CLONING AND CHARACTERIZATION OF THE 5'-FLANKING REGION OF THE MOUSE DIASTROPHIC DYSPLASIA SULFATE TRANSPORTER GENE

Dyastrophic dysplasia sulfate transporter (DTDST) plays an important r ole in proteoglycan synthesis in the extracellular matrix of bone and cartilage. Recently, we found that the mouse DTDST gene was induced in pluripotent C3H10T1/2 cells during differentiation by bone morphogene tic protein-2 (BMP-2). To clarify the transcriptional regulation of th e DTDST gene, we have cloned the 5'-flanking region of the mouse DTDST gene by the PCR based gene walking method. Sequence analysis revealed the presence of the TATA box followed by GC rich sequences containing two Sp-1 binding sites and a CBFA1 binding site. Transient transfecti on assays demonstrated that the basal transcriptional activity in oste oblastic MC3T3-E1 cells was mainly present between -309 and -275 bp up stream of the transcription start site (Segment -309/-275) which conta ined the consensus sequence for the xenobiotic-responsible element (XR E). Nuclear proteins from MC3T3-E1 cells and C3H10T1/2 cells could bin d to this short segment in vitro. BMP-2 increased the promoter activit y as well as the nuclear protein binding to the sequence in C3H10T1/2 cells. The present data suggest that the DTDST gene expression in oste oblasts and differentiating precursor cells to osteoblast/chondrocyte lineage would be mainly regulated by undetermined XRE binding transcri ption factors. (C) 1997 Academic Press.