Characterization of the human IRF-3 promoter and its regulation by the transcription factor E2F1

Interferon regulatory factor 3 (IRF-3), an essential transcriptional regulator of the interferon genes, has been implicated in virus and double-stranded RNA mediated induction of IFN-α, IFN-β and RANTES, in virus-induced apoptosis and in tumor suppression. Promoter plays an important role in the regulation of gene expression, but the characterization of the human IRF-3 promoter has not been systematically analyzed in HEK 293 cells. To characterize the human IRF-3 promoter, we have isolated a genomic clone of the human IRF-3 gene promoter region containing 1,000 nucleotides of the 5'- flanking region. Transient transfection of 5'-deleted promoter-reporter constructs and luciferase assay illustrated the region -149/-93 relative to the transcription start site (TSS) is sufficient for full promoter activity. This region contains HSF, E2F, CdxA and c-Myb transcription factor binding sites. The E2F sites are highly conserved among IRF-3 promoter regions of mouse, rat and human. Therefore, it was suggested that this E2F site may be essential for basal promoter activity. Surprisingly, mutation of this E2F site increased the promoter activity by 2-fold. Furthermore, overexpression of E2F1 reduced the transcription activity by 80%. These results indicated that human IRF-3 gene core promoter was located within the region -149/-93 relative to the TSS. E2F1 transcription factor negatively regulates human IRF-3 gene promoter.