Hepatitis B virus X gene differentially modulates subgenotypes F1b and F4 replication

CAMPOS H RODOLFO; SPERONI, MICAELA; ELIZALDE, MARÍA MERCEDES; FLICHMAN DIEGO

Melbourne

Congreso; The International HBV Meeting; 2019

Hepatitis B Foundation

Introduction. Hepatitis B virus (HBV) is classified into ten genotypes and numerous subgenotypes (sgt). SgtF1b and sgtF4, native of Latin America, have been associated with differences in clinical and virological characteristics. Particularly, sgtF1b has been related with progression to hepatocellular carcinoma. Hepatitis B virus X protein (HBx) is a multifunctional regulatory protein associated with the modulation of viral transcription and replication. Objective. To analyze the role of HBx in sgtF1b and sgtF4 replication. Methodology. Huh7 cells were transfected with sgtF1b and sgtF4 wild type, HBx minus (HBV X(-)), HBx chimera wild type (F1b/XF4, F4/XF1b) and HBx minus chimera (F1b/X(-)F4 and F4/X(-)F1b) full-length genomes. Intracellular replicative intermediates and viral progeny were evaluated by Southern Blot and qPCR, respectively. Pregenomic mRNA levels were analyzed by RT-qPCR. Results. Silencing of HBx revealed remarkably differences in the replicative capacity of sgtF1b and sgtF4 mutants. The absence of HBx increased sgtF1b X(-) transcription and replication by more than 2.5 fold compared to the wild type variant, while it decreased sgtF4 X(-) transcription and replication by more than 3 fold. Trans-complementation of HBx was able to restore sgtF1b and sgtF4 wild type transcription and replication levels. In addition, transfection with chimeric variants, carrying the wild type or mutated X gene of one sgt in the backbone of the other sgt, suggested that the X gene sequence, that includes regulatory elements that modulate pgRNA transcription, and not HBx, was the responsible for the disparity observed between sgtF1b X(-) and sgtF4 X(-) variants. Conclusion. These results suggest that sgtF1b and sgtF4 viruses might exploit different pathways to regulate its transcription and replication, which eventually lead to a common purpose, to modulate wild type replication levels of sgtF1b and sgtF4 viruses.