Regulation of prolactin gene expression by basic fibroblast growth factor
Date of Completion
January 1998
Keywords
Biology, Molecular|Biology, Cell|Chemistry, Biochemistry
Degree
Ph.D.
Abstract
Expression of the prolactin (PRL) gene is regulated by multiple extracellular factors in the pituitary. These studies examined the molecular mechanisms by which fibroblast growth factor regulates PRL gene expression, using the rat somatolactotropic GH3 cell line, as a model system. Treatment of GH3 cells for 1 day with bFGF (1-5 ng/ml) increased mRNA levels for PRL, but had no effect on growth hormone (GH). Transient transfection assays with a PRL promoter-reporter construct indicated that bFGF increases PRL gene transcription. Basic FGF increased PRL mRNA after a delay of several hours, and this increase was dependent on new protein synthesis. Inhibitor studies revealed that neither protein kinase C nor phosphoinositide 3-kinase is required for the ability of bFGF to increase PRL gene expression. However, PP1, which is an inhibitor of src-related protein tyrosine kinases (SR-PTKs), blocked bFGF actions on PRL. Utilizing a targeted differential display reverse-transcriptase polymerase chain reaction (RT-PCR) approach, we observed that the SR-PTK, Lck, is expressed in GH$\sb3$ cells. Further RT-PCR analysis, using Lck-specific primers, revealed Lck expression in both GH$\sb3$ cells and rat pituitary. Analysis of Lck gene expression by ribonuclease protection assay further confirmed Lck expression in GH$\sb3$ cells, and indicated that Lck mRNA levels are increased by bFGF by 2- to 3-fold. Immunoblot analysis revealed the expected 56 kDa form of Lck protein in lysates from GH$\sb3$ cells and rat pituitary. Additionally, immunoblot analysis detected a smaller form of about 45 kDa, which may represent a pituitary-specific isoform of Lck. Finally, chronic treatment of GH$\sb3$ cells induced lactotrope differentiation, as evidenced by an increase in PRL and a repression of GH expression. These studies indicate that bFGF actions on PRL gene transcription are mediated by a SR-PTK, which may be Lck. Further, these studies represent the first demonstration of expression of Lck, which is thought to be a lymphocyte-specific SR-PTK, in the pituitary. ^
Recommended Citation
Hu, Meimei, "Regulation of prolactin gene expression by basic fibroblast growth factor" (1998). Doctoral Dissertations. AAI9832011.
https://digitalcommons.lib.uconn.edu/dissertations/AAI9832011