AN ALTERNATIVELY SPLICED FORM OF PIT-1 REPRESSES PROLACTIN GENE-EXPRESSION

The pituitary-specific transcription factor Pit-1 is required for expr ession of the PRL gene. Transcription of the PRL gene in the anterior pituitary is both activated and repressed in response to neuroendocrin e signals. The molecular events that mediate repression are unknown. T ransplantation of GH3 pituitary tumor cells from culture to female Wis tar-Furth rats resulted in repression of PRL gene expression. When the transplanted cells were returned to culture, PRL gene expression was rapidly activated. We used this model to study potential mechanisms by which PRL gene expression was silenced. In addition to the appropriat e size Pit-1 proteins of 33 and 31 kilodaltons, smaller forms of the t ranscription factor, migrating at approximately 27 and 24 kilodaltons, were found in transplanted cells in which PRL gene expression was rep ressed. These smaller forms of Pit-1 protein were observed to disappea r when transplanted cells were returned to culture, coincident with th e activation of PRL gene expression. A transcript approximately 170 ba se pairs shorter than expected for that encoding full-length Pit-1 was detected in transplanted GH3 cell RNA by polymerase chain reaction. T he shorter Pit-1 transcript was abundant only in GH3 cells after in vi vo passage and was not readily detected in transplanted cells after as little as 12 h in culture. This shorter transcript was found to resul t from excision of sequence corresponding to exon iv and encodes a Pit -1 protein lacking 54 amino acids of the POU-specific domain. Gene tra nsfer studies demonstrated this alternative form of Pit-1 inhibited PR L promoter activity. The results implicate an alternatively spliced fo rm of Pit-1 as a potential mediator of repression of PRL gene expressi on.