THE TRANSCRIPTION-FACTOR-SPL REGULATES THE MYELOID-SPECIFIC EXPRESSION OF THE HUMAN HEMATOPOIETIC-CELL KINASE (HCK) GENE THROUGH BINDING TO2 ADJACENT GC BOXES WITHIN THE HCK PROMOTER-PROXIMAL REGION

The human hemopoietic cell kinase (HCK) is a member of the src family of protein tyrosine kinases specifically expressed in myeloid cells an d to a minor extent in B-lymphoid cells. HCK expression is up-regulate d at the transcriptional level during myeloid differentiation of hemat opoietic cells. To elucidate the molecular basis of the differential H CK: gene expression, the genomic region containing the HCK promoter wa s isolated and functionally characterized. A DNA fragment containing 1 01 base pairs of the 5'-flanking sequence showed strong promoter activ ity in the macrophage cell line RAW264 but was inactive in the non-mon ocytic cell lines HUT-78 and NIH-3T3. Site-directed mutagenesis of the proximal promoter region showed that two GC-rich sequence elements ar e essential for transcriptional activity in myeloid cells. Electrophor etic mobility shift analysis using nuclear extracts obtained from RAW2 64 cells and from the promonocytic cell line U-937 revealed the format ion of at least three distinct protein-DNA complexes at each of these sites, one of which was found to contain the transcription factor Spl. Expression of a reporter gene linked to the -101 HCK promoter region was up-regulated by Spl, but not by other members of the Spl family of transcription factors, in Drosophila Schneider cells. A synergistic e ffect on HCK promoter activity was observed at high concentrations of Spl. Our results show that Spl plays an essential role in the regulati on of the differential gene expression of the HCK gene.