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The -1G mutant HBV is more prevalent in individuals co-infected with HIV/HBV than in individuals infected with HBV alone and in some cases is the dominant virus in circulation. This mutant is created by the deletion of a dGMP (-1G) from the guanine rich homopolymer sequence located at nts 2,085-2,090 (numbering from EcoRI site as position 1) in the HBV core gene. This deletion causes a frameshift generating a premature stop codon at (64) Asn in the HBV core gene (codon 93 in the precore gene), that truncates the precore protein, precursor of the secreted hepatitis B "e" antigen (HBeAg), and the core protein which forms the viral nucleocapsid. However, the replication phenotype of the -1G mutant HBV is unknown. An in vitro cell culture model in which hepatoma cells were transiently transfected with infectious cDNAs was used to show that the -1G mutant HBV is incapable of autonomous replication and, as expected, replication was restored to wild-type (wt) levels by supplying HBV core protein in trans. Although the -1G mutation had no deleterious effect on intracellular HBV-DNA levels, high levels of -1G mutant HBV relative to wt HBV reduced virus secretion and HBeAg secretion relative to empty vector controls. Importantly, the -1G mutant HBV also caused intracellular retention of truncated precore protein in the endoplasmic reticulum (ER) and Golgi apparatus. Together, these effects may be contributing to the increased pathology observed in the setting of HIV/HBV co-infection., (Copyright © 2012 Wiley Periodicals, Inc.)

Unlabelled: Defective hepatitis B virus DNA (dDNA) is reverse-transcribed from spliced hepatitis B virus (HBV) pregenomic messenger RNA (pgRNA) and has been identified in patients with chronic HBV (CH-B). The major 2.2-kb spliced pgRNA encodes a novel HBV gene product, the hepatitis B splice protein (HBSP) via a deletion and frame shift within the polymerase. Although spliced RNA and HBSP expression have been associated with increased HBV DNA levels and liver fibrosis, the role of dDNA in HBV-associated disease is largely undefined. Our aims were to (1) compare the relative proportions of dDNA (% dDNA) in a range of HBV-infected serum samples, including patients with human immunodeficiency virus (HIV)/HBV coinfection and HBV-monoinfected persons with differing severities of liver disease, and (2) determine the effect of mutations associated with drug resistance on defective DNA production. Defective DNA was detected in 90% of persons with CH-B. There was no significant difference in the relative abundance of dDNA between the monoinfected and HIV/HBV-coinfected groups. We also found no association between the % dDNA and alanine aminotransferase, hepatitis B e antigen status, HBV DNA levels, fibrosis levels, compensated or decompensated liver cirrhosis, genotype, or drug treatment. However, the % dDNA was significantly lower in individuals infected with lamivudine-resistant (LMV-R) HBV compared with wild-type HBV (P < 0.0001), indicating that antiviral drug resistance alters the balance between defective and genomic length DNA in circulation. Experiments in vitro using HBV encoding LMV-R mutations confirmed these results., Conclusion: Our results identified no association between dDNA and parameters associated with disease status and suggested that the relative abundance of dDNA is largely dependent on the integrity of the HBV polymerase and is unrelated to the severity of liver disease.

Hepatitis B virus (HBV) infection is a major cause of liver-related morbidity and mortality. Toll-like receptors (TLRs) have recently been recognized to play an important role in the pathogenesis of chronic hepatitis B (CH-B). Furthermore, manipulation of TLR signalling pathways shows potential as an antiviral therapeutic strategy. Whether hepatocytes themselves possess intact TLR signalling pathways remains controversial. It is critical that cell culture models be developed to allow investigation of the interaction between HBV and the TLR signalling pathways. We have screened three hepatocyte cell lines for the integrity of pro-inflammatory responses and antiviral cytokines following stimulation with interleukin-1 (IL-1) and different TLR ligands. We observed that Huh-7, HepG2 and PH5CH8 cells selectively responded to IL-1 and TLR2 ligands, leading to the activation of NF-κB. In addition, the PH5CH8 cell lines were able to induce type 1 interferon (IFN) via both TLR3 and RIG-I following stimulation with poly I:C, HepG2 cells mounted an IFN response via RIG-I only, whereas Huh-7 cells were unresponsive. We conclude that the hepatocyte cell lines investigated display a repertoire of TLR signalling, albeit limited, suggesting that hepatocytes may themselves play an active role in innate immune responses to viruses such as HBV. Furthermore, particular hepatoma cell lines are suitable for investigating the interaction between HBV and hepatocyte-expressed pattern recognition receptors. [ABSTRACT FROM AUTHOR]