title: Pit-1 binding sequences permit calcium regulation of human prolactin gene expression.
abstract: This study examines the regulation of the human PRL (hPRL) gene promoter by intracellular calcium. Deletants of the 5'-flanking region of the hPRL gene and constructs consisting of the thymidine kinase promoter linked to the first or second proximal Pit-1 binding site were fused to the bacterial chloramphenicol acetyl transferase (CAT) reporter gene. With the complete 5-kilobase pair (kbp) hPRL promoter sequence the calcium channel agonist Bay K8644 induced a significant 2-fold increase in CAT reporter gene expression and the antagonist verapamil a 4.5-fold reduction, using GH3 cells cultured in physiological levels of calcium. The transcriptional response to calcium influx was similar with a series of 5'-deleted hPRL-CAT constructs including those that comprised the proximal (up to 740 bp) or distal (-1300- to -1700-bp) sequences alone. When treating cells cultured in low calcium conditions the induction with the hPRL promoter increased to 5-fold on the addition of exogenous calcium and Bay K8644. The pituitary-specific expression of the hPRL gene is conferred by the interaction of the pituitary-specific factor Pit-1 with several binding sites located in the 5'-flanking DNA, of which three are located in the proximal region. This suggested that Pit-1 binding sites may be involved in the calcium response.(ABSTRACT TRUNCATED AT 250 WORDS)
authors.affiliation: Department of Medicine, University of Manchester, United Kingdom.
authors.forename: N
authors.initials: N
authors.lastname: Hoggard
authors.forename: J R
authors.initials: JR
authors.lastname: Davis
authors.forename: M
authors.initials: M
authors.lastname: Berwaer
authors.forename: P
authors.initials: P
authors.lastname: Monget
authors.forename: B
authors.initials: B
authors.lastname: Peers
authors.forename: A
authors.initials: A
authors.lastname: Belayew
authors.forename: J A
authors.initials: JA
authors.lastname: Martial
mesh_terms: D000634:Aminoquinolines; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D002118:Calcium / Q000502:physiology*; D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics / Q000378:metabolism; D002872:Chromosome Deletion; D004268:DNA-Binding Proteins / Q000378:metabolism*; D005456:Fluorescent Dyes; D005822:Genetic Vectors; D006801:Humans; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides; D010911:Pituitary Neoplasms; D011388:Prolactin / Q000235:genetics*; D011401:Promoter Regions, Genetic; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D013050:Spectrometry, Fluorescence; D013937:Thymidine Kinase / Q000235:genetics / Q000378:metabolism; D050817:Transcription Factor Pit-1; D014157:Transcription Factors / Q000378:metabolism*; D014162:Transfection
chemical_list: D000634:Aminoquinolines; D004268:DNA-Binding Proteins; D005456:Fluorescent Dyes; D009838:Oligodeoxyribonucleotides; C494187:POU1F1 protein, human; C494186:Pou1f1 protein, rat; D011993:Recombinant Fusion Proteins; D050817:Transcription Factor Pit-1; D014157:Transcription Factors; C035738:Quin2-acetoxymethyl ester; D011388:Prolactin; D015500:Chloramphenicol O-Acetyltransferase; D013937:Thymidine Kinase; D002118:Calcium