A natural hepatitis B virus (HBV) variant associated with seroconversion from HBeAg to anti-HBe antibody contains two nucleotide substitutions (A1764T and G1766A) in the proximal nuclear hormone receptor binding site in the nucleocapsid promoter. These nucleotide substitutions prevent the binding of the retinoid X receptor ␣ (RXR␣)-peroxisome proliferator-activated receptor ␣ (PPAR␣) heterodimer without greatly altering the efficiency of binding of hepatocyte nuclear factor 4 (HNF4) to this recognition sequence. In addition, these nucleotide substitutions create a new binding site for HNF1. Analysis of HBV transcription and replication in nonhepatoma cells indicates that RXR␣-PPAR␣ heterodimers support higher levels of pregenomic RNA transcription from the wild-type than from the variant nucleocapsid promoter, producing higher levels of wildtype than of variant replication intermediates. In contrast, HNF4 supports higher levels of pregenomic RNA transcription from the variant than from the wild-type nucleocapsid promoter, producing higher levels of variant than of wild-type replication intermediates. HNF1 can support variant virus replication at a low level but is unable to support replication of the wild-type HBV genome. These observations indicate that the replication of wild-type and variant viruses can be differentially regulated by the liver-specific transcription factors that bind to the proximal nuclear hormone receptor binding site of the nucleocapsid promoter. Differential regulation of viral replication may be important in the selection of specific viral variants as a result of an antiviral immune response.The hepatitis B virus (HBV) genome is a partially doublestranded 3.2-kb DNA molecule (14, 30). The unusual structure of the viral genome reflects the replication cycle of the hepadnaviruses (30, 54). Nuclear covalently closed circular 3.2-kbp HBV DNA is transcribed to produce a greater-than-genomelength pregenomic 3.5-kb RNA that is reverse transcribed by the HBV polymerase to generate encapsidated viral genomic DNA (14,30,54). The core polypeptide and the viral polymerase are both encoded by the HBV 3.5-kb transcripts (36). Therefore, transcriptional regulation of the synthesis of the HBV 3.5-kb RNAs also controls the level of viral DNA synthesis.The level of 3.5-kb HBV RNA synthesis is determined by the rate of transcription from the nucleocapsid promoter. The regulatory elements controlling transcription from the nucleocapsid promoter have been extensively characterized (17, 23, 26-28, 39, 58-61). Recently it was shown that nuclear hormone receptors are a major determinant in regulating HBV 3.5-kb RNA synthesis and viral replication (52). The most important recognition element involved in controlling viral pregenomic RNA synthesis and viral replication is the proximal nuclear hormone receptor binding site in the nucleocapsid promoter (52).Viral variants containing two nucleotide substitutions (A1764T and G1766A) in the proximal nuclear hormone receptor binding site in the nucleocapsid promoter ar...
