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Results for “Syk Kinase”

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PubMed2004

Zeta-associated protein of 70 kDa (ZAP-70), but not Syk, tyrosine kinase can mediate apoptosis of T cells through the Fas/Fas ligand, caspase-8 and caspase-3 pathways.

The TCR zeta-chain-associated protein of 70 kDA (ZAP-70) and Syk tyrosine kinases play critical roles in regulating TCR-mediated signal transduction. They not only share some overlapped functions but also may play unique roles in regulating the function and development of T cells. However, it is not known whether they have different effects on the activation and activation-induced cell death of T cells. To address this question, we generated cDNAs encoding chimeric molecules that a tailless TCR zeta-chain was directly linked to truncated ZAP-70 (Z/ZAP) or Syk (Z/Syk) molecules lacking the two Src homology 2 domains. Transfection of these molecules into zeta-chain-deficient cells restored their TCR expression. In addition, Z/ZAP and Z/Syk transfectants but not control cells demonstrated kinase activities in phosphorylating an exogenous substrate specific for ZAP-70 and Syk kinases. Z/ZAP transfectants activated through TCRs underwent a faster time course of apoptosis and had a greater percentage of apoptotic cells than that of Z/Syk and control cells. Activated Z/ZAP transfectants increased Fas and Fas ligand (FasL) expression 3- and 40-fold, respectively. Blocking of the Fas/FasL interaction could inhibit the apoptosis of Z/ZAP transfectants. In contrast, although activated Z/Syk transfectants could increase FasL expression, their Fas expression actually decreased and the percentage of apoptotic cells did not increase. Further studies of the mechanisms revealed that activation of Z/ZAP but not Z/Syk transfectants resulted in rapid activation of caspase-3 and caspase-8 that could also be inhibited by blocking Fas/FasL interaction. These results demonstrated that ZAP-70 and Syk play distinct roles in T cell activation and activation-induced cell death.

Animals↗

PubMed1996

p120cbl is a major substrate of tyrosine phosphorylation upon B cell antigen receptor stimulation and interacts in vivo with Fyn and Syk tyrosine kinases, Grb2 and Shc adaptors, and the p85 subunit of phosphatidylinositol 3-kinase.

We and others have demonstrated that the c-cbl proto-oncogene product is one of the earliest targets of tyrosine phosphorylation upon T cell receptor stimulation. Given the similarities in the B and T lymphocyte antigen receptors, and the induction of pre-B leukemias in mice by the v-cbl oncogene, we examined the potential involvement of Cbl in B cell receptor signaling. We demonstrate prominent and early tyrosine phosphorylation of Cbl upon stimulation of human B cell lines through surface IgM. Cbl was associated in vivo with Fyn and, to a lesser extent, other Src family kinases. B cell activation also induced a prominent association of Cbl with Syk tyrosine kinase. A substantial fraction of Cbl was constitutively associated with Grb2 and this interaction was mediated by Grb2 SH3 domains. Tyrosine-phosphorylated Shc, which prominently associated with Grb2, was detected in association with Cbl in activated B cells. Thus, Grb2 and Shc adaptors, which associate with immunoreceptor tyrosine based activation motifs, may link Cbl to the B cell receptor. B cell activation also induced a prominent association between Cbl and the p85 subunit of phosphatidylinositol (PI) 3-kinase resulting in the association of a substantial fraction of PI 3-kinase activity with Cbl. Thus, Cbl is likely to play an important role to couple the B cell receptor to the PI 3-kinase pathway. Our results strongly suggest a role for p120cbl in signaling downstream of the B cell receptor and support the idea that Cbl participates in a general signal transduction function downstream of the immune cell surface receptors.

Adaptor Proteins, Signal Transducing↗

PubMed2005

Syk tyrosine kinase expression during multistep mammary carcinogenesis.

