Phosphorylation of Syk activation loop tyrosines is essential Syk function: An in vivo study using a specific anti-Syk activation loop phosphotyrosine antibody

Receptors and Signal Transduction Section, Oral Infection and Immunity Branch, NIDCR, National Institutes of Health, Bethesda, Maryland 20892, USA.
Journal of Biological Chemistry (Impact Factor: 4.57). 12/2000; 275(45):35442-7. DOI: 10.1074/jbc.M004549200
Source: PubMed


Syk is an important protein-tyrosine kinase in immunoreceptor signaling. FcepsilonRI aggregation in mast cells induces tyrosine phosphorylation and increased enzymatic activity of Syk. The two adjacent tyrosines in the Syk activation loop are thought to be important for the propagation of FcepsilonRI signaling. To evaluate the phosphorylation of these tyrosines in vivo and further understand the relationship of Syk tyrosine phosphorylation with its function, an antibody was developed specific for phosphorylated tyrosines in the activation loop of Syk. FcepsilonRI aggregation on mast cells induced the phosphorylation of both tyrosine residues of the activation loop. The kinase activity of Syk played the major role in phosphorylating its activation loop tyrosines both in vivo and in vitro. In FcepsilonRI-stimulated mast cells, the total Syk tyrosine phosphorylation paralleled the phosphorylation of its activation loop tyrosines and downstream propagation of signals for histamine release. In contrast, the cell surface binding of anti-ganglioside monoclonal antibody AA4 induced only strong general tyrosine phosphorylation of Syk and minimal histamine release and weak phosphorylation of activation loop tyrosines. These results demonstrate that phosphorylation of the activation loop tyrosines is important for mediating receptor signaling and is a better marker of Syk function than is total Syk tyrosine phosphorylation.

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    • "SYK is associated with the γ chain of the FcεRI. Binding of SYK to phosphorylated γ chain ITAM through the SYK SH2 domains induces a conformational change in the kinase, leading to its increased enzymatic activity (Zhang et al., 2000; Siraganian et al., 2010). Tec family kinases represent another class of non-receptor protein tyrosine kinases that are implicated in FcεRI-mediated activation. "
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    • "FcεRI signaling relies on Lyn-dependent phosphorylation of ITAMs on the cytoplasmic portion of the β and γ receptor subunits. The protein kinase Syk is recruited to the phosphorylated ITAMs where it becomes activated and autophosphorylated (Zhang et al. 2000; de Castro et al. 2010 "
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    • "These two rules differ in whether Syk is phosphorylated at its activation loop tyrosine residues Y519 and Y520, which are treated as a single site for simplicity. Phosphorylation of the activation loop enhances the catalytic activity of Syk (48). Rate constants consistent with this regulatory mechanism are given after each rule, and are assigned values in the “parameters” block of the model specification (File S1 in Supplementary Material). "
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