Functional Interaction Between a RARE and an AP-2 Binding Site in the Regulation of the Human HOX A4 Gene Promoter

Abstract

HOX A genes are induced in a temporal fashion after retinoic acid (RA) treatment in non-N-ras-transformed PA-1 human teratcarcinoma cells. However, 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 transcriptional activity is inhibited relative to non ras-transformed PA-1 cells. Constitutive 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-binding sites in the DNA sequence. Transcription assays with variably sized 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 region 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.

Evolutionary Conservation of Regulatory Elements in Vertebrate Hox Gene Clusters

“…Upstream of the HoxA4 gene we identified a stretch 154 bp that has a similarity of 85% containing a RARE element (17 bp) that is part of the HoxA4 promoter, described by Doerksen et al (1996). In the intron of gene HoxA4 a 68 bp long stretch was found containing the previously described HB1 element (Haerry and Gehring 1996).…”

CREG1 enhances p16INK4a-induced cellular senescence

“…The cyclin A2 promoter reporter was generated by cloning of a 347 bp fragment (-75 to +272) of cyclin A2 promoter into the KpnI and HindIII restriction sites of the pGL3-basic vector. 43,44 U2OS cells (40,000) were transfected with 2 μg of each expression vector in a six-well cultured plate in triplicate. Selecting media was changed at 48 hours after transfection.…”

Skeletal and pigment cell defects in the lockjaw mutant reveal multiple roles for zebrafish tfap2a in neural crest development

“…Given the restricted defects in the pharyngeal arches of low and Tfap2a mutant mice, we predict that Tfap2a and other AP-2 genes have a general role in regulating Hox genes in the NCC. Indeed, AP-2 binding motifs have been described in the promotor of mouse HoxA4 and shown to be necessary for transcription in cell lines (Doerksen et al, 1996). In mouse, expression of Tfap2b and Tfap2g has been described in the NCC and may have redundant roles with Tfap2a in patterning NCC (Chazaud et al, 1996;Moser et al, 1997).…”