Abstract
Although RNA splicing of hepatitis B virus (HBV) is a commonly observed in livers of hepatitis B patients as well as in the cultured cells replicating the viral genome, its biological significance in the HBV life cycle and the detailed regulatory mechanisms are still largely unclear. In this study, we found cell-type dependency of HBV splicing of the 3.5 kb pregenomic RNA, which is efficiently spliced in human hepatoma cells but not in cells derived from human hepatic stellate, mouse hepatoma and human non-hepatic cells. It may be likely that RNA splicing is one of the determinants of host range restriction of HBV. Given the finding indicating the difference in cell-type dependency of the splicing efficiency between HBV and simian virus 40, we carried out intron-swapping experiments. The results suggest the presence of putative exonic splicing enhancer that possibly works in the cell-type dependent fashion. Together with further mutational analyses, a novel 50-nt intronic splicing silencer, whose secondary structure is well conserved among the HBV strains, was identified. It appears that this intronic silencer functions effectively independent of cell backgrounds.
Highlights
Hepatitis B virus (HBV) infection is a major global problem despite the availability of an efficacious vaccine
Northern blotting with pregenome/precore probe showed that two bands corresponding to unspliced 3.5 kb RNA and 3.5 kb RNA -derived, 2.2 kb spliced RNA (SP1) lacking ∼1.2 kb intron, respectively, were more abundant in HepG2 cells compared to HEK293 cells (Figure 1A)
It is of interest that the ratio of spliced/unspliced 3.5 kb RNAs was higher in all five kinds of cell lines derived from human hepatocellular carcinoma (HCC) (HepG2, HuH-7, PLC/PRF/5, FLC7, and ORL8c), compared to other cell types; mouse hepatoma (Hepa1c1c7), human hepatic stellate cell derived (TWNT4), human non-hepatic cells (A549, HEK293, HEK293T, Caco2, C33A and HeLa)
Summary
Hepatitis B virus (HBV) infection is a major global problem despite the availability of an efficacious vaccine. HBV is a member of the Hepadnaviridae family and contains a 3.2 kb partially double-stranded relaxed circular DNA genome with four open reading frames encoding seven proteins. Precore mRNA codes for precore antigen or HBeAg. The pregenomic RNA serves as a template for the synthesis of HBV DNA and as the mRNA of core antigen (HBcAg) and polymerase. In cultured human hepatoma cells transfected with the viral genome, synthesis of multiple spliced RNAs derived from 3.5 kb RNA has been commonly observed among HBV isolates. It has been reported that up to 80% of intracellular capsids contain the viral DNAs originated from the spliced RNAs in HBV genome-replicating hepatoma cells
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
