REDUCED ANTIGEN PRODUCTION BY HEPATITIS-B VIRUS HARBORING NUCLEOTIDE DELETIONS IN THE OVERLAPPING X-GENE AND PRECORE-CORE PROMOTER

Hepatitis B virus (HBV) genomes with deletions in the precore-core (pr eC-C) promoter have been detected in HBV infections without serologica l markers, To address whether the mutations are responsible for the re duced production of virus antigens, either an 8 bp (8d, position 1763 to 1770) or a 20 bp (20d, 1753 to 1772) deletion was created in a wild -type (wt) HBV clone, Both mutations cause premature termination of th e overlapping X ORF, When introduced into HepG2 cells, both mutants pr oduced reduced amounts of HBsAg, HBcAg and HBeAg, but released the sam e or more virion-associated DNA compared with the wt, A co-transfectio n of the 20d mutant with a small amount of intact X gene resulted in a 3-fold increase of HBcAg production compared to transfection with eit her the 20d or wt alone. When the promoter region was cloned into CAT plasmids, the 8d preC promoter showed weak activity and its initiation site was shifted 6 to 10 bp downstream. The preC promoter activity of 20d was not detectable by CAT ELISA and 5' RACE, The levels of C tran scripts of both mutants were higher than that of the wt, and their sta rt sites were not altered, Therefore, the deletions cause the reductio n of HBsAg, HBcAg and HBeAg although the mutant viruses can still repl icate in cultured cells. The reduction of HBeAg is due to both the red uced preC promoter activity and the defect in HBx. The reduction of HB cAg is due to the disrupted X gene, despite augmented C promoter activ ity.