Role of core protein mutations in the development of occult HBV infection

• Published in: Journal of hepatology 2021-06, Vol.74 (6), p.1303-1314
• Main Authors: Chen, Jingna, Liu, Bochao, Tang, Xi, Zheng, Xin, Lu, Jinhui, Zhang, Ling, Wang, Wenjing, Candotti, Daniel, Fu, Yongshui, Allain, Jean-Pierre, Li, Chengyao, Li, Linhai, Li, Tingting
• Format: Article
• Language: English
• Subjects: Adult; Amino Acid Substitution - genetics; Amino acids; Animals; Cell Line, Tumor; Core protein; Disease Models, Animal; DNA, Viral - blood; DNA, Viral - genetics; Female; Genomes; Genotype; Genotypes; Hepatitis B; Hepatitis B - blood; Hepatitis B - genetics; Hepatitis B - virology; Hepatitis B Core Antigens - biosynthesis; Hepatitis B e antigen; Hepatitis B e Antigens - biosynthesis; Hepatitis B surface antigen; Hepatitis B Surface Antigens - blood; Hepatitis B virus - genetics; Hepatocellular carcinoma; Humans; Infections; Life Sciences; Mice; Mice, Inbred BALB C; Microbiology and Parasitology; Mutagenesis, Site-Directed - methods; Mutants; Mutation; Nucleotide sequence; Occult HBV infection; Proteins; Regulatory sequences; Replication; Replicon; Site-directed mutagenesis; Transfection; Viral Core Proteins - genetics; Virology; Virus Replication - genetics
• ISSN: 0168-8278; 1600-0641
• DOI: 10.1016/j.jhep.2020.12.023

Occult HBV infection (OBI) is associated with transfusion-transmitted HBV infection and hepatocellular carcinoma. Studies on OBI genesis have concentrated on mutations in the S region and the regulatory elements. Herein, we aimed to determine the role of mutations in the core region on OBIs. An OBI strain (SZA) carrying 9 amino acid (aa) substitutions in the core protein/capsid (Cp) was selected by sequence alignment and Western blot analysis from 26 genotype B OBI samples to extensively explore the impact of Cp mutations on viral antigen production in vitro and in vivo. A large panel of 30 Cp replicons were generated by a replication-competent pHBV1.3 carrying SZA or wild-type (WT) Cp in a 1.3-fold over-length of HBV genome, in which the various Cp mutants were individually introduced by repairing site mutations of SZA-Cp or creating site mutations of WT-Cp by site-directed mutagenesis. The expression of HBcAg, HBeAg, and HBsAg and viral RNA was quantified from individual SZA and WT Cp mutant replicons in transfected Huh7 cells or infected mice, respectively. An analysis of the effect of Cp mutants on intracellular or extracellular viral protein production indicated that the W62R mutation in Cp had a critical impact on the reduction of HBcAg and HBeAg production during HBV replication, whereas P50H and/or S74G mutations played a limited role in influencing viral protein production invivo. W62R and its combination mutations in HBV Cp might massively affect HBcAg and HBeAg production during viral replication, which, in turn, might contribute to the occurrence of OBI. Occult hepatitis B virus infections (OBIs) have been found to be associated with amino acid mutations in the S region of the HBV, but the role of mutations in the core protein (Cp) remains unclear. In this study, an OBI strain (SZA) carrying 9 amino acid substitutions in Cp has been examined comprehensively in vitro and in vivo. The W62R mutation in Cp majorly reduces HBcAg and HBeAg production during HBV replication, potentially contributing to the occurrence of OBI. [Display omitted] •Amino acid (aa) substitutions in the core protein (Cp) of occult HBV infection (OBI) strains were identified.•An OBI mutant strain (SZA) carrying 9 aa substitutions in Cp was characterised in vitro (cells) and in vivo (mice).•The functional impact of individual aa substitutions in pHBV1.3-SZA Cp replicons was assessed in vitro and in vivo.•The Cp W62R mutation significantly reduced HBV protein production.•The Cp