MODULATION OF THE STRESS-INDUCED SYNTHESIS OF STRESS PROTEINS BY A PHORBOL ESTER AND OKADAIC ACID

The expression of alpha B crystallin, hsp27, and hsp70 in C6 cells increased when the cells were exposed to arsenite (50 mu M for 1h) or heat (42 degrees C for 30 min), as detected by specific immunoassays, Western blot analysis, and Northern blot analysis. When cells were exposed to arsenite in the presence of 0.1 mu M phorbol 12-myristate 13-acetate (PMA), an activator of protein kinase C, or 0.2 mu M okadaic acid, an inhibitor of phosphoserine/phosphothreonine protein phosphatases, expression of alpha B crystallin was markedly enhanced. The induction of hsp27 and hsp70 expression was also stimulated to a considerable extent in the same cells. The stimulatory effect of PMA was further enhanced in the presence of okadaic acid, but it was strongly inhibited in the presence of 0.5 mu M staurosporine, an inhibitor of protein kinase C, PMA and okadaic acid also stimulated the response to heat stress of the expression of alpha B crystallin, but they barely stimulated the response to heat stress of hsp27. The extent of stimulation of the arsenite-induced responses by PMA and okadaic acid was greater when the concentration of arsenite (i.e. the magnitude of the stress) was relatively low (25-50 mu M). The arsenite-induced release of arachidonic acid from cells was also stimulated in the presence of PMA and/or okadaic acid, and the stimulatory effects of PMA and okadaic acid on the arsenite-induced accumulation of alpha B crystallin and hsp27 were strongly suppressed by quinacrine, an inhibitor of phospholipase A(2). These results suggest that the stimulatory effects of PMA and okadaic acid on the stress responses are caused, in part, by the increased metabolic activity of the arachidonic acid cascade, as a consequence of the activation of phospholipase A(2).