Uckun F M, Schieven G L, Dibirdik I, Chandan-Langlie M, Tuel-Ahlgren L, Ledbetter J A
Department of Therapeutic Radiology-Radiation Oncology, University of Minnesota Health Sciences Center, Minneapolis 55455.
J Biol Chem. 1991 Sep 15;266(26):17478-85.
CD40/Bp50 B-cell receptor has been implicated as having an important function for the regulation of human B-cell growth and maturation as well as antigen-driven selection of tonsillar B-cells in germinal centers. The purpose of the present study was to examine the biochemical events triggered by the engagement of the CD40 receptor in human B-lineage lymphoid cells corresponding to discrete developmental stages of human B-cell ontogeny. The engagement of the CD40 receptor on pro-B-, pre-pre-B-, pre-B-, or activated mature B-cells but not on resting mature B-cells with the agonistic anti-CD40 monoclonal antibody G28-5 resulted in enhanced tyrosine phosphorylation of four distinct phosphoproteins with molecular masses of 67, 72, 96, and 113 kDa and induced a rapid increase in the production of inositol 1,4,5-trisphosphate. Further, we have identified five electrophoretically distinct renaturable CD40-regulated serine/threonine-specific protein kinases (PK120, PK93, PK76, PK55, and PK48) that showed markedly increased in vitro activity after CD40 stimulation. Protein kinase C inhibitor 1-(5-isoquinolinylsulfonyl)-2-methylpiperazine (H7) abrogated the stimulation of the in vitro activity of PK120, PK93, PK55, and PK48 and attenuated the stimulation of the in vitro activity of PK76 in response to the engagement of the CD40 receptor but did not influence the enhanced tyrosine phosphorylation of cellular substrates after CD40 stimulation. Notably, genistein and herbimycin A, two potent inhibitors of tyrosine-specific protein kinases, not only abrogated the CD40-induced enhanced tyrosine phosphorylation on the 67-, 72-, 96-, and 113-kDa substrates, but they also inhibited the CD40-induced stimulation of phosphoinositide turnover as well as the CD40-induced increase of the in vitro activity of renaturable serine/threonine-specific protein kinases.(ABSTRACT TRUNCATED AT 400 WORDS)
CD40/Bp50 B细胞受体已被认为在调节人类B细胞生长和成熟以及生发中心扁桃体B细胞的抗原驱动选择方面具有重要功能。本研究的目的是检测与人类B细胞个体发育离散阶段相对应的人类B系淋巴细胞中，CD40受体激活所引发的生化事件。用激动性抗CD40单克隆抗体G28-5作用于前B细胞、前前B细胞、前B细胞或活化的成熟B细胞上的CD40受体，而不是静息成熟B细胞上的该受体，导致4种分子量分别为67、72、96和113 kDa的不同磷蛋白酪氨酸磷酸化增强，并诱导肌醇1,4,5-三磷酸的产生迅速增加。此外，我们鉴定出5种电泳性质不同的可复性CD40调节的丝氨酸/苏氨酸特异性蛋白激酶（PK120、PK93、PK76、PK55和PK48），它们在CD40刺激后体外活性显著增加。蛋白激酶C抑制剂1-(5-异喹啉磺酰基)-2-甲基哌嗪（H7）消除了PK120、PK93、PK55和PK48体外活性的刺激，并减弱了PK76体外活性对CD40受体激活的反应，但不影响CD40刺激后细胞底物酪氨酸磷酸化的增强。值得注意的是，酪氨酸特异性蛋白激酶的两种有效抑制剂染料木黄酮和赫司特霉素A，不仅消除了CD40诱导的67、72、96和113 kDa底物上酪氨酸磷酸化的增强，而且还抑制了CD40诱导的磷酸肌醇代谢周转刺激以及CD40诱导的可复性丝氨酸/苏氨酸特异性蛋白激酶体外活性的增加。（摘要截断于400字）
