HBV T1719G mutation reduced HBV replication through mutant Enh II and HBx protein in vitro

Abstract

Summary It was repeatedly reported that the hepatitis B virus (HBV) T1719G mutation was very common and related to progression and malignancy of liver disease. However, its effect on viral replication efficiency remains unclear. In this study, we aimed to evaluate the function and mechanisms of the T1719G mutation on viral replication capacity. Wild‐type and T1719G mutation‐bearing HBV1.2× plasmids were transfected into Huh7 and HepG2 cells, respectively, and HBV total RNA, 3.5 kb RNA and supernatant HBV DNA were assessed using real‐time PCR, hepatitis B surface antigen (HBsAg) and hepatitis B e antigen (HBeAg) levels were measured by time‐resolved fluoroimmunoassay. In order to assess Enh II activity and the binding capacity of HNF3β to Enh II sequence, dual‐luciferase assay and Chromatin immunoprecipitation (ChIP)‐PCR were employed, respectively. Simultaneously, the HBx or HBx‐mut (T1719G) plasmid was co‐transfected to evaluate the effect of HBx on viral replication. Our results showed that the T1719G mutation impaired viral replication efficacy compared with the wild type both by reducing Enh II activity and binding capacity of HNF3β with Enh II. And such reduction caused by T1719G mutation could be rescued by HBx protein. Our results show that the T1719G mutation decreases HBV viral replication capacity possibly by mutant HBx protein and altered Enh II activity.

Nucleos(t)ide analogues altered quasispecies composition of hepatitis B virus (HBV)‐resistant mutations in serum HBV DNA and serum HBV RNA

“…[13][14][15] There are few studies on the monitoring of HBV RNA resistance, and one study showed that patients with YMDD site mutations within 1 year of NAs treatment had significantly higher patients were HBeAg-negative, possibly due to extremely low cccDNA levels or mutations or epigenetic modifications that resulted in inactive cccDNA transcription. 21,22 One study has found that serum HBV RNA sequences are more similar to cccDNA sequences in liver than serum HBV DNA sequences. 23 Another study has also shown that high-throughput sequencing of serum HBV DNA and HBV RNA from HBV and HIV co-infected patients has shown that HBV RNA detects a higher proportion of wild quasispecies.…”

“…Most of these patients were HBeAg‐positive and had poor response to NAs antiviral therapy, resulting in insignificant depletion of cccDNA in hepatocytes. In Group 3, serum HBV RNA was not consistently detectable, and most of these patients were HBeAg‐negative, possibly due to extremely low cccDNA levels or mutations or epigenetic modifications that resulted in inactive cccDNA transcription 21,22 …”