Phorbol ester inhibits granulocyte-macrophage colony-stimulating factor binding and tyrosine phosphorylation.
Exposure of human polymorphonuclear neutrophils to phorbol 12-myristate 13-acetate (PMA) results in a 70-75% reduction in the specific binding of 125I-granulocyte-macrophage colony-stimulating factor (GM-CSF) to its receptors. The PMA-induced reduction in 125I-GM-CSF binding is due to a decrease in the number of available GM-CSF receptors, as derived from Scatchard analysis of the binding data. On the other hand, the phorbol ester 4-alpha-phorbol 12,13-didecanoate (4 alpha-PDD) fails to affect 125I-GM-CSF binding. PMA promotes phosphorylation on tyrosine residues of several proteins, as demonstrated by Western blotting analysis using antiphosphotyrosine antibodies. The molecular masses of those proteins are 41, 55, 66, 78, 85, 104, and 115 kDa. GM-CSF increases the levels of the tyrosine phosphorylation of several proteins, the majority of which have similar Mr to those found in PMA-stimulated neutrophils. This increase, on all but the 41-kDa protein, is partially prevented by treatment of the cells with PMA. The inhibition by PMA of GM-CSF binding to its receptors and its phosphorylated effects is partially prevented by the protein kinase C inhibitor 1-(5-isoquinolinylsulfonyl)-2-methylpiperazine and, to a greater extent, by staurosporine. It is suggested that PMA, through the activation of protein kinase C, interrupts the excitation-response sequence initiated by GM-CSF, which includes tyrosine phosphorylation, and that the earliest altered step is the binding of GM-CSF to its receptor.
Gomez-Cambronero
J
J
Department of Physiology, University of Connecticut Health Center, Farmington 06030.
Huang
C K
CK
Yamazaki
M
M
Wang
E
E
Molski
T F
TF
Becker
E L
EL
Sha'afi
R I
RI
D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; D000470:Alkaloids / Q000494:pharmacology; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000378:metabolism*; D006801:Humans; D007546:Isoquinolines / Q000494:pharmacology; D009504:Neutrophils / Q000378:metabolism; D010766:Phosphorylation; D010879:Piperazines / Q000494:pharmacology; D011493:Protein Kinase C / Q000037:antagonists & inhibitors; D011506:Proteins / Q000378:metabolism; D019311:Staurosporine; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*; D014443:Tyrosine / Q000378:metabolism*
D000470:Alkaloids; D007546:Isoquinolines; D010879:Piperazines; D011506:Proteins; D014443:Tyrosine; D016178:Granulocyte-Macrophage Colony-Stimulating Factor; D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; D011493:Protein Kinase C; D019311:Staurosporine; D013755:Tetradecanoylphorbol Acetate