C Couture

McGill University, Montréal, Quebec, Canada

Are you C Couture?

Claim your profile

Publications (26)165.71 Total impact

  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: T-cell activation leads to signaling pathways that ultimately result in induction of gene transcription from the interleukin-2 (IL-2) promoter. We hypothesized that the IL-2 promoter or its synthetic derivatives can lead to T-cell specific, activation-induced transgene expression. Our objective was to develop a retroviral vector for stable and activation-induced transgene expression in T-lymphocytes. First, we compared the transcriptional potency of the full-length IL-2 promoter with that of a synthetic promoter composed of 3 repeats of the Nuclear Factor of Activated T-Cells (NFAT) element following activation of transfected Jurkat T-cells expressing the large SV40 T antigen (Jurkat TAg). Although the NFAT3 promoter resulted in a stronger induction of luciferase reporter expression post stimulation, the basal levels of the IL-2 promoter-driven reporter expression were much lower indicating that the IL-2 promoter can serve as a more stringent activation-dependent promoter in T-cells. Based on this data, we generated a self-inactivating retroviral vector with the full-length human IL-2 promoter, namely SINIL-2pr that incorporated the enhanced green fluorescent protein (EGFP) fused to herpes simplex virus thymidine kinase as a reporter/suicide "bifunctional" gene. Subsequently, Vesicular Stomatitis Virus-G Protein pseudotyped retroparticles were generated for SINIL-2pr and used to transduce the Jurkat T-cell line and the ZAP-70-deficient P116 cell line. Flow cytometry analysis showed that EGFP expression was markedly enhanced post co-stimulation of the gene-modified cells with 1 muM ionomycin and 10 ng/ml phorbol 12-myristate 13-acetate (PMA). This activation-induced expression was abrogated when the cells were pretreated with 300 nM cyclosporin A. These results demonstrate that the SINIL-2pr retrovector leads to activation-inducible transgene expression in Jurkat T-cell lines. We propose that this design can be potentially exploited in several cellular immunotherapy applications.
    Virology Journal 02/2006; 3:97. · 2.09 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Regulation of protein tyrosine kinases (PTKs) by tyrosine phosphorylation is well recognized; in fact, nearly all PTKs require phosphorylation of tyrosine residues in their "activation loop" for catalytic activity. In contrast, the phosphorylation of PTKs on serine and threonine residues has not been studied nearly as much. We report that the ZAP-70 PTK contains predominately phosphoserine in normal T lymphocytes as well as in Jurkat T leukemia cells. We have identified one site of phosphorylation as Ser-520 and find this site to be important for the recruitment and activation of ZAP-70 in T cells. Mutant ZAP-70-S520A had reduced ability to autophosphorylate and to mediate antigen receptor-induced interleukin 2 gene activation and was not enriched at the plasma membrane. These defects were rescued by addition of a myristylation signal to the N terminus of ZAP-70-S520A to force its plasma membrane and lipid raft localization. We conclude that phosphorylation of ZAP-70 at Ser-520 plays an important role in the correct localization of ZAP-70 and in priming ZAP-70 for its acute recruitment and activation upon antigen receptor ligation.
    Molecular and Cellular Biology 12/2003; 23(21):7667-77. · 5.37 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: The ZAP-70 protein-tyrosine kinase plays a central role in signaling from the T cell antigen receptor. Recruitment and activation of ZAP-70 are transient and are terminated by phosphorylation of negative regulatory tyrosine residues and dephosphorylation of positively acting sites. We report that the low molecular weight protein-tyrosine phosphatase (LMPTP) specifically dephosphorylates the negative regulatory Tyr-292 of ZAP-70, thereby counteracting inactivation of ZAP-70. Expression of low levels of LMPTP resulted in increased ZAP-70 phosphorylation, presumably at the activating Tyr-493 and other sites, increased kinase activity, and augmented downstream signaling to the mitogen-activated protein kinase pathway. The ZAP-70 Y292F mutant was not affected by LMPTP. Our results indicate that LMPTP, like CD45, dephosphorylates a negative regulatory tyrosine site in a protein-tyrosine kinase and thereby strengthens T cell receptor signaling.
