TH2-SPECIFIC PROTEIN DNA INTERACTIONS AT THE PROXIMAL NUCLEAR FACTOR-AT SITE CONTRIBUTE TO THE FUNCTIONAL-ACTIVITY OF THE HUMAN IL-4 PROMOTER/

IL-4 is a pleiotropic immunoregulatory cytokine secreted by. activated Th2, but not Th1, cells. The proximal IL-4 promoter contains MARE, C/ EBP, P0, octamer-like, P1, and activating protein-1 elements. The half c-Maf binding site (MARE), P0, and P1 sites were previously shown to be involved in Th2-specific transcriptional activity, Except the MARE and P1 site, the molecular basis for Th2 specificity of the P0 site ha s not been analyzed. Here, we provide the first detailed analysis of t he P0 binding factors and show that in Th2, but not in Th1, cells, NF- AT and proteins of the activating protein-1 family are involved in coo perative binding to the P0 and the adjacent octamer-like site. In the mouse Th2 D10 cells, Oct-1/Oct-2 are also found to participate in form ation of the P0-binding complexes, Mutation, deletion, and methylation interference analysis demonstrate that both the P0 and the octamer-li ke sequence are required for inducible binding. Furthermore,,ve provid e the first report of the functional relevance of each site in the hum an IL-4 promoter by mutagenesis/transfection analysis and demonstrate that the octamer-like, P0 and P1 sites are important for the biologic function of the IL-4 promoter. The MARE site, although it was shown to be critical for the function of the murine IL-4 promoter, does not ap pear essential for human IL-4 promoter activity in Jurkat T cells. The se findings suggest that besides c-Maf, another Th2-specific factor(s) may be involved in tissue-specific expression of the IL-4 gene.