Human infection with hepatitis B virus (HBV) results in a broad range of clinical symptoms, from mild, inapparent disease, fulminant hepatitis to hepatocellular carcinoma. Due to lack of an in vitro system for HBV, the events of viral life cycle and infectious process are poorly understood. Human hepatocytes are the primary site of HBV infection. The expression of the HB viral genes displays preference for liver cells. The basis of this activity is the DNA-protein interaction at the transcription control elements. In the HBV genome, the enhancer element 1 which is located strategically between nucleotides 966-1236 has been shown to be the master regulatory element with a potential to control the overall viral gene expression. Studies from several laboratories including ours, have mapped the protein binding sites within the enhancer and have demonstrated the liver cell-specificity of this element. Our interest in this study is to continue to investigate the regulatory functions of proteins that bind within the core element of the HBV enhancer. We also propose to study the promoter that lies adjacent to this enhancer, and is implicated in the biosynthesis of a HB viral regulatory protein, HBx. HBx transactivates the enhancer/X promoter activity via a c-AMP inducible motif (CRE) through CREB/ATF-2 proteins. Studies proposed here are aimed at exploring the mechanism(s) by which HBx; regulates its own biosynthesis (autoregulation). The second principal focus of this application is on the purification of a liver-enriched factor, PBF (palindrome-binding factor) which binds to a motif within the enhancer that is critical in the regulation for the overall HBV gene expression. PBF co-purified during oligonucleotide-affinity purification of HNF-3beta. Since the mutation of PBF-binding site affect enhancer function, we propose to undertake the isolation of the gene that encodes PBF.
The aim of this project is to obtain sufficient amounts of PBF for microsequencing and subsequently pursue the isolation of the gene encoding that factor. Finally, the proposed studies are aimed at understanding key DNA-protein interactions that are functionally relevant to HB viral gene expression, consequences of which manifest in liver disease pathogenesis including hepatocellular carcinoma.
| Waris, Gulam; Siddiqui, Aleem (2002) Interaction between STAT-3 and HNF-3 leads to the activation of liver-specific hepatitis B virus enhancer 1 function. J Virol 76:2721-9 |
