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

线粒体前序加工在线粒体蛋白的生物发生和蛋白质稳态中起着核心作用。必需的线粒体加工肽酶MPP在绝大多数进入的前体蛋白从细胞质输入时切割其前体肽。被切割的序列随后被Cym1/PreP寡肽酶降解。MPP活性受损触发线粒体应激反应，然而，在这种应激反应中平衡线粒体功能障碍的信号传导和实现机制在很大程度上是未知的。MPP突变和PreP活性下降与癫痫和阿尔茨海默病有关，其病理机制和病理生理后果尚不清楚。这项emmy - noether研究计划的目的是对线粒体蛋白酶MPP和Cym1/PreP进行综合分析。为此，我将(I)分析MPP损伤引起的线粒体应激反应，（ii）剖析MPP突变导致人类癫痫的病理生理后果，（iii）研究Cym1/PreP对淀粉样蛋白- β肽的转化以及线粒体淀粉样蛋白- β在阿尔茨海默病中的病理机制。我将结合体外、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）来破译线粒体前序处理机制在健康和疾病中的新作用。mpp相关的应激反应将使用全局蛋白质组学和转录组学分析进行研究，随后对相关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线粒体-细胞核信号传导和应激反应诱导的缓解也将对理解人类疾病中的MPP功能障碍具有很高的价值。特别地，我的目标是通过生化和细胞生物学的方法来机械地分析MPP亚基PMPCB中引起癫痫的突变。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通过体内模型进行仔细研究，该模型能够评估各种淀粉样蛋白- β物种的毒性，并研究它们被线粒体寡肽酶Cym1/PreP和Ste23/IDE降解的情况。由此产生的数据将被转移到人类组织培养细胞和小鼠模型上，以破译线粒体前序转换机制在阿尔茨海默病发病和进展中的作用。综上所述，amy - noether的提议旨在揭示线粒体应激信号和反应的机制原理，以及线粒体前序处理机制功能失调引起的神经系统疾病的发病机制。

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.