Hanada Takao, Noda Nobuo N, Satomi Yoshinori, Ichimura Yoshinobu, Fujioka Yuko, Takao Toshifumi, Inagaki Fuyuhiko, Ohsumi Yoshinori
Division of Molecular Cell Biology, National Institute for Basic Biology, Myodaiji, Okazaki, Japan.
J Biol Chem. 2007 Dec 28;282(52):37298-302. doi: 10.1074/jbc.C700195200. Epub 2007 Nov 6.
Autophagy is a bulk degradation process in eukaryotic cells; autophagosomes enclose cytoplasmic components for degradation in the lysosome/vacuole. Autophagosome formation requires two ubiquitin-like conjugation systems, the Atg12 and Atg8 systems, which are tightly associated with expansion of autophagosomal membrane. Previous studies have suggested that there is a hierarchy between these systems; the Atg12 system is located upstream of the Atg8 system in the context of Atg protein organization. However, the concrete molecular relationship is unclear. Here, we show using an in vitro Atg8 conjugation system that the Atg12-Atg5 conjugate, but not unconjugated Atg12 or Atg5, strongly enhances the formation of the other conjugate, Atg8-PE. The Atg12-Atg5 conjugate promotes the transfer of Atg8 from Atg3 to the substrate, phosphatidylethanolamine (PE), by stimulating the activity of Atg3. We also show that the Atg12-Atg5 conjugate interacts with both Atg3 and PE-containing liposomes. These results indicate that the Atg12-Atg5 conjugate is a ubiquitin-protein ligase (E3)-like enzyme for Atg8-PE conjugation reaction, distinctively promoting protein-lipid conjugation.
自噬是真核细胞中的一种大量降解过程；自噬体包裹细胞质成分以便在溶酶体/液泡中进行降解。自噬体的形成需要两个类泛素偶联系统，即Atg12和Atg8系统，它们与自噬体膜的扩张紧密相关。先前的研究表明这些系统之间存在层级关系；在Atg蛋白组织的背景下，Atg12系统位于Atg8系统的上游。然而，具体的分子关系尚不清楚。在这里，我们使用体外Atg8偶联系统表明，Atg12-Atg5偶联物，而非未偶联的Atg12或Atg5，强烈增强了另一种偶联物Atg8-PE的形成。Atg12-Atg5偶联物通过刺激Atg3的活性促进Atg8从Atg3转移至底物磷脂酰乙醇胺（PE）。我们还表明Atg12-Atg5偶联物与Atg3和含PE的脂质体相互作用。这些结果表明，Atg12-Atg5偶联物是一种用于Atg8-PE偶联反应的类泛素-蛋白连接酶（E3），独特地促进蛋白-脂质偶联。
