EFFECT OF AN ECTOKINASE INHIBITOR, K252B, ON DEGRANULATION AND CA2-2H3 CELLS AND HUMAN BASOPHILS( SIGNALS OF RBL)

We examined the effects of K252b, an ectoprotein kinase inhibitor of m icrobial origin, on the activation process of RBL-2H3 cells by cross-l inking of IgE receptors by the endoplasmic reticulum Ca2+-ATPase inhib itor 2,5-di(tert-butyl)-1,4-hydroquinone or by the Ca2+ ionophore A231 87, Analysis of phosphorylation of ectoproteins following IgE receptor cross-linking revealed that K252b mainly inhibited the phosphorylatio n of a 130-kDa protein, The inhibitor simultaneously inhibited degranu lation and the sustained increase in the cytosolic calcium ion concent ration even after addition of Ag, In contrast, K252b did not inhibit t he increase in degranulation and cytosolic calcium ion concentration c aused by stimulation with 2,5-di(tert-butyl)-1,4-hydroquinone and A231 87, Permeation of K252b into RBL-2H3 cells, assessed by fluorescence i ntensity, was very low, K252b also inhibited degranulation caused by I gE receptor cross-linking in human basophils, but did not inhibit the degranulation caused by A23187, Thus, our findings suggest that the ef fects of K252b may be mediated by outer surface-bound or -anchored K25 2b-sensitive molecules on RBL-2H3 cells and human basophils, and that the phosphorylation of ectoprotein may involve a transmembrane influx of Ca2+ by IgE receptor cross-linking.