AIM: To analyze the expression of Syk tyrosine kinase, recently implicated as a tumor suppressor in mammary carcinogenesis, during the multi step development of human breast carcinoma. METHODS: Syk expression was examined in invasive carcinomas of the breast and in corresponding premalignant epithelial lesions in 50 women, using immunohistochemical method and semi-quantitative scoring system (H-score). The results were correlated with the expression of Syk in the lymph node metastases. RESULTS: Syk was strongly and uniformly expressed in normal mammary epithelium (H score=2.36-/+0.75), and no significant reduction in Syk expression was observed in hyperplastic lesions (H=2.31-/+0.87), carcinoma in situ (H=1.90-/+0.93), or invasive carcinomas (H=1.83-/+0.88). Loss of Syk expression (defined as H< or =1.0) was seen in approximately 1/5 of invasive breast carcinomas, but the majority of metastatic carcinomas (15/21) still strongly expressed Syk, irrespective of Syk expression in primary tumor. CONCLUSION: The loss of Syk characterizes a subset of breast carcinomas but does not apparently contribute to the development of the metastatic potential of these carcinomas, limiting the potential diagnostic utility of immunohistochemical test in the prediction of malignant behavior.

Breast↗

PubMed2001

Exposure of RBL-2H3 mast cells to Ag(+) induces cell degranulation and mediator release.

There is a growing need to understand the impact of environmental sulfhydryl group-reactive heavy metals on the immune system. Here we show that Ag(+) induces mast cell degranulation, as does the aggregation of the high affinity immunoglobulin E receptor (FcepsilonRI). Micromolar quantities of Ag(+) specifically induced degranulation of mast cell model rat basophilic leukemia (RBL-2H3) cells without showing cytotoxicity. The Ag(+)-mediated degranulation could be observed as rapidly as 5 min after the addition of the ions. Ag(+) also induced a rapid change in tyrosine phosphorylation of multiple cellular proteins including the focal adhesion kinase but not Syk kinase. The Syk-selective inhibitor piceatannol and the Src family-selective tyrosine kinase inhibitor PP1 dose-dependently inhibited FcepsilonRI-mediated degranulation, whereas neither compound inhibited the Ag(+)-mediated degranulation. Furthermore, likewise FcepsilonRI aggregation, Ag(+) also induced leukotriene secretion. These results show that Ag(+) activates RBL-2H3 mast cells through a tyrosine phosphorylation-linked mechanism, which is distinct from that involved in FcepsilonRI-mediated activation.

Animals↗

PubMed1997

Functional role for Syk tyrosine kinase in natural killer cell-mediated natural cytotoxicity.

Natural killer (NK) cells are named based on their natural cytotoxic activity against a variety of target cells. However, the mechanisms by which sensitive targets activate killing have been difficult to study due to the lack of a prototypic NK cell triggering receptor. Pharmacologic evidence has implicated protein tyrosine kinases (PTKs) in natural killing; however, Lck-deficient, Fyn-deficient, and ZAP-70-deficient mice do not exhibit defects in natural killing despite demonstrable defects in T cell function. This discrepancy implies the involvement of other tyrosine kinases. Here, using combined biochemical, pharmacologic, and genetic approaches, we demonstrate a central role for the PTK Syk in natural cytotoxicity. Biochemical analyses indicate that Syk is tyrosine phosphorylated after stimulation with a panel of NK-sensitive target cells. Pharmacologic exposure to piceatannol, a known Syk family kinase inhibitor, inhibits natural cytotoxicity. In addition, gene transfer of dominant-negative forms of Syk to NK cells inhibits natural cytotoxicity. Furthermore, sensitive targets that are rendered NK-resistant by major histocompatibility complex (MHC) class I transfection no longer activate Syk. These data suggest that Syk activation is an early and requisite signaling event in the development of natural cytotoxicity directed against a variety of cellular targets.

Animals↗

PubMed1996

Abrogation of the Fc gamma receptor IIA-mediated phagocytic signal by stem-loop Syk antisense oligonucleotides.

