Four Tyrosine Residues in Phospholipase C- 2, Identified as Btk-dependent Phosphorylation Sites, Are Required for B Cell Antigen Receptor-coupled Calcium Signaling

Osaka University, Suika, Ōsaka, Japan
Journal of Biological Chemistry (Impact Factor: 4.57). 11/2001; 276(42):38595-601. DOI: 10.1074/jbc.M103675200
Source: PubMed


Activation of phospholipase C-γ2 (PLCγ2) is the critical step in B cell antigen receptor (BCR)-coupled calcium signaling.
Although genetic dissection experiments on B cells have demonstrated that Bruton's tyrosine kinase (Btk) and Syk are required
for activating PLCγ2, the exact activation mechanism of PLCγ2 by these kinases has not been established. We identify the tyrosine
residues 753, 759, 1197, and 1217 in rat PLCγ2 as Btk-dependent phosphorylation sites by using an in vitro kinase assay. To evaluate the role of these tyrosine residues in phosphorylation-dependent activation of PLCγ2, PLCγ2-deficient
DT40 cells were reconstituted with a series of mutant PLCγ2s in which the phenylalanine was substituted for tyrosine. Substitution
of all four tyrosine residues almost completely eliminated the BCR-induced PLCγ2 phosphorylation, indicating that these residues
include the major phosphorylation sites upon BCR engagement. Cells expressing PLCγ2 with a single substitution exhibited some
extent of reduction in calcium mobilization, whereas those expressing quadruple mutant PLCγ2 showed greatly reduced calcium
response. These findings indicate that the phosphorylations of the tyrosine residues 753, 759, 1197, and 1217, which have
been identified as Btk-dependent phosphorylation sites in vitro, coordinately contribute to BCR-induced activation of PLCγ2.

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