Transcription repression of human hepatitis B virus genes by negative regulatory element-binding protein/SON

A negative regulatory element (NRE) is located immediately upstream of the upstream regulatory sequence of core promoter and second enhancer of human hepatitis B virus (HBV), NRE represses the transcription activation functio n of the upstream regulatory sequence of core promoter and the second enhan cer. In this study, we described the cloning and characterization of an NRE -binding protein (NREBP) through expression cloning. NREBP cDNA is 8266 nuc leotides in size and encodes a protein of 2386 amino acids with a predicted molecular mass of 262 kDa, Three previously described cDNAs, DBP-5, SONB, and SONA, are partial sequence and/or alternatively spliced forms of NREBP, The genomic locus of the NREBP/SON gene is composed of 13 exons and 12 int rons, The endogenous NREBP protein is localized in the nucleus of human hep atoma HuH-7 cells. Antibody against NREBP protein can specifically block th e NRE binding activity present in fractionated nuclear extracts in gel shif ting assays, indicating that NREBP is the endogenous nuclear protein that b inds to NRE sequence. By polymerase chain reaction-assisted binding site se lection assay, we determined that the consensus sequence for NREBP binding is GA(GPT)AN(C/G)(A/G)CC, Overexpression of NREBP enhances the repression o f the HBV core promoter activity via NRE. Overexpression of NREBP can also repress the transcription of HBV genes and the production of HBV virions in a transient transfection system that mimics the viral infection in vivo.