Lipid flux during autophagosome membrane biogenesis

摘要 巨细胞自噬是细胞内的应激反应途径，细胞通过它包装部分胞浆。 输送到溶酶体中。这种包装是由一种新的细胞器重新形成的，称为 自噬小体生长并包裹胞浆物质以供最终溶酶体降解的自噬小体多么 自噬小体的形成，尤其包括自噬小体为了扩张而提取脂类的地方 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 并提出要么自噬小体只是从预先存在的隔室(如内质网)中生长出来，要么是自噬小体 自噬小体是由来自不同部位的单个囊泡继续融合而形成的。 手机。此外，脂质生物发生途径与自噬密切相关，这表明一个或 更多的细胞器参与脂质的产生也可能与植物的生长密切相关 自噬小体。十多年前的时间研究表明，需要自噬蛋白ATG2 在膜的扩张过程中，ATG2如何促进膜的生长仍然是个谜。 我们的初步结果表明，ATG2是一个新的脂质转运蛋白家族的成员，并且 提出了膜扩张的第三种模式；通过蛋白质从细胞器大量输送脂肪- 经调解的联系地点。事实上，ATG2一次可以结合多达20种脂类，比虚拟的多一个数量级 任何其他脂类运输蛋白，因此在生物发生过程中有能力运输大量的脂类。我们展示了 在细胞中，ATG2聚集在自噬小体和内质网之间的交界处，强烈表明这一点 细胞器-细胞器接触部位可能是脂质转移的部位。此外，我们还开发了基因- 编辑了人类每种ATG2蛋白的敲除，并发现在没有ATG2的情况下，不仅 自噬小体不会膨胀，但数百个小泡聚集在自噬小体生物发生的位置。这 令人惊讶的观察表明，囊泡介导的膜传递也可能是必不可少的，而且 当ATG2介导的脂质运输缺失时，这些囊泡的融合就会被特别地破坏。 有了这个提议，我们希望描述ATG2如何与自噬体上的蛋白质和 ER来驱动脂质流动。这很可能涉及稳定细胞器-细胞器接触部位所需的蛋白质，以及 也可能涉及感受或调节内质网中脂质产生的蛋白质。然后我们将确定这种脂质是如何 流动与运输囊泡的招募和利用有关，以描述支持自噬体 成长。

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.