Transcriptional regulation of the murine surfactant protein-A gene by B-myb

Surfactant protein A (SP-A) is selectively synthesized in subsets of cells lining the respiratory epithelium, where its expression is regulated by var ious transcription factors including thyroid transcription factor-1 (TTF-1) , Cell-specific transcription of the mouse SP-A promoter is mediated by bin ding of TTF-1 at four distinct cis-active sites located in the 5'-flanking region of the gene, Mutation of TTF-1-binding sites (TBE) 1, 3, and 4 in co mbination markedly decreased transcriptional activity of SP-A promoter-chlo ramphenicol acetyltransferase constructs containing SP-A gene sequences fro m -256 to +45, In contrast, the same mutations enhanced transcriptional act ivity in constructs containing additional 5' SP-A sequences from -399 to +4 5 suggesting that cis-acting elements within the region -399 to -256 influe nce effects of TTF-1 on SP-A promoter activity. A consensus Myb-binding sit e was identified within the region, located at positions -380 to -371 in th e mouse gene. Mutation of the Myb-binding site decreased activity of SP-A p romoter constructs in MLE-15 cells. MLE-15 cells, a cell line expressing SP -A mRNA, also expressed B-Myb. B-Myb bound to the MBS in the SP-A gene as a ssessed by electrophoretic mobility shift assay. While co-transfection of H eLa cells with a E-Myb expression plasmid activated the transfected SP-A pr omoter about 3-fold, co-transfection of B-myb with cyclin A and cdk-2, to e nhance phosphorylation of B-Myb, increased transcriptional activity of SP-A constructs approximately 20-fold. Taken together, the data support activat ion of SP-A gene promoter activity by E-Myb which acts at a cis-acting elem ent in the SP-A gene.