RAB2 regulates the formation of autophagosome and autolysosome in mammalian cells
RAB2 regulates the formation of autophagosome and autolysosome in mammalian cells
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RAB2 调节哺乳动物细胞自噬体和自溶酶体的形成
DOI：
10.1080/15548627.2019.1596478
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发表时间：
2019-04-07
期刊：
影响因子：
13.3
通讯作者：
Sun, Qiming
中科院分区：
文献类型：
--
作者：
Ding, Xianming;Jiang, Xiao;Sun, Qiming
摘要多种来源的膜和蛋白质机制有助于构建功能性巨自噬/自噬结构。然而，潜在的分子机制仍然难以捉摸。在这里，我们表明，RAB 2连接高尔基体网络的自噬途径，通过提供膜和顺序从事不同的自噬机制。在未应激的细胞中，RAB 2主要存在于高尔基体中，如其与GOLGA 2/GM 130的相互作用和共定位所证明的。重要的是，自噬刺激使RAB 2与GOLGA 2解离以与ULK 1复合物相互作用，这促进了ULK 1复合物的募集以形成吞噬细胞。有趣的是，RAB 2似乎调节ULK 1激酶活性以传播自噬体形成的信号。随后，RAB 2转而与自噬体RUBCNL/PACER和STX 17相互作用，以进一步指定HOPS复合物的募集以形成自噬体。总之，我们的研究揭示了大量自噬调控的多价途径，并提供了高尔基体如何有助于形成不同自噬结构的机制见解。缩略语：会计师事务所：肌动蛋白β; ATG9：自噬相关9A; ATG14：自噬相关14例; ATG16L1：自噬相关16样1; BCAP 31：B细胞受体相关蛋白31; BECN 1：beclin 1; Ctrl：对照; CQ：氯喹; CTSD：组织蛋白酶D; DMSO：二甲亚砜; EBSS：Earle平衡盐溶液; EEA 1：早期内体抗原1; GDI：鸟嘌呤核苷酸解离抑制剂; GFP：绿色荧光蛋白; GOLGA 2：golgin A2; HOPS：同型融合和蛋白分选复合物; IP：免疫沉淀; KD：敲低; KO：敲除; LAMP 1：溶酶体相关膜蛋白1; LC 3：微管相关蛋白1轻链3; OE：过表达; PtdIns 3 K：III类磷脂酰肌醇3-激酶; SQSTM 1/p62：隔离体1; RAB 2：RAB 2A，RAS癌基因家族成员; RAB 7：RAB 7A，RAS癌基因家族成员; RAB 11：RAB 11 A，RAS癌基因家族成员; RUBCNL/PACER：Rubicon样自噬增强子; STX 17：突触融合蛋白17; TBC 1D 14：TBC 1结构域家族成员14; TFRC：转铁蛋白受体; TGOLN 2：trans-Golgi网络蛋白2; TUBB：微管蛋白β I类; ULK 1：unc-51样自噬激活激酶1; VPS 41：VPS 41，HOPS复合物亚基; WB：蛋白质印迹; WT：野生型; YPT 1：GTP结合蛋白YPT 1。
ABSTRACT Multiple sources contribute membrane and protein machineries to construct functional macroautophagic/autophagic structures. However, the underlying molecular mechanisms remain elusive. Here, we show that RAB2 connects the Golgi network to autophagy pathway by delivering membrane and by sequentially engaging distinct autophagy machineries. In unstressed cells, RAB2 resides primarily in the Golgi apparatus, as evidenced by its interaction and colocalization with GOLGA2/GM130. Importantly, autophagy stimuli dissociate RAB2 from GOLGA2 to interact with ULK1 complex, which facilitates the recruitment of ULK1 complex to form phagophores. Intriguingly, RAB2 appears to modulate ULK1 kinase activity to propagate signals for autophagosome formation. Subsequently, RAB2 switches to interact with autophagosomal RUBCNL/PACER and STX17 to further specify the recruitment of HOPS complex for autolysosome formation. Together, our study reveals a multivalent pathway in bulk autophagy regulation, and provides mechanistic insights into how the Golgi apparatus contributes to the formation of different autophagic structures. Abbreviations: ACTB: actin beta; ATG9: autophagy related 9A; ATG14: autophagy related 14; ATG16L1: autophagy related 16 like 1; BCAP31: B cell receptor associated protein 31; BECN1: beclin 1; Ctrl: control; CQ: chloroquine; CTSD: cathepsin D; DMSO: dimethyl sulfoxide; EBSS: Earle’s balanced salt solution; EEA1: early endosome antigen 1; GDI: guanine nucleotide dissociation inhibitor; GFP: green fluorescent protein; GOLGA2: golgin A2; HOPS: homotypic fusion and protein sorting complex; IP: immunoprecipitation; KD: knockdown; KO: knockout; LAMP1: lysosomal associated membrane protein 1; LC3: microtubule-associated protein 1 light chain 3; OE: overexpression; PtdIns3K: class III phosphatidylinositol 3-kinase; SQSTM1/p62: sequestosome 1; RAB2: RAB2A, member RAS oncogene family; RAB7: RAB7A, member RAS oncogene family; RAB11: RAB11A, member RAS oncogene family; RUBCNL/PACER: rubicon like autophagy enhancer; STX17: syntaxin 17; TBC1D14: TBC1 domain family member 14; TFRC: transferrin receptor; TGOLN2: trans-golgi network protein 2; TUBB: tubulin beta class I; ULK1: unc-51 like autophagy activating kinase 1; VPS41: VPS41, HOPS complex subunit; WB: western blot; WT: wild type; YPT1: GTP-binding protein YPT1.