Role of mitochondria in neurodegenerative diseases

我们已经探索了线粒体在帕金森病（PD）中的作用。目前已知家族性PD中至少有两种突变的基因产物PINK 1和Parkin介导缺陷线粒体的自噬去除，这表明PD的一个原因是线粒体质量控制受损。PINK 1是位于线粒体上的激酶，而Parkin是通常位于细胞溶质中的E3泛素连接酶。 在线粒体损伤后，Pink 1将胞质Parkin招募到线粒体中以介导线粒体自噬，揭示了哺乳动物细胞中Pink 1在Parkin上游工作的细胞生物学途径。PINK 1作为线粒体功能的传感器，帕金诱导了这些受损线粒体的消除。一个积累线粒体DNA突变的动物模型证实了帕金介导质量控制并从受损的线粒体中拯救神经元的模型。我们进行了全基因组RNAi筛选，以鉴定参与PINK 1将Parkin募集到线粒体和Parkin刺激线粒体自噬体吞噬的基因产物。 第一章 遗传筛选使我们在Parkin-/-和PINK 1-/-小鼠与Mutator小鼠杂交时鉴定出强烈的炎症表型，这些小鼠随着年龄的增长而积累线粒体DNA突变。 这种炎症通过同时失去STING（I型干扰素对胞质DNA的反应的中心调节物）而完全被挽救。 来自SNc的DA神经元的损失和在年老的Parkin-/-; Mutator小鼠中观察到的运动缺陷也通过STING的损失而被挽救，这表明炎症促进了这种表型。在与一个拥有大量PINK 1和Parkin突变型PD患者的领先德国小组合作时，我们发现双等位基因Parkin突变的人也显示出高于健康对照或特发性PD患者的循环细胞因子水平升高。此外，这些双等位基因Parkin和PINK 1患者的线粒体DNA循环水平高于健康对照或特发性PD患者。这些结果支持PINK 1和Parkin介导的线粒体自噬抑制先天免疫作为预防PD的一种方式的模型。 （二） 与小鼠和人相反，黑腹果蝇中Parkin的隐性突变显示出由过量线粒体损伤引起的严重的身体和运动缺陷，导致肌肉缺陷和多巴胺能神经元的进行性变性。包括STING和NF-kB途径在内的先天免疫途径在果蝇中是保守的，然而，先天免疫信号在Pink 1/Parkin突变果蝇病理学中的作用尚未确定。我们发现保守的免疫调节因子STING介导Parkin功能丧失表型。产生了两种遗传上独立的STING和Parkin双敲除菌株。STING的缺失显著降低了主要帕金空蝇表型的突变率，包括飞行肌肉缺陷、翅膀姿势和攀爬能力。令人惊讶的是，帕金突变果蝇中潜在的线粒体形态缺陷在这些双突变系中也被抑制。 第三章 我们探讨了STING如何在线粒体损伤和mtDNA释放到胞质溶胶中的下游发挥作用，激活先天免疫。我们发现了STING在激活LC 3的脂化中的一种新的和意想不到的功能，这被经典地认为与双膜自噬体有关。我们发现STING将LC 3脂化到单膜高尔基体相关膜上，而不是像主导模型那样诱导自噬。我们和其他人正在探索STING活性中这一新步骤的先天免疫功能。

We have explored the role of mitochondria in Parkinson's disease (PD). At least two gene products mutated in familial PD, PINK1 and Parkin, are now known to mediate autophagic removal of defective mitochondria suggesting that one cause of PD is an impairment of mitochondrial quality control. PINK1 is a kinase located on mitochondria whereas Parkin is an E3 ubiquitin ligase normally located in the cytosol. Upon mitochondrial damage Pink1 recruits cytosolic Parkin to mitochondria to mediate mitophagy revealing a cell biology pathway in mammalian cells where Pink1 works upstream of Parkin. PINK1 acts as a sensor of mitochondria function and Parkin induced the elimination of those damaged mitochondria. An animal model that accumulates mitochondrial DNA mutations corroborates the model that Parkin mediates quality control and rescues neurons from damaged mitochondria. We preformed full genome RNAi screens to identify gene products participating in PINK1 recruitment of Parkin to mitochondria and Parkin stimulation of autophagosome engulfment of mitochondria. 1) The genetic screens led us to identify a strong inflammatory phenotype in both Parkin-/- and PINK1-/- mice when crossed with Mutator mice, which accumulate mitochondrial DNA mutations with age. This inflammation is completely rescued by concurrent loss of STING, a central regulator of the type I Interferon response to cytosolic DNA. The loss of DA neurons from the SNc and the motor defect observed in aged Parkin-/-; Mutator mice are also rescued by loss of STING, suggesting that inflammation facilitates this phenotype. In collaboration with a leading German group with large cohorts of PINK1 and Parkin mutant PD patients we discovered that people with biallelic Parkin mutations also display elevated circulating cytokines at levels higher than either healthy controls or those with idiopathic PD. Furthermore, these biallelic Parkin and PINK1 patients had higher circulating levels of mitochondrial DNA than either healthy controls or idiopathic PD patients. These results support the model that PINK1- and Parkin-mediated mitophagy restrains innate immunity as a way to prevent PD. 2) In contrast to mice and man, recessive mutations in Parkin in Drosophila melanogaster display severe physical and locomotion defects caused by excess mitochondria damage, leading to muscle defects and progressive degeneration of dopaminergic neurons. Innate immunity pathways including STING and NF-kB pathways are conserved in flies, however, the role of innate immune signaling in Pink1/Parkin mutant fly pathology has not been established. We found the conserved immunity regulator STING mediates the Parkin loss of function phenotypes. Two genetically independent STING and Parkin double knockout strains were generated. Deletion of STING with significantly reduces the penetrance of the major Parkin null fly phenotypes including flight muscle defects, wing posture, and climbing ability. Surprisingly, the underlying mitochondria morphology defects in Parkin mutant flies were also suppressed in these double mutant lines. 3) We explored how STING, functioning downstream of mitochondrial damage and mtDNA release into the cytosol, activates innate immunity. We discovered a new and unanticipated function of STING in activating the lipidation of LC3, which is classically thought to be linked to double membrane autophagosomes. Instead of inducing autophagy as had been the dominant model, we found STING lipidates LC3 onto single membrane Golgi associated membranes. The innate immune function of this novel step in STING activity is being explored by us and others.