Hepatitis C virus depends on E-cadherin as an entry factor and regulates its expression in epithelial-to-mesenchymal transition

Abstract

Hepatitis C virus (HCV) enters the host cell through interactions with a cascade of cellular factors. Although significant progress has been made in understanding HCV entry, the precise mechanisms by which HCV exploits the receptor complex and host machinery to enter the cell remain unclear. This intricate process of viral entry likely depends on additional yet-to-be-defined cellular molecules. Recently, by applying integrative functional genomics approaches, we identified and interrogated distinct sets of host dependencies in the complete HCV life cycle. Viral entry assays using HCV pseudoparticles (HCVpps) of various genotypes uncovered multiple previously unappreciated host factors, including E-cadherin, that mediate HCV entry. E-cadherin silencing significantly inhibited HCV infection in Huh7.5.1 cells, HepG2/miR122/CD81 cells, and primary human hepatocytes at a postbinding entry step. Knockdown of E-cadherin, however, had no effect on HCV RNA replication or internal ribosomal entry site (IRES)-mediated translation. In addition, an E-cadherin monoclonal antibody effectively blocked HCV entry and infection in hepatocytes. Mechanistic studies demonstrated that E-cadherin is closely associated with claudin-1 (CLDN1) and occludin (OCLN) on the cell membrane. Depletion of E-cadherin drastically diminished the cell-surface distribution of these two tight junction proteins in various hepatic cell lines, indicating that E-cadherin plays an important regulatory role in CLDN1/OCLN localization on the cell surface. Furthermore, loss of E-cadherin expression in hepatocytes is associated with HCVinduced epithelial-to-mesenchymal transition (EMT), providing an important link between HCV infection and liver cancer. Our data indicate that a dynamic interplay among E-cadherin, tight junctions, and EMT exists and mediates an important function in HCV entry.epatitis C virus (HCV), a member of the Hepacivirus genus in the Flaviviridae family, is an enveloped, single-stranded and positive-sense RNA virus that infects humans and other higher primates, with a selective tropism to the liver. The virus is estimated to infect 2.8% of the world's population (1), and has evolved into a major causative agent of end-stage liver diseases, including cirrhosis and hepatocellular carcinoma (HCC) (2). Chronic hepatitis C is also the leading indication for liver transplantation in the United States (3). To date, a protective vaccine is not available. Current therapeutic regimens applying direct-acting antivirals with or without ribavirin have made it possible to cure the majority of patients with HCV (4).HCV infection gains chronicity in ∼75-85% of patients, facilitated by various viral mechanisms to evade host immune responses and exploit the cellular machinery (5). The replication cycle of HCV in the host cell consists of multiple sequential steps, beginning with the lipo-viro-particle binding and entry, followed by viral RNA translation and replication, packaging and assembly of the virion, and finally secretion from host cells (6, 7). Each...

Infection of Hepatocytes With HCV Increases Cell Surface Levels of Heparan Sulfate Proteoglycans, Uptake of Cholesterol and Lipoprotein, and Virus Entry by Up-regulating SMAD6 and SMAD7

“…Among them, TGF-β and NF-κB components were shown to be upregulated by HCV infection. Indeed, both pathways have previously been shown to be activated by HCV 4, 31–33 . To explore whether these signaling pathways are involved in the HCV-triggered induction of I-SMADs, we first treated Huh7 cells with SB-431542, a potent and specific inhibitor of TGF-β signaling.…”

Mice Expressing Minimally Humanized CD81 and Occludin Genes Support Hepatitis C Virus Uptake In Vivo

“…In mouse cells, the HCV life cycle is blocked or inefficiently supported at multiple steps, in particular, viral entry and HCV RNA replication (reviewed in reference 5). A surprisingly large number of host factors have been shown to be important in the uptake of HCV into human hepatocytes, including glycosaminoglycans (GAGs) present on heparan sulfate proteoglycans (HSPGs) (6), low-density-lipoprotein receptor (LDLR) (7), CD81 (8), scavenger receptor class B member 1 (SCARB1) (9), the tight junction (TJ) proteins claudin-1 (CLDN1) (10) and occludin (OCLN) (11,12), the receptor tyrosine kinases epidermal growth factor receptor (EGFR) and ephrin receptor A2 (EphA2) (13), the cholesterol transporter Niemann-Pick C1-like 1 (NPC1L1) (14), transferring receptor 1 (TfR1) (15), the cell death-inducing DFFA-like effector b (CIDEB) (16), and E-cadherin (17). Of those, differences in the sequences of CD81 and OCLN between the murine and human orthologues can at least in part explain the lower efficiency of HCV uptake by rodent versus human cells.…”

Claudins in viral infection: from entry to spread

“…As an RNA virus, HCV exists as a highly variable population of quasispecies and is classified into seven major genotypes. HCV infects hepatocytes via a complex, multi-step process involving several host proteins, including glycosaminoglycans [8, 9, 57, 83], low density lipoprotein receptor [2], very-low-density lipoprotein receptor [95], tetraspanins CD81 [76] and CD63 [74], scavenger receptor class B type 1 [82, 103], tight junction proteins CLDN1 [25] and occludin [77], E-cadherin [58], receptor tyrosine kinases such as epidermal growth factor receptor (EGFR) [59, 106], serum response factor binding protein 1 [35], cholesterol transporter Niemann-Pick C1-like 1 [80] and transferrin receptor 1 [62]. Following virion internalization [26, 68], fusion is triggered by low pH in the endosome [56].…”