Stimulation of protein tyrosine phosphorylation, phosphoinositide turnover, and multiple previously unidentified serine/threonine-specific protein kinases by the Pan-B-cell receptor CD40/Bp50 at discrete developmental stages of human B-cell ontogeny.
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)
Uckun
F M
FM
Department of Therapeutic Radiology-Radiation Oncology, University of Minnesota Health Sciences Center, Minneapolis 55455.
Schieven
G L
GL
Dibirdik
I
I
Chandan-Langlie
M
M
Tuel-Ahlgren
L
L
Ledbetter
J A
JA
D015703:Antigens, CD / Q000378:metabolism*; D000944:Antigens, Differentiation, B-Lymphocyte / Q000378:metabolism*; D001402:B-Lymphocytes / Q000166:cytology / Q000378:metabolism*; D019013:CD40 Antigens; D002455:Cell Division; D002478:Cells, Cultured; D006801:Humans; D010716:Phosphatidylinositols / Q000378:metabolism*; D010766:Phosphorylation; D011494:Protein Kinases / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases; D015398:Signal Transduction
D015703:Antigens, CD; D000944:Antigens, Differentiation, B-Lymphocyte; D019013:CD40 Antigens; D010716:Phosphatidylinositols; D011494:Protein Kinases; D017346:Protein Serine-Threonine Kinases