Hepatitis B Virus-Specific miRNAs and Argonaute2 Play a Role in the Viral Life Cycle

Abstract

Disease-specific serum miRNA profiles may serve as biomarkers and might reveal potential new avenues for therapy. An HBV-specific serum miRNA profile associated with HBV surface antigen (HBsAg) particles has recently been reported, and AGO2 and miRNAs have been shown to be stably associated with HBsAg in serum. We identified HBV-associated serum miRNAs using the Toray 3D array system in 10 healthy controls and 10 patients with chronic hepatitis B virus (HBV) infection. 19 selected miRNAs were then measured by quantitative RT-PCR in 248 chronic HBV patients and 22 healthy controls. MiRNA expression in serum versus liver tissue was also compared using biopsy samples. To examine the role of AGO2 during the HBV life cycle, we analyzed intracellular co-localization of AGO2 and HBV core (HBcAg) and surface (HBsAg) antigens using immunocytochemistry and proximity ligation assays in stably transfected HepG2 cells. The effect of AGO2 ablation on viral replication was assessed using siRNA. Several miRNAs, including miR-122, miR-22, and miR-99a, were up-regulated at least 1.5 fold (P<2E-08) in serum of HBV-infected patients. AGO2 and HBcAg were found to physically interact and co-localize in the ER and other subcellular compartments. HBs was also found to co-localize with AGO2 and was detected in multiple subcellular compartments. Conversely, HBx localized non-specifically in the nucleus and cytoplasm, and no interaction between AGO2 and HBx was detected. SiRNA ablation of AGO2 suppressed production of HBV DNA and HBs antigen in the supernatant.ConclusionThese results suggest that AGO2 and HBV-specific miRNAs might play a role in the HBV life cycle.

Hepatitis B Viral RNA Directly Mediates Down-regulation of the Tumor Suppressor MicroRNA miR-15a/miR-16-1 in Hepatocytes

“…Our data suggest a net negative impact of the microRNA machinery on HBV replication, which is contradictory to a recent report indicating a positive role of Ago2 in HBV replication (23). To address this discrepancy, we further tested knocking down the essential component of the microprocessor DGCR8 and the microRNA maturation factor Dicer, both of which were efficiently down-regulated in siRNA-treated cells (data not shown).…”

“…by knocking down Ago2), we demonstrated that inactivation of the microRNA machinery benefits HBV replication in HepG2 cells. Our results disagree with a recent report indicating that a siRNA against Ago2 impaired HBV replication in T23 cells (23). Although the source for this discrepancy remains unclear, we note that T23 cells carry an integrated HBV genome whereas our experiments were performed on freshly transfected HepG2 cells.…”

“…We first established that the microRNA machinery is inhibitory to HBV infection on the basis of multiple lines of functional evidence, which is contrary to the conclusion reached in a recent report (23). We found that miR-15a/miR-16-1 is able to directly target HBV transcripts, particularly that encoding for HBx, which is also responsible for reprogramming the expression of multiple microRNAs, including miR-15a/miR-16-1.…”

Differences in serum microRNA profiles in hepatitis B and C virus infection

“…Analysis of differential microRNA expression in liver tissues has revealed HCV-and HBV-specific microRNAs as well as microRNAs associated with the stage of liver disease. 5e9 MicroRNA levels in the liver have been found to be correlated with serum levels for a number of microRNAs, 10,11 suggesting that serum microRNAs might act as a surrogate measure of microRNA activity in the liver. While RNA typically has a short-half life and is quickly degraded by RNases, microRNAs tend to exist stably in serum when bound to argonaute proteins such as AGO2 as part of the RNA-induced silencing complex, the molecular scaffold that facilitates interaction of a microRNA with its target sequence.…”

Manipulation of Cellular Processing Bodies and Their Constituents by Viruses

“…This interaction is presumed to be important for the intracellular movement of viral components. Hepatitis B virus (HBV), core (HBc), and surface (HBs) proteins but not HBx protein interact and colocalize with Ago2 in the ER and other subcellular compartments, while Ago2 appears to play a role in HBV life cycle (Hayes et al, 2012). The Kaposi's sarcoma-associated herpesvirus SOX protein induces internal cleavage of host mRNA, which are then degraded by the 5¢-3¢ cellular exonuclease Xrn1 (Covarrubias et al, 2011).…”