium. The studies will also provide insights into whether these EVs can potentially function as early diagnostic biomarkers of cardiac stress prior to development of disease.