Molecular basis of ESCRT-mediated sealing of the nuclear envelope.

ESRT 介导的核膜封闭的分子基础。

• 批准号: 395534068
• 负责人: Dr. Alexander von Appen
• 金额: --
• 依托单位: Max-Planck-Institut für molekulare Zellbiologie und Genetik (MPI-CBG)
• 依托单位国家: 德国
• 项目类别: Research Fellowships
• 财政年份: 2018
• 资助国家: 德国
• 起止时间: 2017-12-31 至 2020-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/395534068?language=en
• 关键词: Molecular; basis; ESCRT; mediated; sealing

The eukaryotic genome is compartmentalized in the nucleus; whose boundary is a double-membrane structure called the nuclear envelope (NE). The nuclear envelope segregates transcription from translation, shields the genome from parasitic nucleic acids and other environmental hazards, regulates gene activity, and prevents DNA damage and malignant transformation as a consequence. Dividing cells of vertebrates go through the process of “open mitosis” in which the NE opens up to such a degree that nucleoplasmic identity is lost. NE membranes including many of their proteins disperse into the connected endoplasmic reticulum (ER) during chromosomes segregation. The process is reversed during mitotic exit when an intact and closed NE reforms. After each cell cycle—and following a variety of mechanical stressors that damage the nuclear envelope—cells need to seal the nuclear envelope to maintain intact compartmentalization. How human cells open and seal the double membrane of the nuclear envelope in the course of each cell division is a long-standing question that still remains poorly understood. Recent studies demonstrated that endosomal sorting complexes required for transport III (ESCRT-III) proteins are involved in the cell cycle-dependent reformation and repair of the nuclear envelope. ESCRT-III proteins and their membrane remodeling functions are conserved in archaea and all eukaryotes. They further drive the formation of multi-vesicular bodies, and egress of enveloped viruses, as well as cytokinetic abscission. The ESCRT-III protein CHMP7 has recently been implicated to play a major role in the process of nuclear envelope reformation. We have recently reported that the inner nuclear membrane protein Lem2 recruits Chmp7 as the initiating step in NE gap closure before other ESCRT-III components, like IST1/CHMP8 and CHMP2A are recruited. Despite their fundamental importance during the cell cycle, we understand very little about how ESCRTIII proteins function during stages.My postdoctoral studies aim to structurally characterize i) how the ESCRT-III machinery is recruited to the nuclear envelope; ii) how ESCRT-III proteins assemble to perform membrane remodeling reactions at the NE; and iii) how this process is regulated during the mammalian cell cycle. In course of this work I will use biochemical reconstitutions, complex characterization by cross-linking mass spectrometry, structure determination by cryo-electron microscopy and genome wide genetic screens in human cells to understand the role of LEM2, CHMP7 and the ESCRT-III machinery at the site of the nuclear envelope.

真核生物的基因组被划分在细胞核中;其边界是称为核膜（NE）的双膜结构。核膜将转录与翻译分离，保护基因组免受寄生核酸和其他环境危害，调节基因活性，并因此防止DNA损伤和恶性转化。脊椎动物的分裂细胞经历“开放有丝分裂”的过程，其中NE开放到核质同一性丧失的程度。在染色体分离过程中，NE膜（包括其许多蛋白质）分散到相连的内质网（ER）中。在有丝分裂退出过程中，当一个完整的和封闭的NE改革逆转。在每个细胞周期后，在各种机械应激损伤核膜后，细胞需要封闭核膜以保持完整的区室化。人类细胞如何在每次细胞分裂过程中打开和密封核膜的双层膜是一个长期存在的问题，仍然知之甚少。最近的研究表明，内体分选复合物所需的运输III（ESCRT-III）蛋白参与细胞周期依赖的改造和修复的核膜。ESCRT-III蛋白及其膜重塑功能在古细菌和所有真核生物中是保守的。它们进一步驱动多囊泡体的形成、包膜病毒的排出以及细胞动力学裂解。ESCRT-III蛋白CHMP 7最近被认为在核膜重组过程中发挥重要作用。我们最近报道，内核膜蛋白Lem 2招募Chmp 7作为NE缺口闭合的起始步骤，然后招募其他ESCRT-III组分，如IST 1/CHMP 8和CHMP 2A。尽管它们在细胞周期中具有根本的重要性，但我们对ESCRTIII蛋白在各个阶段的功能知之甚少。我的博士后研究旨在从结构上表征i）ESCRT-III机器如何被招募到核膜; ii）ESCRT-III蛋白如何组装以在NE进行膜重塑反应; iii）这一过程在哺乳动物细胞周期中如何被调节。在这项工作的过程中，我将使用生化重组，复杂的表征交联质谱，结构测定冷冻电子显微镜和全基因组遗传筛选在人类细胞中了解LEM 2的作用，CHMP 7和ESCRT-III机器在核膜的网站。