Department of Biochemistry and Department of Cardiology of the Second Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, China.
National Center for Protein Science Shanghai, Institute of Biochemistry and Cell Biology, Shanghai Institutes of Biological Sciences, Chinese Academy of Sciences, Shanghai, China.
Autophagy. 2021 May;17(5):1157-1169. doi: 10.1080/15548627.2020.1752471. Epub 2020 Apr 15.
The fusion of autophagosomes and endosomes/lysosomes, also called autophagosome maturation, ensures the degradation of autophagic cargoes. It is an important regulatory step of the macroautophagy/autophagy process. STX17 is the key autophagosomal SNARE protein that mediates autophagosome maturation. Here, we report that the acetylation of STX17 regulates its SNARE activity and autophagic degradation. The histone acetyltransferase CREBBP/CBP and the deacetylase HDAC2 specifically regulate the acetylation of STX17. In response to cell starvation and MTORC1 inhibition, the inactivation of CREBBP leads to the deacetylation of STX17 at its SNARE domain. This deacetylation promotes the interaction between STX17 and SNAP29 and the formation of the STX17-SNAP29-VAMP8 SNARE complex with no effect on the recruitment of STX17 to autophagosomal membranes. Deacetylation of STX17 also enhances the interaction between STX17 and the tethering complex HOPS, thereby further promoting autophagosome-lysosome fusion. Our study suggests a mechanism by which acetylation regulates the late-stage of autophagy, and possibly other STX17-related intracellular membrane fusion events. ACTB: actin beta; CREBBP/CBP: CREB binding protein; Ctrl: control; GFP: green fluorescent protein; GST: glutathione S-transferase; HDAC: histone deacetylase; HOPS: homotypic fusion and protein sorting complex; KO: knockout; LAMP1/2: lysosomal associated membrane protein 1/2; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MEFs: mouse embryonic fibroblasts; MS: mass spectrometry; MTORC1: mechanistic target of rapamycin kinase complex 1; NAM: nicotinamide; PtdIns3K: phosphatidylinositol 3-kinase; RFP: red fluorescent protein; SNAP29: synaptosome associated protein 29; SNARE: soluble N-ethylamide-sensitive factor attachment protein receptor; SQSTM1/p62: sequestosome 1; STX17: syntaxin 17; TSA: trichostatin A; TSC1/2: TSC complex subunit 1/2; VAMP8: vesicle associated membrane protein 8; WT: wild type.
自噬体与内体/溶酶体融合，也称为自噬体成熟，确保了自噬 cargo 的降解。它是巨自噬/自噬过程的一个重要调控步骤。STX17 是介导自噬体成熟的关键自噬体 SNARE 蛋白。在这里，我们报告 STX17 的乙酰化调节其 SNARE 活性和自噬降解。组蛋白乙酰转移酶 CREBBP/CBP 和去乙酰化酶 HDAC2 特异性调节 STX17 的乙酰化。在细胞饥饿和 MTORC1 抑制的反应中，CREBBP 的失活导致 STX17 的 SNARE 结构域去乙酰化。这种去乙酰化促进了 STX17 与 SNAP29 的相互作用，并形成 STX17-SNAP29-VAMP8 SNARE 复合物，而对 STX17 招募到自噬体膜没有影响。STX17 的去乙酰化也增强了 STX17 与连接复合物 HOPS 之间的相互作用，从而进一步促进自噬体-溶酶体融合。我们的研究提出了一种机制，即乙酰化调节自噬的晚期阶段，可能还有其他与 STX17 相关的细胞内膜融合事件。ACTB：肌动蛋白β；CREBBP/CBP：CREB 结合蛋白；Ctrl：对照；GFP：绿色荧光蛋白；GST：谷胱甘肽 S-转移酶；HDAC：组蛋白去乙酰化酶；HOPS：同源融合和蛋白分选复合物；KO：敲除；LAMP1/2：溶酶体相关膜蛋白 1/2；MAP1LC3/LC3：微管相关蛋白 1 轻链 3；MEFs：小鼠胚胎成纤维细胞；MS：质谱法；MTORC1：雷帕霉素激酶复合物 1 的机械靶标；NAM：烟酰胺；PtdIns3K：磷酸肌醇 3-激酶；RFP：红色荧光蛋白；SNAP29：突触相关蛋白 29；SNARE：可溶性 N-乙基酰胺敏感因子附着蛋白受体；SQSTM1/p62：自噬体 1；STX17：突触融合蛋白 17；TSA：曲古抑菌素 A；TSC1/2：TSC 复合物亚基 1/2；VAMP8：囊泡相关膜蛋白 8；WT：野生型。
