Systematic analysis of the role of CD19 cytoplasmic tyrosines in enhancement of activation in Daudi human B cells: clustering of phospholipase C and Vav and of Grb2 and Sos with different CD19 tyrosines.

Brooks SR, Li X, Volanakis EJ, Carter RH
J Immunol. 2000 164 (6): 3123-31

PMID: 10706702 · DOI:10.4049/jimmunol.164.6.3123

CD19 is a coreceptor on B cells that enhances the increase in cytoplasmic calcium and ERK2 activation when coligated with the B cell Ag receptor. Constructs containing point mutations and truncations were expressed in Daudi human B lymphoblastoid cells to systematically determine the requirement for individual CD19 cytoplasmic tyrosines in these responses. Evidence for activity was found for Y330, Y360, and Y421 as well as that previously published for Y391. Precipitates formed with phosphopeptides consisting of CD19 sequences flanking these residues were used to screen for cytoplasmic proteins that mediate signaling. Phosphopeptide Y330 precipitated Grb2 and Sos, whereas phosphopeptides Y391 and Y421 both precipitated Vav and phospholipase C-gamma2. These molecules also were found associated with native CD19. In mapping studies with altered constructs, CD19 Y330 and/or Y360 were necessary for binding Grb2 and Sos. Vav associated with CD19 constitutively in unstimulated cells by a tyrosine-independent mechanism requiring the portion of CD19 encoded by exons 9-12. After B cell Ag receptor stimulation, Vav association was tyrosine-dependent, but binding was influenced by multiple residues. However, when maximally phosphorylated by pervanadate, Y391 and, to a lesser extent, Y421 were sufficient. CD19 Y391 was also both necessary and sufficient for binding phospholipase C-gamma2. Thus, different tyrosines along the CD19 cytoplasmic domain provide scaffolding for the formation of complexes of different signaling molecules.

MeSH Terms (28)

Adaptor Proteins, Signal Transducing Adjuvants, Immunologic Antigens, CD19 B-Lymphocytes Calcium Signaling Cell Cycle Proteins Cytoplasm Exons GRB2 Adaptor Protein Humans Isoenzymes Lymphocyte Activation MAP Kinase Signaling System Mitogen-Activated Protein Kinase 1 Molecular Weight Mutagenesis, Site-Directed Peptide Mapping Phospholipase C gamma Phosphopeptides Proteins Proto-Oncogene Proteins Proto-Oncogene Proteins c-vav Receptors, Antigen, B-Cell Son of Sevenless Protein, Drosophila Tumor Cells, Cultured Type C Phospholipases Tyrosine Vanadates

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