ERManI (Endoplasmic Reticulum Class I α-Mannosidase) Is Required for HIV-1 Envelope Glycoprotein Degradation via Endoplasmic Reticulum-associated Protein Degradation Pathway*
ERManI (Endoplasmic Reticulum Class I α-Mannosidase) Is Required for HIV-1 Envelope Glycoprotein Degradation via Endoplasmic Reticulum-associated Protein Degradation Pathway*
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DOI：
10.1074/jbc.m115.675207
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发表时间：
2015-07
期刊：
影响因子：
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通讯作者：
Tao Zhou;Dylan A Frabutt;K. Moremen;Yong-Hui Zheng
中科院分区：
文献类型：
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作者：
Tao Zhou;Dylan A Frabutt;K. Moremen;Yong-Hui Zheng
背景资料：HIV-1包膜糖蛋白（Env）在感染细胞后靶向内质网（ER）相关蛋白降解（ERAD）途径进行降解。结果：ER Ⅰ类α-甘露糖苷酶（ERManI）与Env相互作用，启动了Env的降解过程。结论：ERManI对Env的降解是必需的。重要性：这些发现定义了一种新的内源性和潜在的治疗适用的抗逆转录病毒机制，通过靶向Env降解。我们以前报道过线粒体转运蛋白（TSPO）通过内质网（ER）相关蛋白降解（ERAD）途径诱导HIV-1包膜（Env）降解，但其机制尚不清楚。在这里，我们研究了四种ER相关糖苷水解酶家族47（GH47）α-甘露糖苷酶、ERManI和ER降解增强α-甘露糖苷酶样（EDEM）蛋白1、2和3如何参与Env降解过程。这四种α-甘露糖苷酶的异位表达揭示了只有ERManI以剂量依赖性方式抑制HIV-1 Env表达。此外，使用CRISPR/Cas9技术基因敲除ERManI基因MAN 1B 1破坏了TSPO介导的Env降解。生化研究表明，HIV-1 Env与ERManI相互作用，并且在ERManI胞质、跨膜、内腔茎和内腔催化结构域之间，催化结构域在Env-ERManI相互作用中起关键作用。此外，功能研究表明，通过定点诱变的催化位点的失活破坏ERManI活性。这些研究确定ERManI是ER相关蛋白降解途径中的关键GH47 α-甘露糖苷酶，其启动Env降解，并表明其催化结构域和酶活性在该过程中发挥重要作用。
Background: HIV-1 envelope (Env) glycoprotein is targeted to endoplasmic reticulum (ER)-associated protein degradation (ERAD) pathway for degradation after infecting cells. Results: ER class I α-mannosidase (ERManI) interacts with Env and initiates this degradation process. Conclusion: ERManI is essential for the Env degradation. Significance: These findings define a novel endogenous and potential therapeutically applicable antiretroviral mechanism by targeting Env for degradation. Previously, we reported that the mitochondrial translocator protein (TSPO) induces HIV-1 envelope (Env) degradation via the endoplasmic reticulum (ER)-associated protein degradation (ERAD) pathway, but the mechanism was not clear. Here we investigated how the four ER-associated glycoside hydrolase family 47 (GH47) α-mannosidases, ERManI, and ER-degradation enhancing α-mannosidase-like (EDEM) proteins 1, 2, and 3, are involved in the Env degradation process. Ectopic expression of these four α-mannosidases uncovers that only ERManI inhibits HIV-1 Env expression in a dose-dependent manner. In addition, genetic knock-out of the ERManI gene MAN1B1 using CRISPR/Cas9 technology disrupts the TSPO-mediated Env degradation. Biochemical studies show that HIV-1 Env interacts with ERManI, and between the ERManI cytoplasmic, transmembrane, lumenal stem, and lumenal catalytic domains, the catalytic domain plays a critical role in the Env-ERManI interaction. In addition, functional studies show that inactivation of the catalytic sites by site-directed mutagenesis disrupts the ERManI activity. These studies identify ERManI as a critical GH47 α-mannosidase in the ER-associated protein degradation pathway that initiates the Env degradation and suggests that its catalytic domain and enzymatic activity play an important role in this process.