Functional Role of Hepatitis C Virus NS5A in the Regulation of Autophagy

Abstract

Many types of RNA viruses, including the hepatitis C virus (HCV), activate autophagy in infected cells to promote viral growth and counteract the host defense response. Autophagy acts as a catabolic pathway in which unnecessary materials are removed via the lysosome, thus maintaining cellular homeostasis. The HCV non-structural 5A (NS5A) protein is a phosphoprotein required for viral RNA replication, virion assembly, and the determination of interferon (IFN) sensitivity. Recently, increasing evidence has shown that HCV NS5A can induce autophagy to promote mitochondrial turnover and the degradation of hepatocyte nuclear factor 1 alpha (HNF-1α) and diacylglycerol acyltransferase 1 (DGAT1). In this review, we summarize recent progress in understanding the detailed mechanism by which HCV NS5A triggers autophagy, and outline the physiological significance of the balance between host-virus interactions.

Keywords: HCV; autophagy; chaperone-mediated autophagy; microautophagy; selective autophagy.

Figure 1

Genome organization of HCV. The viral genome of HCV consists of a positive-sense, single-stranded RNA with a length of approximately 9.6 kb. The viral genome of HCV includes an open reading frame (ORF) that is flanked by 5′- and 3′-untranslated regions (UTRs). The 5′-UTR contains an internal ribosome entry site (IRES) for stimulating translation, whereas the 3′-UTR harbors a pathogen-associated molecular pattern (PAMP) for inducing innate antiviral immunity. The ORF encodes a single polypeptide that is processed by proteases encoded by host cellular RNA and viral RNA to form structural proteins (core, E1, and E2) and nonstructural proteins (p7, NS2, NS3, NS4A, NS4B, NS5A, and NS5B). The core, E1, and E2 proteins are the major constituents of the HCV virion, and other viral proteins differentially regulate the replication of viral RNA and the assembly of infectious particles. The enzymatic activities of these viral proteins are indicated.

Figure 2

Domain structure of HCV NS5A. HCV NS5A contains an amphipathic helix (AH) at its N-terminal region and three specific domains: I, II, and III. They are flanked by two low-complexity sequences (LCSs), LCSI and LCSII. The functional role(s) of each domain in the HCV life cycle are specifically indicated. The interferon sensitivity-determining region (ISDR; a.a. 237~276) and the protein kinase R-binding domain (PKRBD; a.a. 237~302) are shown. The serine (S), threonine (T), and tyrosine (Y) residues within HCV NS5A that can be phosphorylated are indicated.

Figure 3

The autophagy process. Autophagy is categorized into three forms: macroautophagy, chaperone-mediated autophagy (CMA), and microautophagy. To initiate macroautophagy, various stressors, such as nutrient deprivation, induce nucleation of the isolation membrane (IM)/phagophore. The IM/phagophore expands and encloses cytosolic components to form mature autophagosomes surrounded by double membranes. The fusion of autophagosomes with lysosomes generates autolysosomes, and lysosomal hydrolases degrade the elements in the autolysosome interior. The termination of autophagy is coupled with the reformation of tubular lysosome (ALR) and autophagosomal component recycling (ACR), thus regenerating lysosomes and autophagic molecules. Selective autophagy is a specific form of macroautophagy that requires cargo receptors to recognize degradative cargoes via the ubiquitination of cargoes and other adaptor proteins, ultimately delivering these targeted proteins for autophagic degradation. CMA selectively targets degradative substrates harboring the pentapeptide “KFERQ”, which is recognized by HSC70. Through its binding to LAMP2 on the lysosomal membrane, the substrate-HSC70 complex is delivered into lysosomal lumen for degradation. Microautophagy occurs in endosomes and lysosomes, and involves the invagination and protrusion of membrane to sequester cargoes within intraluminal vesicles; these cargoes are then degraded by lysosomal hydrolases.

Figure 4

Current model of HCV NS5A-regulated autophagy. HCV NS5A induces mitochondrial depolarization and fission and triggers mitochondrial translocation to remove deformed mitochondria via mitophagy. Additionally, HCV NS5A acts as a bridge to connect HNF-1α and HSC70 on lysosomes, allowing the degradation of HNF-1α by LAMP2A-mediated CMA. In addition, HCV NS5A promotes the degradation of DGAT1 via endosomal microautophagy (eMI) through the binding of HSC70 and functional VPS4B.