THE FUNCTIONAL IMPORTANCE OF A CAP SITE-PROXIMAL REGION OF THE HUMAN PROINTERLEUKIN-1-BETA GENE IS DEFINED BY VIRAL PROTEIN TRANSACTIVATION

Prointerleukin 1-beta (IL-1-beta) is a cytokine that mediates a broad range of biological activities. Genomic sequences that regulate IL-1-beta transcription include both inducible regulatory elements located more than 2,700 bp upstream of the transcriptional start site (cap site) and proximal elements located near the TATA box of this gene. In this study, we focused on the identification and characterization of trans-acting nuclear regulatory proteins that bind to the cap site-proximal region of the human IL-1-beta gene. We identified a protein, termed NFIL-1-beta-A (NF-beta-A), that binds to a highly conserved 12-bp DNA sequence (-49 to -38) located upstream of the TATA box motif in both the human and murine IL-1-beta genes. The IL-1-alpha gene, which lacks a TATA motif, does not possess an NF-beta-A-binding sequence within the promoter region, suggesting that NF-beta-A may selectively regulate IL-1-beta expression. Using electrophoretic mobility shift assays, we identified several distinct DNA-protein complexes that are expressed in a cell-type-specific manner. In monocytic cell lines, the relative abundance of these complexes varies rapidly following stimulation of the cells with phorbol esters or lipopolysaccharide. UV cross-linking analysis identified two distinct DNA-binding polypeptides that comprise distinct complexes. The functional role of NF-beta-A was assessed in transient transfection assays. These data indicate that NF-beta-A is required for both basal and inducible promoter activity in monocytic cells. Furthermore, the human cytomegalovirus immediate-early 1 gene product requires the presence of NF-beta-A in order to trans-activate the proximal IL-1-beta promoter in a monocytic cell line. We propose that NF-beta-A is a factor that mediates either direct or indirect activation by the immediate-early 1 gene product. The proximity of this essential factor to the TATA motif suggests a possible role in transcriptional initiation.