The role of Syk kinase in Fc gamma receptor (Fc gamma R) IIA-mediated phagocytosis was examined with two forms of antisense oligodeoxynucleotides (ODNs) designed to hybridize to human Syk mRNA. Monocytes were incubated with linear and stem-loop antisense ODNs targeted to Syk mRNA. When complexed with cationic liposomes, stem-loop Syk antisense ODN with phosphorothioate modification exhibited stability in fetal bovine and human serum. The stem-loop Syk antisense ODN at a concentration of 0.2 microM inhibited Fc gamma RIIA-mediated phagocytosis by 90% and completely eliminated Syk mRNA and protein in monocytes, whereas scrambled-control ODNs had no effect. The Syk antisense ODNs did not change beta-actin mRNA levels and Fc gamma RII cell-surface expression. In addition, stem-loop Syk antisense ODN inhibited Fc gamma RI and Fc gamma RIIIA-mediated phagocytosis. These data indicate the efficacy of stem-loop Syk antisense ODN for targeting and degrading Syk mRNA and protein and the importance of Syk kinase in Fc gamma receptor-mediated phagocytosis. Immunoblotting assay demonstrated that Fc gamma RII tyrosine phosphorylation after Fc gamma RII cross-linking did not change in the absence of Syk protein. These results indicate that Syk kinase is required for Fc gamma RIIA-mediated phagocytic signaling and that Fc gamma RII cross-linking leads to tyrosine phosphorylation of Fc gamma RII independent of Syk kinase.

Animals↗

PubMed1999

Xenopus laevis occludin. Identification of in vitro phosphorylation sites by protein kinase CK2 and association with cingulin.

Occludin is a protein component of the membrane domain of tight junctions, and has been shown to be phosphorylated in vivo in cultured cells and Xenopus laevis embryos. However, nothing is known about the identity of specific occludin kinase(s) and occludin phosphorylation site(s). Furthermore, nothing is known about the interaction of occludin with cingulin, a cytoplasmic plaque component of tight junctions. Here we report the isolation and sequencing of a complete X. laevis occludin cDNA, and experiments aimed at mapping X. laevis occludin in vitro phosphorylation site(s) and characterizing occludin interaction with cingulin. The sequence of Xenopus occludin is homologous to that of occludins from other species, with identities ranging from 41% to 58%. Bacterially expressed domain E of Xenopus occludin (amino acids 247-493) was a good substrate for protein kinase CK2 (stoichiometry 10.8%, Km 8.4 microM) but not for CK1 kinase, protein kinase A, cdc2 kinase, MAP kinase or syk kinase. Residues Thr375 and Ser379 were identified as potential CK2 phosphorylation sites in this region based on sequence analysis. Mutation of Ser379 to aspartic acid or alanine reduced phosphorylation by CK2 by approximately 50%, and double mutation of Ser379 into aspartic acid and Thr375 into aspartic acid essentially abolished phosphorylation. Glutathione S-transferase (GST) pull-down experiments using extracts of Xenopus A6 epithelial cells showed that constructs of GST fused to wild-type and mutant forms of the C-terminal region of X. laevis occludin associate with several polypeptides, and immunoblot analysis showed that one of these polypeptides is cingulin. GST pull-down experiments using in vitro translated, full-length Xenopus cingulin indicated that cingulin interacts directly with the C-terminal region of occludin.

Amino Acid Sequence↗

PubMed2002

Adenosine diphosphate (ADP)-induced thromboxane A(2) generation in human platelets requires coordinated signaling through integrin alpha(IIb)beta(3) and ADP receptors.

