Pituitary-specific expression and Pit-1 regulation of the rat growth hormone-releasing hormone receptor gene.
The GHRH receptor is expressed in the somatotroph cell of the anterior pituitary, where it functions to mediate GHRH-stimulated GH release. To study pituitary and somatotroph cell-specific expression of this gene, a transgenic mouse model and complementary cell culture experiments were developed. The activity of the 1.6-kb proximal rat GHRH receptor promoter was examined in vivo by generating transgenic mice with the promoter directing expression of a luciferase reporter. The promoter directs tissue-specific expression; luciferase is highly expressed in the pituitary but absent from 14 other tissues. Immunocytochemistry experiments show that transgene expression is targeted to GH-expressing somatotroph cells. The transgene is 5-fold more highly expressed in males than females, and there is an increase in transgene expression leading up to the onset of puberty. The 1.6-kb promoter was further examined in cell culture experiments, which revealed that the promoter is selectively activated in pituitary cells and that promoter-reporter expression in nonpituitary cells can be enhanced by the pituitary-specific transcription factor Pit-1. EMSAs identified 10 short regions that specifically bind Pit-1 with highly variable relative affinities. The highest affinity site was previously identified and is required for Pit-1 activation of the promoter. Four additional sites contribute to Pit-1 regulation of the promoter and are important to achieving full activation of the gene. The results show that the 1.6-kb promoter is sufficient to direct tissue- and cell-specific expression in vivo and is regulated by Pit-1.
McElvaine
Allison T
AT
Department of Biochemistry, Molecular Biology, and Cell Biology, Northwestern University, Hogan 4-112, Evanston, Illinois 60208, USA.
Korytko
Andrew I
AI
Kilen
Signe M
SM
Cuttler
Leona
L
Mayo
Kelly E
KE
D000818:Animals; D030801:Animals, Genetically Modified; D005786:Gene Expression Regulation / Q000502:physiology*; D017930:Genes, Reporter; D006367:HeLa Cells; D006801:Humans; D051379:Mice; D008822:Mice, Transgenic; D010903:Pituitary Gland, Anterior / Q000378:metabolism*; D051381:Rats; D018013:Receptors, Neuropeptide / Q000096:biosynthesis* / Q000235:genetics*; D018017:Receptors, Pituitary Hormone-Regulating Hormone / Q000096:biosynthesis* / Q000235:genetics*; D050817:Transcription Factor Pit-1 / Q000502:physiology*; D019076:Transgenes
D018013:Receptors, Neuropeptide; D018017:Receptors, Pituitary Hormone-Regulating Hormone; D050817:Transcription Factor Pit-1; C048250:somatotropin releasing hormone receptor