THE MOUSE BONE MORPHOGENETIC PROTEIN-4 GENE - ANALYSIS OF PROMOTER UTILIZATION IN FETAL-RAT CALVARIAL OSTEOBLASTS AND REGULATION BY COUP-TFI ORPHAN RECEPTOR

Bone morphogenetic protein-4 (BMP-4) is one of a member of related pol ypeptides that are important in bone formation and other developmental processes, We isolated the BMP-4 gene from a mouse genomic library an d characterized the exon-intron structure and one of the candidate pro moters. Two alternative 5'-noncoding exons, 1A and 1B, were identified by reverse transcription polymerase chain reaction assays, Quantitati ve competitive polymerase chain reaction using Exon 1A, Exon 1B, and E xon 3 primers indicate the 1A-containing transcript is the primary BMP -4 mRNA expressed in bone cell cultures. Primer extension analysis sup ports that 1A is the major promoter utilized in bone cell cultures as well as in 9.5-day mouse embryos, 1A promoter activity indicate select ive DNA regions functional in bone cells. We found potential regulator y response regions in the IA 5'-flanking region of the BMP-4 gene for the chicken ovalbumin upstream-transcription factor I (COUP TFI). Spec ific binding to the COUP-TFI response regions in the BMP-4 1A promoter was demonstrated, By co transfection of a COUP-TFI expression plasmid with the BMP-4 1A promoter in fetal rat calvarial osteoblasts, we dem onstrated that COUP-TFI inhibits the BMP-4 promoter activity, This sug gests that COUP-TFI could act as a silencer for BMP-4 transcription in vivo.