Phospholipase Cγ1 I interacts with conserved phosphotyrosyl residues in the linker region of Syk and is a substrate for Syk

Department of Microbiology, University of Washington, Seattle, USA.
Molecular and Cellular Biology (Impact Factor: 4.78). 05/1996; 16(4):1305-15. DOI: 10.1128/MCB.16.4.1305
Source: PubMed


Antigen receptor ligation on lymphocytes activates protein tyrosine kinases and phospholipase C-gamma (PLC-gamma) isoforms. Glutathione S-transferase fusion proteins containing the C-terminal Src-homology 2 [SH2(C)] domain of PLC-gamma1 bound to tyrosyl phosphorylated Syk. Syk isolated from antigen receptor-activated B cells phosphorylated PLC-gamma1 on Tyr-771 and the key regulatory residue Tyr-783 in vitro, whereas Lyn from the same B cells phosphorylated PLC-gamma1 only on Tyr-771. The ability of Syk to phosphorylate PLC-gamma1 required antigen receptor ligation, while Lyn was constitutively active. An mCD8-Syk cDNA construct could be expressed as a tyrosyl-phosphorylated chimeric protein tyrosine kinase in COS cells, was recognized by PLC-gamma1 SH2(C) in vitro, and induced tyrosyl phosphorylation of endogenous PLC-gamma1 in vivo. Substitution of Tyr-525 and Tyr-526 at the autophosphorylation site of Syk in mCD8-Syk substantially reduced the kinase activity and the binding of this variant chimera to PLC-gamma1 SH2(C) in vitro; it also failed to induce tyrosyl phosphorylation of PLC-gamma1 in vivo. In contrast, substitution of Tyr-348 and Tyr-352 in the linker region of Syk in mCD8-Syk did not affect the kinase activity of this variant chimera but almost completely eliminated its binding to PLC-gamma1 SH(C) and completely eliminated its ability to induce tyrosyl phosphorylation of PLC-gamma1 in vivo. Thus, an optimal kinase activity of Syk and an interaction between the linker region of Syk with PLC-gamma1 are required for the tyrosyl phosphorylation of PLC-gamma1.

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    • "Src and Fyn are regulated by phosphorylation on two sites: one at tyrosine 416 which is activatory, and the other at tyrosine 527 which is inhibitory (reviewed by Hunter, 1987). Similar phosphorylation sites are found in other SFK members: activation of phosphorylation are at tyrosines 394 for Lck, at tyrosines 323 and 352 for Syk, and inhibition of phosphorylation are at tyrosines 505 for Lck, and at tyrosines 525 and 526 for Syk (Chow et al., 1993; Law et al., 1996; Deckert et al., 1998; Zhang et al., 2000; Rao et al., 2001). Activating phosphorylation often results from autophosphorylation (Chiang and Sefton, 2000; reviewed by Roskoski, 2004; reviewed by Roskoski, 2005). "
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    Ageing research reviews 06/2012; 12(1):289-309. DOI:10.1016/j.arr.2012.06.003 · 4.94 Impact Factor
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    • "Three major MAP kinases, ERK1/2, p38, and JNK, were dose-dependently inhibited by 4-chlorotetrazolo[1,5-a]quinoxaline (Fig. 4B). It is generally believed that Lyn initially phosphorylates the tyrosine 346 residue (Y346) of murine Syk in antigen-stimulated mast cells (Chu et al., 1998; Law et al., 1996). Thus, we examined whether phosphorylation of Syk Y346 was inhibited by 4-chlorotetrazolo[1,5- a]quinoxaline. "
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    ABSTRACT: 4-Chlorotetrazolo[1,5-a]quinoxaline is a quinoxaline derivative. We aimed to study the effects of 4-chlorotetrazolo[1,5-a]quinoxaline on activation of mast cells in vitro and in mice. 4-Chlorotetrazolo[1,5-a]quinoxaline reversibly inhibited degranulation of mast cells in a dose-dependent manner, and also suppressed the expression and secretion of TNF-α and IL-4 in mast cells. Mechanistically, 4-chlorotetrazolo[1,5-a]quinoxaline inhibited activating phosphorylation of Syk and LAT, which are crucial for early FcεRI-mediated signaling events, as well as Akt and MAP kinases, which play essential roles in the production of various pro-inflammatory cytokines in mast cells. Notably, although 4-chlorotetrazolo[1,5-a]quinoxaline inhibited the activation of Fyn and Syk, minimal inhibition was observed in mast cells in the case of Lyn. Furthermore, consistent with its in vitro activity, 4-chlorotetrazolo[1,5-a]quinoxaline significantly suppressed mast cell-mediated passive cutaneous anaphylaxis in mice. In summary, the results from this study demonstrate that 4-chlorotetrazolo[1,5-a]quinoxaline shows an inhibitory effect on mast cells in vitro and in vivo, and that this is mediated by inhibiting the activation of Syk in mast cells. Therefore, 4-chlorotetrazolo[1,5-a]quinoxaline could be useful in the treatment of mast cell-mediated allergic diseases.
    Toxicology and Applied Pharmacology 09/2011; 257(2):235-41. DOI:10.1016/j.taap.2011.09.009 · 3.71 Impact Factor
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    • "These tyrosines have been reported to be involved in the interaction of Syk with PLCγ and Vav. Thus, in COS cells expression of Syk with both Tyr-342 and Tyr-346 mutated to Phe results in loss of interaction of Syk with PLCγ [55]. Experiments using two-hybrid system suggest that phosphorylated Tyr-342 of Syk is a binding site for VAV [56]. "
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    ABSTRACT: The protein tyrosine kinase Syk plays a critical role in FcεRI signaling in mast cells. Binding of Syk to phosphorylated immunoreceptor tyrosine-based activation motifs (p-ITAM) of the receptor subunits results in conformational changes and tyrosine phosphorylation at multiple sites that leads to activation of Syk. The phosphorylated tyrosines throughout the molecule play an important role in the regulation of Syk-mediated signaling. Reconstitution of receptor-mediated signaling in Syk(-/-) cells by wild-type Syk or mutants which have substitution of these tyrosines with phenylalanine together with in vitro assays has been useful strategies to understand the regulation and function of Syk.
    05/2011; 2011(6760):507291. DOI:10.1155/2011/507291
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