Alteration of B-cell antigen receptor signaling by CD19 co-ligation. A study with bispecific antibodies.
The activation of B-cell antigen receptor-associated protein tyrosine kinases is an early and crucial event in B-cell signaling. Apart from the B-cell antigen receptor (BCR), the B-cell-specific transmembrane glycoprotein CD19 has also been shown to directly activate intracellular signaling cascades. In addition, because CD19 and the BCR are associated on the surface of activated B-cells, it has been proposed that close approximation between these two entities is crucial for optimal B-cell triggering. To test this hypothesis, bispecific antibodies were generated that bind membrane IgM and CD19 simultaneously. Although CD19 bispecific antibodies strongly induced tyrosine phosphorylation, they were, in contrast to muF(ab)2 fragments, unable to induce a proliferative response. Detailed analysis of the early signaling events showed that compared with muF(ab)2 fragments CD19 bispecific antibodies potently raised the intracellular [Ca2+], which was correlated with an efficient tyrosine phosphorylation of syk. Strikingly, the assembly of Grb2 complexes that may couple the BCR to p21(ras) was clearly altered by the CD19 bispecific antibody. In addition to the reported Shc and 145-kDa phosphoproteins, a prominent 90-95-kDa phosphoprotein resembling CD19 was detected in the Grb2 complexes. Thus, studies with CD19 bispecific antibodies show that CD19 co-ligation both quantitatively and qualitatively alters BCR signaling.
Lankester
A C
AC
Central Laboratory of the Netherlands Red Cross Blood Transfusion Service and Laboratory for Experimental and Clinical Immunology, University of Amsterdam, 1066 CX Amsterdam, The Netherlands.
Rood
P M
PM
van Schijndel
G M
GM
Hooibrink
B
B
Verhoeven
A J
AJ
van Lier
R A
RA
D048868:Adaptor Proteins, Signal Transducing; D018033:Antibodies, Bispecific / Q000378:metabolism*; D018941:Antigens, CD19 / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism; D002455:Cell Division / Q000187:drug effects; D002460:Cell Line; D004591:Electrophoresis, Polyacrylamide Gel; D004789:Enzyme Activation; D004792:Enzyme Precursors / Q000378:metabolism; D051380:GRB2 Adaptor Protein; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D010766:Phosphorylation; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D011506:Proteins / Q000378:metabolism; D011947:Receptors, Antigen, B-Cell / Q000378:metabolism*; D015398:Signal Transduction; D013379:Substrate Specificity; D000072377:Syk Kinase
D048868:Adaptor Proteins, Signal Transducing; D018033:Antibodies, Bispecific; D018941:Antigens, CD19; D004792:Enzyme Precursors; D051380:GRB2 Adaptor Protein; C495948:GRB2 protein, human; D047908:Intracellular Signaling Peptides and Proteins; D011506:Proteins; D011947:Receptors, Antigen, B-Cell; D011505:Protein-Tyrosine Kinases; C000606896:SYK protein, human; D000072377:Syk Kinase; D002118:Calcium