RESPIRATORY BURST AND TYROSINE PHOSPHORYLATION BY VANADATE

We studied involvement of tyrosine-phosphorylated proteins in activati on of NADPH oxidase in guinea pig neutrophils, Pervanadate, which is t he oxidized form of orthovanadate, induced O-2(-) production and prote in tyrosine phosphorylation in neutrophils. O-2(-) production induced by pervanadate was more sensitive to the tyrosine kinase-specific inhi bitor, ST-638, as compared with the production induced by PMA, On the other hand, staurosporine more selectively inhibited PMA-induced O-2(- ) production than pervanadate-induced production. These results indica te that tyrosine kinase, not protein kinase C, is involved in pervanad ate-induced O-2(-) production. The tyrosine-phosphorylated proteins we re detected in both the cytosol and membrane fractions prepared from p ervanadate-induced neutrophils, In order to examine if tyrosine residu es of some components of NADPH oxidase were directly phosphorylated, t yrosine-phosphorylated proteins were removed from solubilized membrane s prepared from the pervanadate-stimulated neutrophils by immunoprecip itation with an anti-phosphotyrosine antibody. NADPH oxidase activity in the solubilized membranes was not decreased by the treatment. These findings suggest that the components of NADPH oxidase are not tyrosin e-phosphorylated by pervanadate treatment, that tyrosine phosphorylati on may be involved in the signal transduction pathway of NADPH oxidase activation by pervanadate, and that this pathway is independent of th e activation by protein kinase C. (C) 1995 Academic Press, Inc.