Department of Biochemistry, and Department of Hepatobiliary and Pancreatic Surgery of the First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, China.
Key Laboratory of Vector Biology and Pathogen Control of Zhejiang Province, School of Life Sciences, Huzhou University, Huzhou, China.
EMBO J. 2023 Oct 4;42(19):e112814. doi: 10.15252/embj.2022112814. Epub 2023 Aug 28.
The regulation of autophagy initiation is a key step in autophagosome biogenesis. However, our understanding of the molecular mechanisms underlying the stepwise assembly of ATG proteins during this process remains incomplete. The Rab GTPase Ypt1/Rab1 is recognized as an essential autophagy regulator. Here, we identify Atg23 and Atg17 as binding partners of Ypt1, with their direct interaction proving crucial for the stepwise assembly of autophagy initiation complexes. Disruption of Ypt1-Atg23 binding results in significantly reduced Atg9 interactions with Atg11, Atg13, and Atg17, thus preventing the recruitment of Atg9 vesicles to the phagophore assembly site (PAS). Likewise, Ypt1-Atg17 binding contributes to the PAS recruitment of Ypt1 and Atg1. Importantly, we found that Ypt1 is phosphorylated by TOR at the Ser174 residue. Converting this residue to alanine blocks Ypt1 phosphorylation by TOR and enhances autophagy. Conversely, the Ypt1 phosphorylation mimic impairs both PAS recruitment and activation of Atg1, thus inhibiting subsequent autophagy. Thus, we propose TOR-mediated Ypt1 as a multifunctional assembly factor that controls autophagy initiation via its regulation of the stepwise assembly of ATG proteins.
自噬体生物发生的自噬起始的调节是一个关键步骤。然而，我们对该过程中 ATG 蛋白逐步组装的分子机制的理解仍不完整。Rab GTPase Ypt1/Rab1 被认为是一种重要的自噬调节剂。在这里，我们确定 Atg23 和 Atg17 是 Ypt1 的结合伴侣，它们的直接相互作用对于自噬起始复合物的逐步组装至关重要。破坏 Ypt1-Atg23 结合会导致 Atg9 与 Atg11、Atg13 和 Atg17 的相互作用显著减少，从而阻止 Atg9 囊泡募集到吞噬体组装位点 (PAS)。同样，Ypt1-Atg17 结合有助于 PAS 募集 Ypt1 和 Atg1。重要的是，我们发现 Ypt1 在 Ser174 残基处被 TOR 磷酸化。将该残基突变为丙氨酸可阻止 TOR 对 Ypt1 的磷酸化，并增强自噬。相反，Ypt1 磷酸化模拟物会损害 PAS 的募集和 Atg1 的激活，从而抑制随后的自噬。因此，我们提出 TOR 介导的 Ypt1 作为一种多功能组装因子，通过调节 ATG 蛋白的逐步组装来控制自噬起始。
