DNA elements with AT-rich core sequences direct pituitary cell-specific expression of the pro-opiomelanocortin gene in transgenic mice.

Abstract

Corticotrophs are the first fully differentiated cells to appear in the anterior pituitary during organogenesis and are distinguished by pro-opiomelanocortin (POMC) gene expression. Earlier studies in our laboratory defined three DNA regions (sites 1, 2 and 3) within promoter sequences at the 5'-end of the rat POMC gene (-323/-34) that cooperatively targeted cell-specific gene expression to corticotrophs and melanotrophs in transgenic mice. In this study we analysed the DNA-nuclear protein interactions underlying this functional activity. We demonstrated that the transcriptional activator SP1 interacts with GC-rich regions in sites 1 (-146/-136) and 2 (-201/-192) and an unidentified protein, which we call PP1 (putative pituitary POMC1), interacts with AT-rich regions in sites 2 (-202/-193) and 3 (-262/-253). The PP1-binding activity appears to be specific to cells that express the POMC gene because it was detected in nuclear extracts prepared from AtT20 corticotroph cells and mouse melanotroph tumours but not from GH4 pituitary tumour cells, HeLa cells or liver. Site-directed mutagenesis of core binding sequences demonstrated that PP1 is required for the correct cell-specific expression of the POMC gene in the pituitary gland of transgenic mice and SP1 appears to support such an expression. The best core binding sequence for PP1 is TAAT, a possible transcription factor homeodomain contact site. However, PP1 is distinct from Brn 3.0, a POU protein that also binds to site 3. We conclude that PP1 is a transcriptional activator for pituitary-specific POMC gene expression.