Platelet-derived growth factor B-chain gene expression in mesangial cells: effect of phorbol ester on gene transcription and mRNA stability.
We have investigated the effect of phorbol 12-myristate 13-acetate (PMA) on platelet-derived growth factor (PDGF) B-chain gene transcription as well as on mRNA stability in cultured human mesangial cells. Addition of actinomycin to cells stimulated with PMA decreases steady state levels of PDGF-B chain mRNA analysed by solution hybridization assay. PDGF-B chain gene transcription was also assayed directly by measuring elongation of transcripts in isolated nuclei followed by hybridization of labeled RNA transcripts to a cDNA encoding for PDGF-B chain. Our data show that PMA induces PDGF-B chain gene transcription by approximately 2-fold. alpha-Amanitin, an RNA polymerase II inhibitor, blocked transcription by more than 70%. In addition, we determined the effect of PMA on the halflife of PDGF-B chain mRNA directly by pulse chase method. In human mesangial cells, the PDGF-B chain mRNA exhibited halflife of approximately 105 min. In the presence of PMA, the halflife of PDGF-B chain mRNA was reduced to approximately 72 min. These studies indicate that regulation of PDGF-B chain gene by PMA in human mesangial cells involves a coordinate effort at the level of transcription and mRNA stability.
Bhandari
B
B
Department of Medicine, University of Texas Health Science Center at San Antonio.
Woodruff
K
K
Abboud
H E
HE
D000546:Amanitins / Q000494:pharmacology; D002478:Cells, Cultured; D003609:Dactinomycin / Q000494:pharmacology; D015870:Gene Expression; D005920:Glomerular Mesangium / Q000166:cytology / Q000378:metabolism*; D006801:Humans; D010982:Platelet-Derived Growth Factor / Q000096:biosynthesis / Q000235:genetics*; D012319:RNA Polymerase II / Q000037:antagonists & inhibitors; D012333:RNA, Messenger / Q000378:metabolism*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*; D014158:Transcription, Genetic / Q000187:drug effects*
D000546:Amanitins; D010982:Platelet-Derived Growth Factor; D012333:RNA, Messenger; D003609:Dactinomycin; D012319:RNA Polymerase II; D013755:Tetradecanoylphorbol Acetate