Lipid flux during autophagosome membrane biogenesis

自噬体膜生物发生过程中的脂质通量

基本信息

批准号：
10331030
负责人：
Thomas James Melia
金额：
$ 33.5万
依托单位：
YALE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-02-19 至 2024-01-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10331030
关键词：
Antibodies Autophagocytosis Autophagosome Binding Biochemistry Biogenesis Carrier Proteins Catalysis Cells Cellular Stress Cellular biology Complex Consumption Coupled Cytoplasm Cytosol Development Disease Docking Electron Microscopy Elements Encapsulated Engineering Enzymes Event Fluorescence Microscopy Genes Growth Harvest Human In Vitro Individual Infection Knock-out Link Lipid Synthesis Pathway Lipids Liposomes Location Lysosomes Malignant Neoplasms Mammalian Cell Mediating Membrane Microscopy Mitochondria Modeling Nerve Degeneration Organelles Pathway interactions Phase Phosphotransferases Process Production Protein Family Proteins Proteome Resources Route Site Source Starvation Structure Synaptic Vesicles System Time Ubiquitin Vesicle Virus Work biological adaptation to stress experimental study light microscopy lipid metabolism lipid transfer protein lipid transport member membrane biogenesis membrane model novel particle pathogen protein aggregation protein function reconstitution recruit response tool trafficking virtual

项目摘要

Summary Macro-autophagy is the intracellular stress-response pathway by which the cell packages portions of the cytosol for delivery into the lysosome. This packaging is carried out by the de novo formation of a new organelle called the autophagosome that grows and encapsulates cytosolic material for eventual lysosomal degradation. How autophagosomes form, including especially from where the autophagosomes extract lipid in order to expand their membranes, has been a core problem in the field for over 50 years. Two competing models have emerged and suggest that either the autophagosome simply grows out of a pre-existing compartment (like the ER) or the autophagosome forms from the continued fusion of individual vesicles recruited from many different sites in the cell. Furthermore, lipid biogenesis pathways are intimately associated with autophagy, suggesting that one or more organelles involved in the production of lipids might also be tightly associated to the growth of the autophagosome. Temporal studies established over a decade ago that the autophagy protein ATG2, is needed during membrane expansion, but how ATG2 facilitates membrane growth has remained elusive. Our preliminary results now demonstrate that ATG2 is a member of a novel lipid-transport family of proteins and suggest a third model for membrane expansion; the bulk delivery of lipid from organelles through protein- mediated contact sites. Indeed, ATG2 binds up to 20 lipids at once, an order of magnitude more than virtually any other lipid transport protein, and thus has the capacity to move a lot of lipid during biogenesis. We show that in cells, ATG2 accumulates at an interface between autophagosomes and the ER, strongly suggesting this organelle-organelle contact site might be the location of lipid transfer. In addition, we have developed gene- edited knockouts of each of the ATG2 proteins in humans and discovered that in the absence of ATG2, not only do autophagosomes not expand, but hundreds of vesicles collect at the site of autophagosome biogenesis. This surprising observation suggests that vesicle-mediated delivery of membrane might also be essential and that the fusion of these vesicles is specifically disrupted when ATG2-mediated lipid transport is absent. With this proposal, we expect to describe how ATG2 works with proteins on both the autophagosome and the ER to drive lipid flow. Likely, this will involve proteins needed to stabilize organelle-organelle contact sites and may also involve proteins sensing or regulating lipid production in the ER. We will then establish how this lipid flow is related to the recruitment and utilization of trafficking vesicles to describe to support autophagosome growth.

概括宏 - 自助噬菌是细胞内应力 - 响应途径，细胞包装部分细胞质的部分用于输送到溶酶体。这种包装是由从头形成的新细胞器进行的生长并封装胞质材料的自噬体，以实现最终的溶酶体降解。如何自噬体形式，尤其是从自噬体提取脂质以扩展的地方他们的膜是该领域的核心问题已有50多年了。出现了两个竞争模型并建议自噬体只是从先前存在的隔室（例如ER）或自噬体形式来自从许多不同地点募集的单个囊泡的持续融合。细胞。此外，脂质生物发生途径与自噬密切相关，这表明一个或涉及脂质生产的更多细胞器也可能与生长紧密相关自噬体。十年前确定的时间研究，需要自噬蛋白ATG2 在膜扩张期间，但是ATG2如何促进膜的生长仍然难以捉摸。我们的初步结果现在表明ATG2是一种新型脂质传播蛋白质家族的成员，建议膜扩张的第三个模型；从细胞器通过蛋白质的大量脂质传递介导的接触站点。实际上，ATG2同时绑定了多达20个脂质，一个数量级比实际上多。任何其他脂质转运蛋白，因此具有在生物发生过程中移动大量脂质的能力。我们显示在细胞中，ATG2在自噬体与ER之间的界面上积聚，强烈建议 Organel-organelle接触位点可能是脂质转移的位置。此外，我们开发了基因对人类中每种ATG2蛋白的编辑敲除，发现在没有ATG2的情况下，不仅可自噬体不扩展，但数百个囊泡在自噬体生物发生部位收集。这令人惊讶的观察表明，囊泡介导的膜的传递也可能是必不可少的，并且当不存在ATG2介导的脂质转运时，这些囊泡的融合被特异性破坏。通过此提案，我们希望描述ATG2如何与自噬体上的蛋白质一起使用 ER驱动脂质流。这可能涉及稳定器官 - 轨道接触位点所需的蛋白质和也可能涉及蛋白质感测或调节ER中的脂质产生。然后，我们将确定这种脂质如何流量与募集和利用贩运囊泡有关支持自噬体的募集和利用有关生长。