School of Life Science and Technology, Tokyo Institute of Technology, Yokohama 226-8501, Japan.
Institute of Innovative Research, Tokyo Institute of Technology, Yokohama 226-8503, Japan.
Proc Natl Acad Sci U S A. 2018 Oct 9;115(41):10363-10368. doi: 10.1073/pnas.1806727115. Epub 2018 Sep 25.
The biogenesis of double-membrane vesicles called autophagosomes, which sequester and transport intracellular material for degradation in lysosomes or vacuoles, is a central event in autophagy. This process requires a unique set of factors called autophagy-related (Atg) proteins. The Atg proteins assemble to organize the preautophagosomal structure (PAS), at which a cup-shaped membrane, the isolation membrane (or phagophore), forms and expands to become the autophagosome. The molecular mechanism of autophagosome biogenesis remains poorly understood. Previous studies have shown that Atg2 forms a complex with the phosphatidylinositol 3-phosphate (PI3P)-binding protein Atg18 and localizes to the PAS to initiate autophagosome biogenesis; however, the molecular function of Atg2 remains unknown. In this study, we show that Atg2 has two membrane-binding domains in the N- and C-terminal regions and acts as a membrane tether during autophagosome formation in the budding yeast An amphipathic helix in the C-terminal region binds to membranes and facilitates Atg18 binding to PI3P to target the Atg2-Atg18 complex to the PAS. The N-terminal region of Atg2 is also involved in the membrane binding of this protein but is dispensable for the PAS targeting of the Atg2-Atg18 complex. Our data suggest that this region associates with the endoplasmic reticulum (ER) and is responsible for the formation of the isolation membrane at the PAS. Based on these results, we propose that the Atg2-Atg18 complex tethers the PAS to the ER to initiate membrane expansion during autophagosome formation.
双层膜囊泡（自噬体）的生物发生，自噬体可以隔离和运输细胞内物质，然后在溶酶体或液泡中降解，这是自噬的一个核心过程。这个过程需要一组称为自噬相关（Atg）蛋白的独特因子。Atg 蛋白组装起来以组织前自噬体结构（PAS），在这个结构中，一个杯状膜，即隔离膜（或噬泡）形成并扩展成为自噬体。自噬体生物发生的分子机制仍知之甚少。以前的研究表明，Atg2 与磷脂酰肌醇 3-磷酸（PI3P）结合蛋白 Atg18 形成复合物，并定位于 PAS 以启动自噬体生物发生；然而，Atg2 的分子功能仍然未知。在这项研究中，我们表明 Atg2 在 N 端和 C 端区域具有两个膜结合结构域，并在出芽酵母中形成自噬体时充当膜连接蛋白；在出芽酵母中形成自噬体时充当膜连接蛋白。C 端区域的一个两亲性螺旋与膜结合，并促进 Atg18 与 PI3P 的结合，将 Atg2-Atg18 复合物靶向 PAS。Atg2 的 N 端区域也参与该蛋白的膜结合，但对于 Atg2-Atg18 复合物靶向 PAS 是可有可无的。我们的数据表明，该区域与内质网（ER）相关联，并且负责在 PAS 处形成隔离膜。基于这些结果，我们提出 Atg2-Atg18 复合物将 PAS 连接到 ER 以在自噬体形成过程中启动膜扩展。
