Okadaic acid induces both augmentation and inhibition of opsonized zymosan-stimulated superoxide production by differentiated HL-60 cells. Possible involvement of dephosphorylation of a cytosolic 21K protein in respiratory burst

Abstract

We found that okadaic acid (OA), a potent tumor promotor and a phosphatase inhibitor, has a unique opposing effect on opsonized zymosan (Op.-zym.)-elicited O 2 . − production by differentiated HL-60 cells in a narrow range of concentration but does not induce any O 2 . − production by itself. Okadaic acid magnified the O 2 . − production 2.5-fold at 1.0 μM, while it inhibited it at 2.0 μM or higher concentrations. This effect of OA did not correspond to the changes in the expression of surface receptors (CD11b/CD18, CR3) for Op.-zym., because they were weakly down-regulated by OA at any concentration. Two-dimensional gel electrophoresis revealed that in the absence of OA, Op.-zym. induced rapid dephosphorylation of a cytosolic 21K protein with a very slight increase in phosphorylation of membranous p47 phox , which is one of the cytosolic factors required for respiratory burst. In the presence of a stimulatory concentration (1.0 μM) of OA, the Op.-zym.-caused dephosphorylation of the 21K protein was still observed and the phosphorylation of p47 phox was enhanced. In the presence of an inhibitory concentration (2.0 or 5.0 μM) of OA, the Op.-zym.-induced dephosphorylation of the 21K protein was strongly inhibited while p47 phox was heavily phosphorylated. Acid hydrolysis of the 21K phosphoprotein yielded only phosphoserine as a phosphoamino acid. Furthermore, at least part of the 21K protein seemed to be associated with p67 phox and p47 phox , because it was co-immunoprecipitated with those cytosolic factors. These results suggest that a cytosolic 21K protein plays an important role in respiratory burst through dephosphorylation by a phosphoserine phosphatase, and that the dephosphorylated 21K protein may work synergistically with the phosphorylated p47 phox on the pathway for activation of the respiratory burst oxidase.