Adenosine diphosphate (ADP) is a platelet agonist that causes platelet shape change and aggregation as well as generation of thromboxane A(2), another platelet agonist, through its effects on P2Y1, P2Y12, and P2X1 receptors. It is now reported that both 2-propylthio-D-beta gamma-dichloromethylene adenosine 5'-triphosphate (AR-C67085), a P2Y12 receptor-selective antagonist, and adenosine-2'-phosphate-5'-phosphate (A2P5P), a P2Y1 receptor-selective antagonist, inhibited ADP-induced thromboxane A(2) generation in a concentration-dependent manner, indicating that coactivation of the P2Y12 and P2Y1 receptors is essential for this event. SC49992, a fibrinogen receptor antagonist, blocked ADP-induced platelet aggregation and thromboxane A(2) production in a concentration-dependent manner. Similarly, P2 receptor antagonists or SC49992 blocked ADP-induced arachidonic acid liberation. Whereas SC49992 blocked arachidonic acid-induced platelet aggregation, it failed to inhibit thromboxane A(2) generation induced by arachidonic acid. Thus, ADP-induced arachidonic acid liberation, but not subsequent conversion to thromboxane A(2), requires outside-in signaling through the fibrinogen receptor. The Fab fragment of ligand-induced binding site-6 (LIBS6) antibody, which induces a fibrinogen-binding site on the integrin alpha(IIb)beta(3), caused both platelet aggregation and thromboxane A(2) generation. Inhibitors of phosphoinositide 3-kinase, Syk, Src kinases, or protein tyrosine phosphatases inhibited platelet aggregation but not thromboxane A(2) generation, indicating that these signaling molecules have no significant role in phospholipase A(2) activation. In the presence of P2 receptor antagonists A2P5P or AR-C67085, LIBS6 failed to generate thromboxane A(2), suggesting that inside-out signaling through ADP receptors is necessary for this event. It was concluded that both outside-in signaling from the fibrinogen receptor and inside-out signaling from the P2Y1 and P2Y12 receptors are necessary for phospholipase A(2) activation, resulting in arachidonic acid liberation and thromboxane A(2) generation.

Adenosine Diphosphate↗

PubMed2002

Specific monitoring of Syk protein kinase activity by peptide substrates including constrained analogs of tyrosine.

The ability of Syk protein tyrosine kinase (PTK) to phosphorylate peptides, where tyrosine had been replaced by conformationally constrained analogs, has been exploited to develop highly selective substrates suitable for the specific monitoring of Syk activity. In particular we have synthesized a peptidomimetic, RRRAAEDDE(L-Htc)EEV (syktide), with the 3(S)-7-hydroxy-1,2,3,4-tetrahydroisoquinoline-3-carboxyl acid residue (L-Htc) replaced for tyrosine, which is phosphorylated by Syk with remarkable efficiency (K(cat)=73 min(-1), K(m)=11 microM), while it is not affected to any appreciable extent by a number of PTKs tested so far. These properties make syktide the first choice substrate for the specific monitoring of Syk.

Animals↗

PubMed2001

The IL-15R alpha chain signals through association with Syk in human B cells.

The alpha-chain of the IL-15R (IL-15Ralpha) serves as the specific, high-affinity receptor for IL-15. It is expressed by lymphoid and nonlymphoid cells, including B cell lymphoma lines. In this study, we have further explored IL-15Ralpha-mediated signaling in activated primary B cells and in Raji cells, a human B-lymphoblastoid cell line which expresses the IL-15Ralpha and IL-2Rgamma chains, but lacks the IL-2Rbeta chain. Stimulation of Raji cells with IL-15 induces their proliferation and rescues them from C2-ceramide-induced apoptosis. By immunoprecipitation and Western blotting, we show that treatment of Raji cells and activated primary B cells with IL-15 induces coprecipitation of Syk kinase with the IL-15Ralpha chain. Upon association, the activated Syk kinase phosphorylates the IL-15Ralpha chain as well as phospholipase Cgamma, which coprecipitates with Syk. Furthermore, transfection of Raji cells with stem-loop Syk antisense oligonucleotides prevents IL-15Ralpha and phospholipase Cgamma phosphorylation as well as the inhibition of apoptosis by IL-15. Mutation of a defined region of the intracellular signaling portion of IL-15Ralpha (Tyr227) abrogates both the IL-15Ralpha/Syk association and IL-15Ralpha phosphorylation. Taken together, this suggests that Syk kinase physically and functionally associates with the IL-15Ralpha chain in B cells and that Syk plays a key role in mediating IL-15-induced signal transduction, thus accounting for the distinct functional consequences of IL-15 vs IL-2 binding to B cells.

