|Reference : The tilapia prolactin I gene: evolutionary conservation of the regulatory elements direc...|
|Scientific journals : Article|
|Life sciences : Biochemistry, biophysics & molecular biology|
|The tilapia prolactin I gene: evolutionary conservation of the regulatory elements directing pituitary-specific expression|
|Poncelet, A. C. [ > > ]|
|Levavi-Sivan, B. [> > > >]|
|Muller, Marc [Université de Liège - ULg > Département des sciences de la vie > GIGA-R : Biologie et génétique moléculaire >]|
|Yaron, Z. [> > > >]|
|Martial, Joseph [Université de Liège - ULg > Département des sciences de la vie > GIGA-R : Biologie et génétique moléculaire >]|
|Belayew, A. [> > > >]|
|DNA & Cell Biology|
|Mary Ann Liebert, Inc.|
|[en] Animals ; Base Sequence ; Binding Sites ; Cell Extracts ; Cells, Cultured ; Cloning, Molecular ; DNA/metabolism ; DNA-Binding Proteins/metabolism ; Gene Expression Regulation/*genetics ; Genes/genetics ; Molecular Sequence Data ; Pituitary Gland/cytology/*physiology ; Prolactin/*genetics ; Promoter Regions, Genetic/genetics ; Rats ; Regulatory Sequences, Nucleic Acid/*genetics ; Sequence Analysis, DNA ; Sequence Deletion ; Sequence Homology, Nucleic Acid ; Species Specificity ; Tilapia/*genetics ; Transcription Factor Pit-1 ; Transcription Factors/metabolism ; Transcription, Genetic/genetics|
|[en] To study the elements involved in the pituitary specific transcriptional regulation of the tilapia prolactin I gene (tiPRL I), we have cloned and entirely sequenced a 3.4-kb genomic fragment immediately upstream from the first exon. In footprinting experiments, three tilapia sequences are protected from DNase I digestion by rat pituitary extracts (base pair coordinates -643 to -593, -160 to -111, and -73 to -46). Computer analysis of the nucleotide sequence reveals significant homology to mammalian binding sites for Pit-1, a transcription factor that is known to mediate pituitary-specific expression of the PRL genes in mammals. The tiPRL I 5'-flanking sequences can direct transient expression of a linked luciferase reporter gene in transfected rat pituitary cell lines and tilapia pituitary primary cell cultures. Transient expression experiments with 5'-deletion mutants reveal three regulatory regions. Two have a stimulatory effect on transcription and one an inhibitory effect. Electrophoretic mobility-shift assays (EMSA) demonstrate that the rat Pit-1 factor specifically binds to tilapia DNA sequences. Several such tilapia Pit-1 binding sites mediate activation of a linked heterologous promoter in transfected rat and tilapia pituitary cells. As evidenced by EMSA, a Pit-1-like protein is present in tilapia pituitary extracts. All these data point to a high conservation of the molecular mechanisms involved in pituitary-specific expression of the PRL genes in vertebrates.|
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