PLATELET-DERIVED GROWTH-FACTOR B-CHAIN GENE-EXPRESSION IN MESANGIAL CELLS - EFFECT OF PHORBOL ESTER ON GENE-TRANSCRIPTION AND MESSENGER-RNASTABILITY

We have investigated the effect of phorbol 12-myristate 13-acetate (PM A) on platelet-derived growth factor (PDGF) B-chain gene transcription as well as on mRNA stability in cultured human mesangial cells. Addit ion 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 measurin g elongation of transcripts in isolated nuclei followed by hybridizati on of labeled RNA transcripts to a cDNA encoding for PDGF-B chain. Our data show that PMA induces PDGF-B chain gene transcription by approxi mately 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 m ethod. In human mesangial cells, the PDGF-B chain mRNA exhibited halfl ife of approximately 105 min;In the presence of PMA, the halflife of P DGF-B chain mRNA was reduced to approximately 72 min. These studies in dicate that regulation of PDGF-B chain gene by PMA in human mesangial cells involves a coordinate effort at the level of transcription and m RNA stability.