Apoptosis↗

PubMed2004

A novel mode of Gleevec binding is revealed by the structure of spleen tyrosine kinase.

Spleen tyrosine kinase (Syk) is a non-receptor tyrosine kinase required for signaling from immunoreceptors in various hematopoietic cells. Phosphorylation of two tyrosine residues in the activation loop of the Syk kinase catalytic domain is necessary for signaling, a phenomenon typical of tyrosine kinase family members. Syk in vitro enzyme activity, however, does not depend on phosphorylation (activation loop tyrosine --> phenylalanine mutants retain catalytic activity). We have determined the x-ray structure of the unphosphorylated form of the kinase catalytic domain of Syk. The enzyme adopts a conformation of the activation loop typically seen only in activated, phosphorylated tyrosine kinases, explaining why Syk does not require phosphorylation for activation. We also demonstrate that Gleevec (STI-571, Imatinib) inhibits the isolated kinase domains of both unphosphorylated Syk and phosphorylated Abl with comparable potency. Gleevec binds Syk in a novel, compact cis-conformation that differs dramatically from the binding mode observed with unphosphorylated Abl, the more Gleevec-sensitive form of Abl. This finding suggests the existence of two distinct Gleevec binding modes: an extended, trans-conformation characteristic of tight binding to the inactive conformation of a protein kinase and a second compact, cis-conformation characteristic of weaker binding to the active conformation. Finally, the Syk-bound cis-conformation of Gleevec bears a striking resemblance to the rigid structure of the nonspecific, natural product kinase inhibitor staurosporine.

Animals↗

PubMed1995

Phosphorylated immunoreceptor signaling motifs (ITAMs) exhibit unique abilities to bind and activate Lyn and Syk tyrosine kinases.

Signal transduction by T and B cell Ag receptors and certain receptors for Ig Fc regions (Fc gamma RI, hFc gamma RIIA, Fc gamma RIII, Fc alpha R, and Fc epsilon RI) involves a conserved sequence motif, termed an immunoreceptor tyrosine-based activation motif (ITAM) and found in multiple receptor chains. Phosphorylation of the two ITAM tyrosines is a critical event in signal transduction. To address the function of this phosphorylation, we assessed the ability of nonphosphorylated and biphosphorylated ((p)2ITAM) ITAM peptides to bind and modify the activity of src and syk family kinases in vivo and in vitro. All (p)2ITAMs, but not their nonphosphorylated counterparts, induced extensive protein tyrosine phosphorylation in permeabilized cells. However, the patterns of proteins phosphorylated differed among (p)2ITAMs. This phosphorylation was found to reflect activation of the src family kinase Lyn, but not Syk. In vitro studies using purified Lyn showed that src family kinase activation resulted from a direct interaction with (p)2ITAM. Binding studies demonstrated clear differences in binding specificity of (p)2ITAMs. Most strikingly, Ig alpha (p)2ITAM and TCR-zeta c and CD3 epsilon (p)2ITAMs exhibit inverse binding preferences for src and syk family kinases. Taken together, these findings demonstrate a novel mechanism by which src family tyrosine kinases are activated, and are consistent with the possibility that different ITAMs may preferentially activate distinct signaling pathways as a consequence of distinct effector Src homology 2 domain (SH2) binding preference.

Amino Acid Sequence↗

PubMed1994

Molecular cloning of human Syk. A B cell protein-tyrosine kinase associated with the surface immunoglobulin M-B cell receptor complex.

