UNDERSTANDING THE ROLE OF MITOCHONDRIAL QUALITY CONTROL

线粒体质量控制(MQC)是线粒体生物学领域中的一个新兴方面，它是指控制线粒体成分的受控降解的机制。事实上，最近已经发现了几种MQC机制，其中最著名的是有丝分裂，但它们在各种细胞类型中的作用、它们之间的相互关系以及它们在体内的生理相关性仍然知之甚少。这个项目旨在利用横纹肌和免疫细胞来回答这些问题，横纹肌和免疫细胞具有不同的设计和功能。我们将集中讨论两个MQC机制：i)有丝分裂吞噬(通过自噬选择性地清除整个线粒体)，以及ii)产生线粒体衍生的小泡(MDV：通过囊泡分泌过程选择性地清除线粒体成分)。*第一部分：肌肉细胞中的线粒体QC：维持适应的和功能最佳的线粒体池对于正常的收缩活动至关重要。出于这个原因，我们假设在心肌细胞中，MQC与线粒体的含量/活性是相称的，并且这些机制以一种分层的方式起作用。更具体地说，我们假设MDV的产生是基线线粒体管家和对线粒体应激的早期反应的主要机制，而当整个细胞器受到过度损伤时，有丝分裂变得更加主导。为了测试这一点，c2c12成肌细胞/肌管将被用来表征线粒体功能、有丝分裂、MDV产生和细胞表型，在基线和已知条件下迫使MQC反应。遗传方法将被用来专门调节有丝分裂和MDV的产生，以阐明它们的作用。这也将在WT和转基因小鼠中使用包括定量形态电子显微镜在内的各种方法在体内进行研究。*第2部分：巨噬细胞中的线粒体QC(MΦ)：在MΦ中，线粒体在促进(M1)和抗(M2)炎症反应的发展中发挥作用，但其潜在机制尚不清楚。我们假设，通过改变线粒体功能和线粒体抗原提呈免疫信号，线粒体吞噬和MDV生产的调节在这一过程中发挥重要作用。为了测试这一点，原始MΦ和原代骨髓来源的MΦ将在体外暴露于促炎或抗炎刺激。线粒体功能将与有丝分裂、MDV产生、线粒体抗原提呈和MΦ分化标记物一起分析。为了阐明它们在M-Φ分化/激活中的作用，将利用遗传方法和转基因小鼠来调节有丝分裂和MDV的产生。还将进行体内研究，以检验MQC在免疫反应中的作用。*总体而言，这项工作将提供有关MQC在线粒体和细胞功能中的作用的重要信息。

Mitochondrial quality control (mQC), which refers to the mechanisms governing the regulated degradation of mitochondrial components, is an emerging facet in the field of mitochondrial biology. Indeed, several mQC mechanisms, the best known being mitophagy, have been uncovered recently, but their role in various cell types, their level of inter-relation, and their physiological relevance in vivo, remain poorly understood. This program aims to answer these questions using striated muscle and immune cells which have different designs and functions. We will focus on two mQC mechanisms: i) mitophagy (the selective clearance of whole mitochondria through autophagy), and ii) production of mitochondrial-derived vesicles (MDVs: the selective clearance of mitochondrial components through a vesicle secretion process).***Part 1: Mitochondrial QC in muscle cells: Maintenance of an adapted and optimally functioning pool of mitochondria is critical for normal contractile activity. For this reason we hypothesize that in myocytes mQC is commensurate to mitochondrial content/activity, and that these mechanisms act in a hierarchical manner. More specifically, we postulate that MDV production is a predominant mechanism for baseline mitochondrial housekeeping, and early response to mitochondrial stress, while mitophagy becomes more predominant when excessive damage to whole organelles is present. To test this, c2c12 myoblasts/myotubes will be used to characterize mitochondrial functions, mitophagy, MDV production, and cellular phenotype at baseline and in response to conditions known to force a mQC response. Genetic approaches will be used to specifically modulate mitophagy and MDV production in order to elucidate their role. This will also be studied in vivo in WT and transgenic mice using various approaches including quantitative morphological electron microscopy.***Part 2: Mitochondrial QC in macrophages (MΦ): In MΦ, mitochondria play a role the development of pro (M1) and anti (M2)-inflammatory responses, but the underlying mechanisms remain unclear. We hypothesize that modulation of mitophagy and MDV production, plays an important role in this process by altering mitochondrial functions, and mitochondrial antigen presentation for immune signalling. To test this Raw MΦ and primary bone marrow-derived MΦ will be exposed to pro- or anti-inflammatory stimuli in vitro. Mitochondrial functions will be analyzed together with mitophagy, MDV production, mitochondrial antigen presentation, and MΦ differentiation markers. Genetic approaches and transgenic mice will be used to modulate mitophagy and MDV production in order to elucidate their role in MΦ differentiation/activation. In vivo studies will also be conducted to examine the role of mQC on the immune response.***Overall this work will provide important information on the role of mQC in mitochondrial and cellular functions.**************