INRS-Institut Armand-Frappier, Institut National de la Recherche Scientifique, Laval, Canada.
Sci Rep. 2017 Jan 9;7:40351. doi: 10.1038/srep40351.
Hepatitis C virus (HCV) infection induces intracellular membrane rearrangements, thus forming a membranous web (MW) in which HCV replication and assembly occur. The HCV-induced MW is primarily composed of double membrane vesicles (DMVs) transfused by multi-membrane vesicles. The autophagy machinery has been proposed to participate in the formation of such vesicles. However, no clear evidence has been found linking autophagy to the formation of these DMVs. In this study, we evaluated the role of the autophagy elongation complex (ATG5-12/16L1) in HCV replication and MW formation. Using a dominant negative form of ATG12 and an siRNA approach, we demonstrated that the ATG5-12 conjugate, but not LC3-II formation, is crucial for efficient viral replication. Furthermore, purification of HCV MW revealed the presence of ATG5-12 and ATG16L1 along with HCV nonstructural proteins. Interestingly, LC3 was not recruited along with the elongation complex to the site of viral replication. Finally, inhibition of the elongation complex, but not LC3, greatly impaired the formation of the wild-type MW phenotype. To our knowledge, this study provides the first evidence of the involvement of autophagy proteins in the formation of wild-type MWs.
丙型肝炎病毒 (HCV) 感染诱导细胞内膜重排，从而形成 HCV 复制和组装发生的膜网络 (MW)。HCV 诱导的 MW 主要由多膜囊泡转导的双膜囊泡 (DMVs) 组成。自噬机制被认为参与了这些囊泡的形成。然而，尚未发现明确的证据将自噬与这些 DMVs 的形成联系起来。在这项研究中，我们评估了自噬伸长复合物 (ATG5-12/16L1) 在 HCV 复制和 MW 形成中的作用。使用 ATG12 的显性负形式和 siRNA 方法，我们证明了 ATG5-12 缀合物，而不是 LC3-II 的形成，对于有效的病毒复制至关重要。此外，HCV MW 的纯化揭示了 ATG5-12 和 ATG16L1 与 HCV 非结构蛋白一起存在。有趣的是，LC3 没有与伸长复合物一起募集到病毒复制部位。最后，伸长复合物的抑制，而不是 LC3，极大地损害了野生型 MW 表型的形成。据我们所知，这项研究首次提供了自噬蛋白参与形成野生型 MW 的证据。