B cell antigen receptors are multicomponent complexes consisting of the surface immunoglobulin and accessory molecules with associating protein-tyrosine kinases. A spleen tyrosine kinase, Syk, in porcine B cells and a 72-kDa protein-tyrosine kinase, PTK72, in murine B cells associate with the B cell antigen receptor. Herein, we report the isolation of a full-length cDNA encoding the human homologue of Syk. This cDNA predicted a polypeptide consisting of two NH2-terminal SH2 domains and a COOH-terminal tyrosine kinase domain. Syk is highly conserved between human and swine and is homologous to the T cell-associated protein-tyrosine kinase ZAP-70. Both Syk mRNA and protein were detected in cells derived from multiple hematopoietic lineages. Within the B cell compartment, Syk was expressed from pro-B cells to plasma cells. In vitro kinase assays conducted on the human Syk protein isolated from B cells revealed the presence of autophosphorylation activity on Syk tyrosine residues. Tyrosine phosphorylation of Syk associating with the B cell receptor complex in human was augmented rapidly after surface immunoglobulin cross-linking. The human SYK locus was mapped to chromosome 9 at band q22.

Amino Acid Sequence↗

PubMed1995

Syk tyrosine kinase required for mouse viability and B-cell development.

The Syk cytoplasmic protein-tyrosine kinase has two amino-terminal SH2 domains and a carboxy-terminal catalytic domain. Syk, and its close relative ZAP-70, are apparently pivotal in coupling antigen- and Fc-receptors to downstream signalling events. Syk associates with activated Fc receptors, the T cell receptor complex and the B-cell antigen-receptor complex (BCR) in immature and mature B lymphocytes. On receptor activation, the tandem SH2 domains of Syk bind dual phosphotyrosine sites in the conserved ITAM motifs of receptor signalling chains, such as the immunoglobulin alpha and beta-chains of the BCR, leading to Syk activation. Here we have investigated Syk function in vivo by generating a mouse strain with a targeted mutation in the syk gene. Homozygous syk mutants suffered severe haemorrhaging as embryos and died perinatally, indicating that Syk has a critical role in maintaining vascular integrity or in wound healing during embryogenesis. Analysis of syk-/- lymphoid cells showed that the syk mutation impaired the differentiation of B-lineage cells, apparently by disrupting signalling from the pre-BCR complex and thereby preventing the clonal expansion, and further maturation, of pre-B cells.

Animals↗

PubMed1996

Syk tyrosine kinase is required for immunoreceptor tyrosine activation motif-dependent actin assembly.

Clustering of several multisubunit receptors on hematopoetic cells results in a signaling cascade involving the phosphorylation of immunoreceptor tyrosine activation motifs, or "ITAMs," and actin polymerization. Recent experiments indicate that direct clustering of the ITAM-binding protein, p72(syk) (Syk), is capable of transmitting a phagocytic signal in COS cells (Greenberg, S., Chang, P., Wang, D., Xavier, R., and Seed, B.(1996) Proc. Natl. Acad. Sci. U. S. A. 93, 1103-1107). However, the possibility of redundant signaling pathways makes it difficult to test the requirement for Syk in ITAM-dependent actin polymerization in hematopoetic cells. We developed a model system to study ITAM-dependent actin assembly. DT40 lymphocytes were transfected with fusion proteins encoding the transmembrane and cytosolic domains of the ITAM-containing gamma subunit of Fc receptors. Clustering the gamma-containing fusion proteins with IgG-coated erythrocytes triggered submembranous actin assembly. This response depended on an intact ITAM, was absent in cell lines that had been engineered to lack Syk, and was augmented in cell lines that stably overexpressed Syk. These experiments demonstrate an absolute requirement for Syk tyrosine kinase in ITAM-dependent actin assembly in transfected lymphocytes.

Actins↗

PubMed1998

Structural basis for Syk tyrosine kinase ubiquity in signal transduction pathways revealed by the crystal structure of its regulatory SH2 domains bound to a dually phosphorylated ITAM peptide.

