SP-A is the major protein of pulmonary surfactant and is encoded by two very similar genes, SP-Al and SP-A2, Human SP-Al and SP-A2 mRNA levels are decreased in the presence of the synthetic glucocorticoid, dexamethasone (dex), in both fetal lung expiants and the lung adenocarcinoma cell line NH441. This inhibition of SP-A gene expression occurs through both transcriptional and post-transcriptional mechanisms. To better define mechanisms involved in the transcriptional inhibition of SP-Al by glucocorticoids, we transiently transfected a number of SP-Al 5'flanking/CAT reporter constructs into NH441 cells and exposed the transfected ceils to 100 nM dex for IShrs. The constructs tested contained the SP-Al 5' flanking regions -1500/4-370, -300/4-370, -57/4-370, or -300/+22 relative to the first SP-Al transcriptional start site. DPX significantly decreased (p<0.01) CAT levels for all SP-Al 51 flanking constructs but not for a CAT control vector driven by the SV40 promoter. These results indicate the presence of a negative glucocorticoid response element within the region -57/4-22. Electromobility shift assays performed using the probe-57/4-22 showed the formation of a DNA/protein complex in the presence of nuclear extracts from NH441 cells exposed to lOOnM dex for I8hrs but not from untreated NH441 cells. Interestingly, the dex induced complex was not supershifted in the presence of antibody specific for the glucocorticoid receptor (GR), indicating that proteins other than GR are involved in the dex induced complex formation. Funded by: NIH HL49823.
|Original language||English (US)|
|State||Published - Dec 1 1998|
All Science Journal Classification (ASJC) codes
- Molecular Biology