Mechanisms of transcriptional activation of the promoter of the rainbow trout prolactin gene by GHF1/Pit1 and glucocorticoid.

The transcription factor GHF1/Pit1, required for the expression of the prolactin (PRL) and other pituitary-specific genes, is highly conserved from fish to mammals but the mechanisms by which it activates transcription are poorly understood. The activity of the promoter (-627/+15 region) of the rainbow trout PRL (tPRL) gene fused to the luciferase reporter gene was studied using GHF1-expressing rat pituitary GC cells. Nuclear extracts of GC cells produced five GHF1-specific footprints in the tPRL promoter, with the position of the two most proximal ones being highly conserved in trout and mammalian GHF1-regulated genes. Deletional and mutational analyses of the tPRL promoter showed that the most proximal GHF1 site alone is sufficient to confer sub-maximal GHF1-dependent transcriptional activity and that a glucocorticoid response element-like motif mediates dexamethasone stimulation. It is suggested that GHF1 molecules bound to different sites of the tPRL promoter cannot interact simultaneously with the transcriptional apparatus. Moreover, GHF1 and the ligand-bound glucocorticoid receptor tethered to their cognate elements in the promoter could cooperate to enhance transcription by interacting simultaneously with different members of the basal transcriptional complex.