Antagonists of phosphatidylinositol 3-kinase block activation of several novel protein kinases in neutrophils.
Several novel protein kinases are known to be rapidly activated in neutrophils stimulated with the chemoattractant fMet-Leu-Phe (fMLP). These kinases include a histone H4 protein kinase and several renaturable kinases with molecular masses of about 69, 63, 49, and 40 kDa. The renaturable kinases can catalyze the phosphorylation of a peptide that corresponds to residues 297-331 of the 47-kDa subunit of the NADPH-oxidase system (p47-phox). Previous studies have indicated that the activation of all of these protein kinases involves an uncharacterized stimulatory pathway and/or novel second messenger. The studies reported herein were undertaken to determine if phosphatidylinositol 3-kinase (PI3-K) is a component of this pathway. We report that certain chromosome derivatives (e.g. 2-(4-morpholinyl)-8-phenylchromone (LY294002)) and wortmannin, which inhibit PI3-K by distinct mechanisms, blocked activation of all of these novel kinases. These antagonists also inhibited the phosphorylation of p47-phox (about 50%) and O2.- release (about 80%) in cells stimulated with fMLP, but not with 4 beta-phorbol 12-myristate 13-acetate. A strong correlation exists between the amounts of these antagonists required to produce 50% inhibition of PI3-K in vitro and O2.- release in vivo. In contrast, a single atom substitution of LY294002 produced a compound (LY303511) that did not inhibit PI3-K. Compound LY303511 did not appreciably inhibit the activation of the novel protein kinases or O2.- generation. These data strongly suggest that PI3-K is involved in the activation of several novel protein kinases in neutrophils, one or more of which may be involved in O2.- release.
Ding
J
J
Boston Biomedical Research Institute, Massachusetts 02114, USA.
Vlahos
C J
CJ
Liu
R
R
Brown
R F
RF
Badwey
J A
JA
D000730:Androstadienes / Q000494:pharmacology*; D000818:Animals; D002867:Chromones / Q000494:pharmacology*; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D006168:Guinea Pigs; D066298:In Vitro Techniques; D007700:Kinetics; D009025:Morpholines / Q000494:pharmacology*; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology*; D009247:NADH, NADPH Oxidoreductases / Q000378:metabolism; D019255:NADPH Oxidases; D009504:Neutrophils / Q000187:drug effects / Q000201:enzymology* / Q000502:physiology; D019869:Phosphatidylinositol 3-Kinases; D010766:Phosphorylation; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000037:antagonists & inhibitors*; D010879:Piperazines / Q000494:pharmacology; D006656:Protamine Kinase / Q000378:metabolism; D011489:Protein Denaturation; D047428:Protein Kinase Inhibitors; D011494:Protein Kinases / Q000378:metabolism*; D013481:Superoxides / Q000378:metabolism; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D000077191:Wortmannin
D000730:Androstadienes; D002867:Chromones; D004791:Enzyme Inhibitors; D009025:Morpholines; D010879:Piperazines; D047428:Protein Kinase Inhibitors; D013481:Superoxides; C093555:LY 303511; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; D009240:N-Formylmethionine Leucyl-Phenylalanine; D009247:NADH, NADPH Oxidoreductases; D019255:NADPH Oxidases; D011494:Protein Kinases; D019869:Phosphatidylinositol 3-Kinases; D017853:Phosphotransferases (Alcohol Group Acceptor); D006656:Protamine Kinase; D013755:Tetradecanoylphorbol Acetate; D000077191:Wortmannin