The mouse disabled 2 (mDab2) gene is a mouse homolog of the Drosophila disabled gene. It is markedly up regulated in retinoic acid (RA)-treated F9 cells, suggesting a role for mDab2 in the cell differentiation. To elucidate the molecular mechanisms that regulate RA-treated F9 cells specific expression of mDab2, we cloned and analyzed its genomic structure. The mDab2 gene spans over 55 kilobases and has 13 exons. The transcription start site, mapped by primer extension and 5'RACE, was located at 53 base pairs (bp) upstream of the most 5'-end of the published cDNA Using reporter gene transfection analysis, we found that a 1-kb mDab2 5'-flanking sequence directed a high level of promoter activity in RA-treated F9 cells but not in untreated cells. Further deletion and mutation analyses identified a direct repeat of 5'-AGG-AGGCGC-3' motif as novel positive regulatory element. Gel retardation assay showed that this element was needed to form specific DNA-protein complexes with factors present in RA-treated F9 cell extracts.