Inhibition of protein kinase C suppresses megakaryocytic differentiation and stimulates erythroid differentiation in HEL cells.
The bisindolylmaleimide, GF109203X (2-[1-(3-dimethylaminopropyl)-1H-indol-3-yl]-3-(1H-indol-3-yl)-maleimide ), a highly selective inhibitor of protein kinase C (PKC), was used to test the role of this enzyme in phorbol ester-induced megakaryocytic differentiation of HEL cells. Treatment of these cells with 10 nmol/L phorbol 12-myristate 13-acetate (PMA) for 3 days caused a complete inhibition of proliferation and a threefold increase in the surface expression of glycoprotein (GP) IIIa, a marker of megakaryocytic differentiation that forms part of the fibrinogen receptor complex, GPIIb/IIIa. A similar effect was observed with phorbol 12,13-dibutyrate, but not with the biologically inactive derivative PMA-4-O-methyl ether. The PMA-induced increase in GPIIIa expression was completely inhibited by GF109203X in a dose-dependent manner (IC50 = 0.5 mumol/L), with a maximal effect at 2.5 to 5.0 mumol/L. GF109203X also blocked the inhibitory effect of PMA on cell growth and inhibited PMA-stimulated phosphorylation of the 47-kD PKC substrate, pleckstrin. Incubation of HEL cells with 25 mumol/L hemin for 3 days caused a fourfold to fivefold increase in expression of the erythroid differentiation marker, glycophorin A. In contrast to the inhibitory effect of GF109203X on GPIIIa expression, hemin induction of glycophorin A was enhanced by this compound. Furthermore, GF109203X alone caused a dose-dependent increase in glycophorin A expression, and induced hemoglobinization. Consistent with these changes, Northern blot analysis revealed that GF109203X treatment reduced the steady-state level of GPIIb mRNA and increased those for glycophorin A and gamma-globin. These results suggest that PKC may act as a developmental switch controlling erythroid/megakaryocytic differentiation.
Department of Medicine, King's College School of Medicine and Dentistry, London, UK.
Y
Y
Hong
J F
JF
Martin
W
W
Vainchenker
J D
JD
Erusalimsky
D016590:Aphidicolin / Q000494:pharmacology; D015415:Biomarkers; D001798:Blood Proteins / Q000378:metabolism; D002453:Cell Cycle / Q000187:drug effects; D002454:Cell Differentiation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D015672:Erythroid Precursor Cells / Q000166:cytology*; D015973:Gene Expression Regulation, Leukemic / Q000187:drug effects; D005914:Globins / Q000096:biosynthesis / Q000235:genetics; D006021:Glycophorins / Q000096:biosynthesis / Q000235:genetics; D006412:Hematopoietic Stem Cells / Q000166:cytology / Q000187:drug effects*; D006427:Hemin / Q000494:pharmacology; D006801:Humans; D007211:Indoles / Q000494:pharmacology*; D004915:Leukemia, Erythroblastic, Acute / Q000473:pathology; D008301:Maleimides / Q000494:pharmacology*; D008533:Megakaryocytes / Q000166:cytology*; D009363:Neoplasm Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D015240:Phorbol 12,13-Dibutyrate / Q000494:pharmacology; D010750:Phosphoproteins; D010766:Phosphorylation; D010980:Platelet Membrane Glycoproteins / Q000096:biosynthesis / Q000235:genetics; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000502:physiology*; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D013755:Tetradecanoylphorbol Acetate / Q000031:analogs & derivatives / Q000494:pharmacology; D014407:Tumor Cells, Cultured
D015415:Biomarkers; D001798:Blood Proteins; D004791:Enzyme Inhibitors; D006021:Glycophorins; D007211:Indoles; D008301:Maleimides; D009363:Neoplasm Proteins; D010750:Phosphoproteins; D010980:Platelet Membrane Glycoproteins; C039077:platelet protein P47; D015240:Phorbol 12,13-Dibutyrate; D016590:Aphidicolin; C020922:4-O-methyl-12-O-tetradecanoylphorbol 13-acetate; D006427:Hemin; D005914:Globins; D011493:Protein Kinase C; C070515:bisindolylmaleimide I; D013755:Tetradecanoylphorbol Acetate