Induction of AP-1 binding to intron 1 of SP-A1 and SP-A2 is implicated in the phorbol ester inhibition of human SP-A promoter activity

A deletional analysis of the SP-A1 promoter in NCI-H441 cells was performed to identify potential cis-acting elements involved in phorbol ester-mediat ed repression of human SP-A transcription. The phorbol ester TPA reduced SP -A1 and SP-A2 promoter activity to approximately 35% to 45% compared to tha t of control cells. The inhibitory effect of TPA was significantly reduced upon removal of the region +64/+394 relative to the SP-A1 transcription sta rt site. Using NCI-H441 nuclear proteins, electromobility shift assay analy sis showed that the intron region +309/+329 of SP-A1 and the corresponding region of SP-A2 formed sequence specific DNA/protein complexes that were in duced by TPA exposure. The region +318/+324 of SP-A1 contains sequences sim ilar to a consensus AP-1 binding site, TGACTGA (TCACTGA for SP-A2), which w hen mutated (TGAGAGT) prevented the formation of the TPA-induced DNA/protei n complex. The TP-induced complex was supershifted in the presence of antib ody against the Jun family of proteins, but not the Fos family of proteins. These results suggest that the binding of AP-1 or an AP-1-like factor to t he first intron of SP-A1 and SP-A2 may be involved in the phorbol ester inh ibition of human SP-A gene expression.