The Syk family of kinases, consisting of ZAP-70 and Syk, play essential roles in a variety of immune and non-immune cells. This family of kinases is characterized by the presence of two adjacent SH2 domains which mediate their localization to the membrane through receptor encoded tyrosine phosphorylated motifs. While these two kinases share many structural and functional features, the more ubiquitous nature of Syk has suggested that this kinase may accommodate a greater variety of motifs to mediate its function. We present the crystal structure of the tandem SH2 domain of Syk complexed with a dually phosphorylated ITAM peptide. The structure was solved by multiple isomorphous replacement at 3.0 A resolution. The asymmetric unit comprises six copies of the liganded protein, revealing a surprising flexibility in the relative orientation of the two SH2 domains. The C-terminal phosphotyrosine-binding site is very different from the equivalent region of ZAP-70, suggesting that in contrast to ZAP-70, the two SH2 domains of Syk can function as independent units. The conformational flexibility and structural independence of the SH2 modules of Syk likely provides the molecular basis for the more ubiquitous involvement of Syk in a variety of signal transduction pathways.

Amino Acid Sequence↗

PubMed1995

Differential binding activity of ARH1/TAM motifs.

T- and B-cell antigen receptors, and certain receptors for IgG and IgE constant regions, transduce signals via a conserved amino acid sequence motif, termed ARH1 or TAM. Receptor ligation leads to phosphorylation of 2 tyrosines found within the motif and this phosphorylation appears critical for signal transduction. Although this 26-residue motif exhibits some functional redundancy, its variability in sequence and occurrence in multiple forms in individual receptor complexes, e.g., as many as 8 copies in TCR, suggests that individual ARH1 motifs may exhibit partially unique function. To begin to address this possibility, we compared the binding activity of doubly phosphorylated and non-phosphorylated Ig alpha, Ig beta, TcR zeta c and CD3 epsilon ARH1 motifs. Results demonstrate a clear difference in binding activity determined by both motif phosphorylation and primary structure. Among non-phosphorylated motifs, Ig alpha exhibits the most readily detectable binding activity; binding src-family kinases [1], CD22, MAPK, PI3-k, and Shc, but not CD19. Among doubly phosphorylated motifs, Ig alpha, Ig beta, TCR zeta c and CD3 epsilon all exhibit binding activity but have distinct effector preferences. For example, while Ig alpha prefers src-family kinases over the Syk kinase and binds Shc avidly, CD3 epsilon prefers Syk over src-kinases and does not bind Shc. TCR zeta c seems to bind Syk, src-kinases and Shc. These data are consistent with the possibility that ARH1 motifs may be coupled to distinct signal propagation mechanisms.

Amino Acid Sequence↗

PubMed1997

Role of Tyr518 and Tyr519 in the regulation of catalytic activity and substrate phosphorylation by Syk protein-tyrosine kinase.

The Syk protein-tyrosine kinase is expressed in many hematopoietic cells and is involved in signaling from various receptors for antigen and Fc portions of IgG and IgE. After cross-linking of these receptors, Syk is rapidly phosphorylated on tyrosine residues. We have previously reported that Syk expressed in COS cells is predominantly phosphorylated at both Tyr518 and Tyr519 at its putative autophosphorylation site. In this study, we have examined the role of each of these two residues for the catalytic activity of Syk in vitro and for the Syk-induced phosphorylation of cellular proteins in intact cells. Mutation of either residue had minor effects on the catalytic activity of Syk, and even the double mutant [F518, F519]Syk was about 60% as active as the wild-type enzyme. In intact cells, however, all three mutants consistently failed to induce the extensive tyrosine phosphorylation of cellular proteins typically observed with wild-type Syk. We have recently shown that the doubly phosphorylated Y518/Y519 site is also the site for association of Syk with the SH2 domain of the Lck kinase, which suggests that although phosphates at Y518/Y519 may enhance the catalytic activity of Syk, its interaction with Src family protein-tyrosine kinases is at least equally important for the induction of downstream substrate phosphorylation.

Amino Acid Sequence↗