Novel roles of the mitochondrial presequence processing machinery in health and disease

线粒体前序列加工在线粒体蛋白质生物合成和蛋白质稳态中起着重要作用。必需的线粒体加工肽酶MPP切割绝大多数进入的前体蛋白质的前序列肽，当它们从胞质溶胶输入时。切割的前序列随后被Cym 1/PreP寡肽酶降解。受损的MPP活性触发线粒体应激反应，然而，在这种应激反应期间平衡线粒体功能障碍的信号传导和实施机制在很大程度上是未知的。MPP的突变和PreP活性的下降与癫痫和阿尔茨海默病有关，其病理机制和病理生理学后果仍然是难以捉摸的。这个Emmy-Noether研究计划的目的是对线粒体蛋白酶MPP和Cym 1/PreP进行综合分析。为此，我将（i）分析MPP损伤后诱导的线粒体应激反应，（ii）剖析导致人类癫痫的MPP突变的病理生理学后果，和（iii）研究Cym 1/Cym 2对淀粉样β肽的转换，PreP与阿尔茨海默病中线粒体淀粉样β蛋白的病理机制我将结合联合收割机在体外，在细胞器和体内的方法，使用不同的模式生物（酵母，组织培养，小鼠和患者来源的细胞），以破译这些线粒体前序列加工机制在健康和疾病的新角色。 MPP相关的应激反应将使用全球蛋白质组学和转录组学分析进行研究，然后对所涉及的线粒体蛋白进行详细的分子和功能分析。MPP功能障碍的后果及其通过神经核信号传导和应激反应诱导的缓解对于理解人类疾病中的MPP功能障碍也将是非常有价值的。特别是我的目的是机械地分析癫痫引起突变的MPP亚基PMPCB的生化和细胞生物学方法。这种突变的病理生理后果将使用患者来源的诱导多能干细胞和长期神经上皮干细胞来破译。线粒体在阿尔茨海默病发病机制中的作用将使用体内模型进行详细检查，该模型能够评估各种淀粉样蛋白-β物质的毒性并研究线粒体寡肽酶Cym 1/PreP和Ste 23/IDE对其的降解。所得数据将转移到人类组织培养细胞和小鼠模型上，以破译线粒体前序列周转机制在阿尔茨海默病发病机制和进展中的作用。总之，这个Emmy-Noether建议旨在揭示线粒体应激信号传导和反应以及由功能失调的线粒体前序列处理机制引起的神经系统疾病的发病机制的机制原理。

Mitochondrial presequence processing plays a central role in mitochondrial protein biogenesis and proteostasis. The essential mitochondrial processing peptidase MPP cleaves the presequence peptides of the vast majority of incoming precursor proteins upon their import from the cytosol. The cleaved presequences are subsequently degraded by the Cym1/PreP oligopeptidase. Impaired MPP activity triggers a mitochondrial stress response, however, the signaling and implementation mechanisms to balance mitochondrial dysfunctions during this stress response are largely unknown. Mutations in MPP and a decline in PreP activity have been implicated in epilepsy and Alzheimer's disease, the pathological mechanisms and pathophysiological consequences remain yet elusive. The objective of this Emmy-Noether-research proposal is the comprehensive analysis of the mitochondrial proteases MPP and Cym1/PreP. To this end I will (i) analyze the mitochondrial stress response induced upon MPP impairment, (ii) dissect the pathophysiological consequences of a mutation in MPP resulting in epilepsy in humans and (iii) investigate turnover of Amyloid-beta peptides by Cym1/PreP and the pathological mechanism of mitochondrial Amyloid-beta in Alzheimer's disease. I will combine in vitro, in organello and in vivo approaches using different model organisms (yeast, tissue culture, mice and patient-derived cells) to decipher these novel roles of the mitochondrial presequence processing machinery in health and disease. The MPP-linked stress response will be investigated using global proteomic and transcriptomic analysis, followed by detailed molecular and functional analysis of the involved mitochondrial proteins. The consequences of MPP dysfunction and its mitigation by mitochondrial-nucleus signaling and stress response induction will also be highly valuable for the understanding of MPP dysfunction in human diseases. In particular I aim to mechanistically analyze an epilepsy-causing mutation in the MPP subunit PMPCB by biochemical and cell biological approaches. The pathophysiological consequences of this mutation will be deciphered using patient-derived induced pluripotent stem cells and long-term neuro-epithelial stem cells. The role of mitochondria in the pathogenesis of Alzheimer's disease will be scrutinized using an in vivo model that enables assessment of the toxicity of various Amyloid-beta species and investigation of their degradation by the mitochondrial oligopeptidases Cym1/PreP and Ste23/IDE. Resulting data will be transferred onto human tissue culture cells and mouse models to decipher the role of the mitochondrial presequence turnover machinery in the pathogenesis and progression of Alzheimer's disease. Taken together, this Emmy-Noether-proposal aims to uncover the mechanistic principles in mitochondrial stress signaling and response and in the pathogenesis of neurological disorders induced by a dysfunctional mitochondrial presequence processing machinery.