Lipid flux during autophagosome membrane biogenesis

自噬体膜生物发生过程中的脂质通量

• 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
• 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

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.

总结 大自噬是细胞内应激反应途径，细胞通过该途径包装部分胞质溶胶 用于递送到溶酶体中。这种包装是通过一种新的细胞器的重新形成来进行的， 生长并包裹胞质物质以最终进行溶酶体降解的自噬体。如何 自噬体形成，尤其包括自噬体为了扩张而提取脂质的地方 它们的膜，50多年来一直是该领域的核心问题。出现了两种相互竞争的模式 这表明，要么自噬体只是从一个预先存在的隔室（如ER）中生长出来，要么自噬体只是从一个预先存在的隔室（如ER）中生长出来。 自噬体的形成是由从细胞内许多不同位点募集的单个囊泡的持续融合引起的。 cell.此外，脂质生物合成途径与自噬密切相关，这表明一种或多种脂质合成途径可能与自噬有关。 更多的细胞器参与脂质的产生也可能与细胞的生长密切相关。 自噬体十多年前的时间研究表明，自噬蛋白ATG 2是必需的， 在膜扩张过程中，ATG 2是如何促进膜生长的仍然是难以捉摸的。 我们的初步结果表明，ATG 2是一个新的脂质转运蛋白家族的成员， 提出了膜扩张的第三种模型;脂质从细胞器通过蛋白质的大量递送- 介导的接触点。事实上，ATG 2一次结合多达20种脂质，比实际上多一个数量级。 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。我们表明 在细胞中，ATG 2在自噬体和ER之间的界面处积累，强烈表明这一点。 细胞器-细胞器接触部位可能是脂质转运的部位。此外，我们还开发了基因- 在人类中编辑敲除每个ATG 2蛋白，并发现在缺乏ATG 2的情况下，不仅 自噬体不扩张，但数百个囊泡聚集在自噬体生物发生的位点。这 令人惊讶的观察表明，囊泡介导的膜递送可能也是必不可少的，并且 当不存在ATG 2介导的脂质转运时，这些囊泡的融合被特异性破坏。 有了这个提议，我们希望描述ATG 2如何与自噬体和自噬体上的蛋白质一起工作。 ER驱动脂质流动。很可能，这将涉及稳定细胞器-细胞器接触位点所需的蛋白质， 也可能涉及在ER中感测或调节脂质产生的蛋白。然后我们将确定这种脂质 流动与募集和利用运输囊泡来描述支持自噬体有关 增长