Teshima R, Saito Y, Ikebuchi H, Rajiva De Silva N, Morita Y, Nakanishi M, Sawada J, Kitani S
Division of Biochemistry and Immunochemistry, National Institute of Health Sciences, Setagaya-ku, Tokyo, Japan.
J Immunol. 1997 Jul 15;159(2):964-9.
We examined the effects of K252b, an ectoprotein kinase inhibitor of microbial origin, on the activation process of RBL-2H3 cells by cross-linking of IgE receptors by the endoplasmic reticulum Ca2+-ATPase inhibitor 2,5-di(tert-butyl)-1,4-hydroquinone or by the Ca2+ ionophore A23187. Analysis of phosphorylation of ectoproteins following IgE receptor cross-linking revealed that K252b mainly inhibited the phosphorylation of a 130-kDa protein. The inhibitor simultaneously inhibited degranulation and the sustained increase in the cytosolic calcium ion concentration even after addition of Ag. In contrast, K252b did not inhibit the increase in degranulation and cytosolic calcium ion concentration caused by stimulation with 2,5-di(tert-butyl)-1,4-hydroquinone and A23187. Permeation of K252b into RBL-2H3 cells, assessed by fluorescence intensity, was very low. K252b also inhibited degranulation caused by IgE receptor cross-linking in human basophils, but did not inhibit the degranulation caused by A23187. Thus, our findings suggest that the effects of K252b may be mediated by outer surface-bound or -anchored K252b-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.
我们研究了微生物来源的胞外蛋白激酶抑制剂K252b对内质网Ca2 + -ATP酶抑制剂2,5 - 二（叔丁基）-1,4 - 对苯二酚或Ca2 +离子载体A23187交联IgE受体而激活RBL - 2H3细胞过程的影响。对IgE受体交联后胞外蛋白磷酸化的分析表明，K252b主要抑制一种130 kDa蛋白的磷酸化。该抑制剂同时抑制脱颗粒以及即使添加抗原后胞质钙离子浓度的持续升高。相比之下，K252b并不抑制由2,5 - 二（叔丁基）-1,4 - 对苯二酚和A23187刺激引起的脱颗粒增加和胞质钙离子浓度升高。通过荧光强度评估，K252b进入RBL - 2H3细胞的渗透率非常低。K252b还抑制人嗜碱性粒细胞中由IgE受体交联引起的脱颗粒，但不抑制由A23187引起的脱颗粒。因此，我们的研究结果表明，K252b的作用可能由RBL - 2H3细胞和人嗜碱性粒细胞表面结合或锚定的K252b敏感分子介导，并且胞外蛋白的磷酸化可能涉及IgE受体交联导致的Ca2 +跨膜内流。
