FUNCTIONAL INTERACTION BETWEEN A RARE AND AN AP-2 BINDING-SITE IN THEREGULATION OF THE HUMAN HOX A4 GENE PROMOTER

HOX A genes are induced in a temporal fashion after retinoic acid (RA) treatment in non-N-ras-transformed PA-1 human teratcarcinoma cells. H owever, in N-ras-transformed PA-1 cells, RA-induced expression of HOX A genes is delayed. The mRNA for the transcriptional activator AP-2 is overexpressed in these ras-transformed cells, but AP-2 transcriptiona l activity is inhibited relative to non ras-transformed PA-1 cells. Co nstitutive expression of AP-2 mimics the effect of ras by transforming cells and inhibiting differentiation in culture. We analyzed 4 kb of the human HOX A4 gene promoter and identified seven putative AP-2-bind ing sites in the DNA sequence, Transcription assays with variably size d HOX A4 promoter reporter constructs revealed that a 365 bp region of the promoter, -2950 to -3315 relative to the mRNA start, controls RA responsiveness and ras-mediated inhibition of HOX A4 activity. This re gion contains an AP-2 binding site and a RARE. Elimination of the AP-2 site by site-directed mutagenesis demonstrated that the AP-2 site is involved in RA-mediated transcriptional activation of the human HOX A4 promoter in combination with the RA receptor response element (RARE). In N-ras-transformed cells, low HOX A4 promoter activity results from ras inhibition of AP-2 transactivation.