Alteration of Mature Nucleocapsid and Enhancement of Covalently Closed Circular DNA Formation by Hepatitis B Virus Core Mutants Defective in Complete-Virion Formation

Assembly of hepatitis B virus (HBV) begins with packaging of the pregenomic RNA (pgRNA) into immature nucleocapsids (NC), which are converted to mature NCs containing the genomic relaxed circular (RC) DNA as a result of reverse transcription. Ma-ture NCs have two alternative fates: (i) envelopment by viral envelope proteins, leading to secretion extracellularly as virions, or (ii) disassembly (uncoating) to deliver their RC DNA content into the host cell nucleus for conversion to the covalently closed circular (CCC) DNA, the template for viral transcription. How these two alternative fates are regulated remains to be better un-derstood. The NC shell is composed of multiple copies of a single viral protein, the HBV core (HBc) protein. HBcmutations lo-cated on the surface of NC have been identified that allow NCmaturation but block its envelopment. The potential effects of some of these mutations on NC uncoating and CCCDNA formation have been analyzed by transfecting HBV replication con-structs into hepatoma cells. All envelopment-defective HBcmutations tested were competent for CCCDNA formation, indicat-ing that core functions in envelopment and uncoating/nuclear delivery of RCDNAwere genetically separable. Some of the envel-opment-defective HBcmutations were found to alter specifically the integrity of mature, but not immature, NCs such that RC DNA became susceptible to nuclease digestion. Furthermore, CCCDNA formation could be enhanced by NC surface mutations that did or did not significantly affect mature NC integrity, indicating that the NC surface residues may be closely involved in NC uncoating and/or nuclear delivery of RCDNA