The POU-homeodomain transcription factor Pit-1 is required for the differentiation of the anterior pituitary cells and the expression of their hormone products. Pit-1 , an alternate splicing isoform, has diametrically different outcomes when it is expressed in different cell types. Pit-1 acts as a transcriptional repressor of prolactin (PRL) and growth hormone genes in pituitary cells, and as a transcriptional activator in non-pituitary cells. In order to explore these differences, we: (1) identified the transcriptional cofactors necessary for reconstitution of repression in non-pituitary cells; (2) tested the effect of the -domain on heterodimerization with Pit-1 and physical interaction with the co-activator CREB binding protein (CBP); and (3) determined the -domain sidechain chemistry requirements for repression. Co-expression of both Pit-1 isoforms reconstituted the repression of the PRL promoter in non-pituitary cells. The -domain allowed heterodimerization with Pit-1 but blocked physical interaction with CBP, and specific chemical properties of the -domain beyond hydrophobicity were dispensable. These data strongly suggest that Pit-1 represses hormone gene expression by heterodimerizing with Pit-1 and interfering with the assembly of the Pit-1-CBP complex required for PRL promoter activity in pituitary cells.

Many transcription factors are expressed as multiple isoforms with distinct effects on the regulation of gene expression, and the functional consequences of structural differences between transcription factor isoforms may allow for precise control of gene expression. The pituitary transcription factor isoforms Pit-1 and Pit-1 differentially regulate anterior pituitary hormone gene expression. Pit-1 is required for the development of and appropriate hormone expression by anterior pituitary somatotrophs and lactotrophs. Pit-1 differs structurally from Pit-1 by the splice-insertion of the 26-residue -domain in the transactivation domain, and it differs functionally from Pit-1 in that it represses expression of the prolactin promoter in a cell-type specific manner. In order to identify signal and promoter context requirements for repression by Pit-1 , we examined its function in the presence of physiological regulatory signals as well as wild-type and mutant Pit-1-dependent target promoters. Here, we demonstrate that Pit-1 impairs recruitment of cAMP response elementbinding protein (CREB)-binding protein to the promoters that it represses. In addition, we show that repression of target promoter activity, reduction in promoter histone acetylation, and decrease of CREB-binding protein recruitment all depend on promoter context. These findings provide a mechanism for promoter-specific repression by Pit-1 .