    Journal of Biological Chemistry 08/2002; 277(27):24220-4. · 4.65 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Transplantable bone marrow stromal cells can be utilized for cell therapy of mesenchymal disorders. They can also be genetically engineered to express synthetic transgenes and subsequently serve as a platform for systemic delivery of therapeutic proteins in vivo. Inducible production of therapeutic proteins would markedly enhance the usefulness of stromal cells for cell therapy applications. We determined whether synthetic corticosteroid hormones can be used to tightly control transgene expression via the glucocorticoid response pathway in primary bone marrow stromal cells. This regulatory mechanism does not require the presence of potentially immunogenic prokaryotic or chimeric "Trans-activators." Further, synthetic corticosteroids are pharmaceutical agents that can be readily used in vivo. We designed a self-inactivating retroviral vector in which expression of the green fluorescent protein (GFP) reporter is controlled by a minimal synthetic promoter composed of five tandem glucocorticoid response elements upstream of a TATAA box. Vesicular stomatitis virus G-pseudotyped retroparticles were synthesized and utilized to transduce cultured cell lines and primary rat bone marrow stromal cells. We have shown that primary rat bone marrow stromal cells could be efficiently engineered with our GRE-containing retrovector, basal reporter expression was low in the absence of exogenous synthetic corticosteroids, and GFP expression was dexamethasone inducible and reversible. To summarize, this strategy allows dexamethasone-induced, "on-demand" transgene expression from transplantable genetically engineered bone marrow stromal cells.
    Human Gene Therapy 10/2000; 11(13):1837-49. · 4.02 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Human immunodeficiency virus type 1 (HIV-1) infection triggers a cytotoxic T-lymphocyte (CTL) response mediated by CD8(+) and perhaps CD4(+) CTLs. The mechanisms by which HIV-1 escapes from this CTL response are only beginning to be understood. However, it is already clear that the extreme genetic variability of the virus is a major contributing factor. Because of the well-known ability of altered peptide ligands (APL) to induce a T-cell receptor (TCR)-mediated anergic state in CD4(+) helper T cells, we investigated the effects of HIV-1 sequence variations on the proliferation and cytotoxic activation of a human CD4(+) CTL clone (Een217) specific for an epitope composed of amino acids 410 to 429 of HIV-1 gp120. We report that a natural variant of this epitope induced a functional anergic state rendering the T cells unable to respond to their antigenic ligand and preventing the proliferation and cytotoxic activation normally induced by the original antigenic peptide. Furthermore, the stimulation of Een217 cells with this APL generated altered TCR-proximal signaling events that have been associated with the induction of T-cell anergy in CD4(+) T cells. Importantly, the APL-induced anergic state of the Een217 T cells could be prevented by the addition of interleukin 2, which restored their ability to respond to their nominal antigen. Our data therefore suggest that HIV-1 variants can induce a state of anergy in HIV-specific CD4(+) CTLs. Such a mechanism may allow a viral variant to not only escape the CTL response but also facilitate the persistence of other viral strains that may otherwise be recognized and eliminated by HIV-specific CTLs.
    Journal of Virology 04/2000; 74(5):2121-30. · 5.08 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Studies with motheaten mice, which lack the SHP1 protein tyrosine phosphatase, indicate that this enzyme plays an important negative role in T cell antigen receptor (TCR) signaling. The physiological substrates for SHP1 in T lymphocytes, however, have remained unclear or controversial. To define these targets for SHP1 we have compared the effects of constitutively active and inactive mutants of SHP1 on TCR signaling. Expression of wild-type SHP1 had a very small effect on the TCR-induced tyrosine phosphorylation of ZAP-70 and Syk, even when SHP1 was overexpressed 20 - 100-fold over endogenous SHP1. Inactive SHP1-D421A and wild-type SHP2 were without effects. Constitutively active SHP1-DeltaSH2 had a more pronounced effect on ZAP-70 and Syk, even when expressed at near physiological levels. SHP1-DeltaSH2 also inhibited events downstream of ZAP-70 and Syk, such as activation of the mitogen-activated protein kinase Erk2 and the transcriptional activation of the interleukin-2 gene. In contrast, a constitutively active SHP2-DeltaSH2 had no statistically significant effect (although it caused a slight augmentation in some individual experiments). None of the constructs influenced the anti-CD3-induced tyrosine phosphorylation of the TCR zeta-chain or phospholipase Cgamma1, indicating that Src family kinase function was intact. Taken together, our findings support the notion that ZAP-70 and Syk can be direct substrates for SHP1 in intact cells. However, the two SH2 domains of SHP1 did not facilitate its recognition of ZAP-70 and Syk as substrates in intact cells. Therefore, we suggest that SHP1 is not actively recruited to inhibit TCR signaling induced by ligation of this receptor alone. Instead, we propose that ligation of a distinct inhibitory receptor leads to the recruitment of SHP1 via its SH2 domains, activation of SHP1 and subsequently inhibition of TCR signals if the inhibitory receptor is juxtaposed to the TCR.
