Abstract
Hepatitis B virus (HBV) genotype C causes prolonged chronic infection and increased risk for liver cancer than genotype B. Our previous work revealed lower replication capacity of wild-type genotype C2 than B2 isolates. HBV DNA replication is driven by pregenomic RNA, which is controlled by core promoter (CP) and further augmented by enhancer I (ENI) and enhancer II (ENII). DNA fragments covering these regulatory elements were amplified from B2 and C2 isolates to generate luciferase reporter constructs. As ENII is fully embedded in CP, we inserted HBV DNA fragments in the sense orientation to determine their combined activities, and in the antisense orientation to measure enhancer activities alone. Genotype B2 isolates displayed higher ENI+ENII+CP, ENII+CP, and ENII activities, but not ENI or ENI+ENII activity, than C2 isolates. The higher ENII+CP activity was partly attributable to 4 positions displaying genotype-specific variability. Exchanging CP region was sufficient to revert the replication phenotypes of several B2 and C2 clones tested. These results suggest that a weaker ENII and/or CP at least partly accounts for the lower replication capacities of wild-type C2 isolates, which could drive the subsequent acquisition of CP mutations. Such mutations increase genome replication and are implicated in liver cancer development.
Highlights
Hepatitis B virus (HBV) isolates worldwide can be classified into eight genotypes (A-H) and further divided into subgenotypes[1,2,3]
Cells seeded at 1–1.5 × 105/wells in 24-well plates were co-transfected with 0.3 μg of HBV reporter construct expressing firefly luciferase and 6.25 ng of Rluc plasmid expressing renilla luciferase, using TransIT-LT1 reagent (Mirus)
The enhancer or promoter/enhancer activity was calculated as the ratio of firefly luciferase activity over renilla luciferase activity, and the results shown were based on three repeat experiments
Summary
Hepatitis B virus (HBV) isolates worldwide can be classified into eight genotypes (A-H) and further divided into subgenotypes[1,2,3]. Genotypes B and C co-circulate in East Asian countries such as China Through their perinatal mode of transmission, these two genotypes are responsible for majority of chronic HBV infection worldwide. Infected individuals are initially positive for hepatitis B e antigen (HBeAg), a secreted version of viral core (capsid) protein, in the bloodstream. Independent studies demonstrated that genotype C patients seroconvert from HBeAg to anti-HBe about 10 years later than genotype B patients[4,5,6], and the prolonged phase of active viral DNA replication and protein expression increases the lifelong risk for liver cirrhosis and HCC7–10. To better understand the contrasting clinical features between these two major HBV genotypes would require their comparative functional studies We previously initiated such a study with a focus on the B2 and C2 subgenotypes prevalent in China. The element found to be more active in genotype B2 was exchanged between clones of the two subgenotypes so as to establish its relevance to the differential replication capacity
Sign-in/Register to view highlights & summary 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 callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
