Earliest hepatitis B virus-hepatocyte genome integration: sites, mechanism, and significance in carcinogenesis

Abstract

Hepatocellular carcinoma (HCC) is the fifth most widespread cancer responsible for one fourth of cancer-related deaths globally. Persistent infection with hepatitis B virus (HBV) remains the main cause of HCC summing up to 50% of its causative etiology. Our recent studies, supported by findings from others, uncovered that HBV and its close relative woodchuck hepatitis virus (WHV) integrate into hepatocyte genome almost immediately, hence in minutes after infection. Retrotransposons and genes with translocation potential were found to be frequent sites of HBV insertions, suggesting a mechanism of HBV DNA spread across liver genome from the earliest stages after virus invasion. Many other genes were identified as the sites of early hepadnavirus merges in human hepatocyte-like lines infected de novo with HBV and in natural woodchuck WHV infection model. It was uncovered that head-to-tail joins (HTJs) prevail among the earliest virus-host fusions, implying their formation via the non-homologous-end-joining (NHEJ) pathway. Overlapping homologous junctions resulting from the micro-homology-mediated-overlapping-joining (MHMOJ) were rarely detected. Formation of the initial HTJs coincided with strong induction of reactive oxygen species (ROS) and transient appearance of inducible nitric oxide (iNOS). This was accompanied by cell DNA damage and activation of the poly(ADP-ribose) polymerase 1 (PARP1)-mediated host DNA repair machinery, which may explain predominant HTJ format of the first virus-host fusions. Identification of initial integration sites and resulting alterations in hepatocyte phenotype may pave a way to discovery of reliable markers of HBV-triggered HCC, including HCC resulting from occult HBV infection. Our research strongly argues that HBV is an ultimate human carcinogen capable of initiation of a pro-oncogenic process immediately after first contact with a susceptible host.

Keywords

Hepatitis B virus / woodchuck hepatitis virus / virus-host genomic integration / virus initial integration sites / retrotransposons / virus-induced oxidative DNA damage / kinetics of DNA repair response / oncogenesis of hepatocellular carcinoma

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