The Functional Importance of a Cap Site-Proximal Region of the Human Prointerleukin 1β Gene Is Defined by Viral Protein trans-Activation

Abstract

Prointerleukin 1β (IL-1β) is a cytokine that mediates a broad range of biological activities. Genomic sequences that regulate IL-1β 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-1β gene. We identified a protein, termed NFIL-1βA (NFβA), 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-1β genes. The IL-1α gene, which lacks a TATA motif, does not possess an NFβA-binding sequence within the promoter region, suggesting that NFβA may selectively regulate IL-1β expression. Using electrophoretic mobility shift assays, we identified several distinct DNAprotein 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 NFβA was assessed in transient transfection assays. These data indicate that NFβA 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 NFβA in order to trans-activate the proximal IL-1β promoter in a monocytic cell line. We propose that NFβA 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.