Secondary structure of the nascent chain as regulatory element for translocation into the endoplasmic reticulum; mechanisms and pathophysiological relevance

新生链的二级结构作为易位到内质网的调节元件；

• 批准号: 408017424
• 负责人: Professor Dr. Jörg Tatzelt
• 金额: --
• 依托单位: Abteilung für Biochemie Neurodegenerativer Erkrankungen
• 依托单位国家: 德国
• 项目类别: Research Grants
• 财政年份: 2018
• 资助国家: 德国
• 起止时间: 2017-12-31 至 2021-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/408017424?language=en
• 关键词: Secondary; structure; nascent; chain; regulatory

Around 25-30% of all proteins synthesized in the cytosol of eukaryotic cells are integrated into the plasma membrane or are secreted. Membrane integration and import into the lumen of the endoplasmic reticulum (ER) is mainly mediated by a heterotrimeric membrane protein complex called Sec61 complex, which is homologous to the SecY complex in bacteria. A major fraction of secretory proteins contains N-terminal signal peptides that mediate transport of the nascent chain to the Sec61 translocon and initiate translocation into the ER lumen. Failure to deliver secretory or membrane proteins to the ER can have fatal consequences. First, these proteins do not reach their final destination (loss of their physiological function) and second, they may form cytosolic aggregates with toxic activities based on the presence of hydrophobic domains (gain of toxic function). For example, mislocalization of the cellular prion protein to the cytosol can induce neuronal cell death in transgenic mice and mammalian cell culture models. Thus, quality control pathways have evolved to eliminate mistargeted proteins. Our work revealed an intrinsic impairment of the Sec61/SecY complex of eukaryotes/bacteria to translocate secretory proteins that are entirely unstructured. The research proposal focusses on a detailed understanding of the mechanisms by which structural features of the polypeptide chain modulate the translocation process. In addition, pathophysiological consequences of a defective ER import will be addressed. Specifically, we will investigate pathways that enable the mammalian translocon to translocate client proteins with extended unstructured domains and study a deregulated translocation during stress and in cells propagating neurotoxic protein conformers. Finally, we will investigate the fate of non-translocated secretory proteins to evaluate the impact of an uncleaved signal peptide on degradation, subsequent trafficking and cytotoxic activity. The experimental approaches include in vitro assays,bacterial, yeast and mammalian cell culture models, including primary neurons, and mouse models of neurodegenerative diseases.

在真核细胞的胞质溶胶中合成的所有蛋白质的约25-30%整合到质膜中或被分泌。膜整合和输入到内质网（ER）的内腔中主要是由异源三聚体膜蛋白复合物（称为Sec 61复合物）介导的，该复合物与细菌中的SecY复合物同源。分泌蛋白的主要部分含有N-末端信号肽，其介导新生链向Sec 61易位子的转运并启动易位进入ER腔。未能将分泌蛋白或膜蛋白递送到ER可能具有致命的后果。首先，这些蛋白质不会到达其最终目的地（失去其生理功能），其次，它们可能会基于疏水性结构域的存在而形成具有毒性活性的胞浆聚集体（获得毒性功能）。例如，在转基因小鼠和哺乳动物细胞培养模型中，细胞朊病毒蛋白到胞质溶胶的错误定位可诱导神经元细胞死亡。因此，质量控制途径已经发展到消除错误的蛋白质。我们的工作揭示了真核生物/细菌的Sec 61/SecY复合物的内在损伤，以转运完全非结构化的分泌蛋白。该研究计划侧重于详细了解多肽链结构特征调节易位过程的机制。此外，一个有缺陷的ER进口的病理生理后果将得到解决。具体来说，我们将调查的途径，使哺乳动物translocon易位客户端蛋白与扩展的非结构化结构域和研究的失调易位过程中的压力和细胞繁殖神经毒性蛋白构象。最后，我们将研究非易位分泌蛋白的命运，以评估未切割的信号肽对降解，随后的贩运和细胞毒活性的影响。实验方法包括体外试验，细菌，酵母和哺乳动物细胞培养模型，包括原代神经元，和神经退行性疾病的小鼠模型。