Double-stranded linear duck hepatitis B virus (DHBV) stably integrates at a higher frequency than wild-type DHBV in LMH chicken hepatoma cells.
Integration of hepadnavirus DNAs into host chromosomes can have oncogenic consequences. Analysis of host-viral DNA junctions of DHBV identified the terminally duplicated r region of the viral genome as a hotspot for integration. Since the r region is present on the 5' and 3' ends of double-stranded linear (DSL) hepadnavirus DNAs, these molecules have been implicated as integration precursors. We have produced a LMH chicken hepatoma cell line (LMH 66-1 DSL) which replicates exclusively DSL duck hepatitis B virus (DHBV) DNA. To test whether linear DHBV DNAs integrate more frequently than the wild type open circular DHBV DNAs, we have characterized the integration frequency in LMH 66-1 DSL cells by using a subcloning approach. This approach revealed that 83% of the LMH 66-1 DSL subclones contained new integrations, compared to only 16% of subclones from LMH-D2 cells replicating wild-type open circular DHBV DNA. Also, a higher percentage of the LMH 66-1 DSL subclones contained two or more new integrations. Mathematical analysis suggests that the DSL DHBV DNAs integrated stably once every three generations during subcloning whereas wild-type DHBV integrated only once every four to five generations. Cloning and sequencing of new integrations confirmed the r region as a preferred integration site for linear DHBV DNA molecules. One DHBV integrant was associated with a small deletion of chromosomal DNA, and another DHBV integrant occurred in a telomeric repeat sequence.
Gong
S S
SS
Marion Bessin Liver Research Center, Department of Medicine, The Jack and Pearl Resnick Campus of the Albert Einstein College of Medicine, Bronx, New York 10461, USA.
Jensen
A D
AD
Chang
C J
CJ
Rogler
C E
CE
D000818:Animals; D001483:Base Sequence; D002645:Chickens; D004270:DNA, Circular; D004279:DNA, Viral; D015611:Hepatitis B Virus, Duck / Q000235:genetics*; D008969:Molecular Sequence Data; D012091:Repetitive Sequences, Nucleic Acid; D016615:Telomere; D014407:Tumor Cells, Cultured; D014771:Virion; D016662:Virus Integration
D004270:DNA, Circular; D004279:DNA, Viral