    European Journal of Immunology 09/1999; 29(8):2539-50. · 4.97 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Malignant cells of patients with cutaneous T cell lymphoma (CTCL) are of monoclonal origin and of the CD4+/CD45RO+ subset. Since unlike their normal counterparts, triggering of their TCR/CD3 in vitro elicits only a weak mitogenic response, we set out to determine which of the signal transduction molecules initiated by anti-CD3E antibodies are affected in neoplastic cells. The results obtained from analysis of tumor cells from four patients show a general reduction in basal and induced tyrosine phosphorylation of a wide range of signaling proteins. Furthermore, the function of members from distinct families of protein tyrosine kinases was altered in neoplastic cells. The enzymatic activity of the membrane-bound fraction of Csk was suppressed, and its association with other cellular proteins was altered. There was a decline in the amount and activity of Syk, and a slight decrease in the specific activity of Lck kinases. Zap70 tyrosyl phosphorylation was reduced or undetectable and the kinase associated weakly, or not at all, with the TCR zeta chain. We propose that dampened TCR-triggered responses in CTCL are caused by suppression of an array of effector molecules required for coupling cell surface receptors to early and late signaling events.
    Leukemia 09/1997; 11(8):1338-46. · 10.16 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: We have begun to explore the mechanisms of apoptosis using a cell-free system based on extracts from Xenopus eggs. Nuclei assembled or placed in these extracts undergo the morphological changes typical of apoptosis and eventually disintegrate. We used this system to investigate the potential involvement in apoptosis of proteins containing Src homology 2 (SH2) domains, which are known to interact with specific tyrosine-phosphorylated ligands. SH2 domains from a number of signaling proteins, including Lck, Src, and Abl, inhibited apoptosis when present at concentrations of 10-100 nM. The inhibition was dependent on specific interaction with endogenous tyrosine-phosphorylated ligands. A synthetic peptide ligand for Src family SH2 domains also inhibited apoptosis in a phosphotyrosine-dependent manner. Kinetic analysis defined three phases in the apoptotic process occurring in this cell-free system. SH2 domains and ceramide act throughout the first 60-90 min of the process (the "initiation" phase). Next, Bcl-2, interleukin-1beta converting enzyme family(CPP32-like) proteases, and the heavy membrane fraction act in a period occurring approximately 90-120 min after the start of incubation (the "sentencing" phase). In the final phase ("execution"), the process of active nuclear destruction ensues.
    The Journal of Cell Biology 07/1997; 137(5):1117-25. · 10.82 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: 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.
    European Journal of Biochemistry 07/1997; 246(2):447-51. · 3.58 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: The two related protein-tyrosine kinases Syk and Zap are rapidly phosphorylated on tyrosine residues and enzymatically activated upon crosslinking of the T cell antigen receptor. We have previously reported that the activation of Syk is less dependent on the Src family kinase Lck than the activation of Zap. Here we report that overexpression of Syk in the Lck-negative JCaM1 cells enabled the T cell antigen receptor/CD3 complex to induce a normal activation of the mitogen-activated protein kinase (MAPK) pathway and expression of a nuclear factor of activated T cells reporter construct. In contrast, Zap and other protein-tyrosine kinases were unable to reconstitute these signaling pathways when expressed at the same levels. In parallel, Syk was phosphorylated on tyrosine, while Zap was not. The Syk-mediated T cell antigen receptor-induced MAPK activation was detectable within 1 min of receptor stimulation and peaked at 3-5 min. The capacity of Syk to reconstitute the MAPK response required the catalytic activity of Syk, an intact autophosphorylation site (Y518 and Y519), both Src homology 2 domains and it was blocked by the inhibitory N17-mutated dominant-negative Ras construct. A Y341-->F mutant of Syk, which is deficient in its interaction with phospholipase Cy1 and Vav, was less efficient than wild-type Syk. Our results suggest that Syk, in contrast to Zap, can transduce signals from the T cell antigen receptor independently of Lck.
    European Journal of Biochemistry 04/1997; 245(1):84-90. · 3.58 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Certain changes in TCR contact residues have been shown to have profound effects on the capacity of a peptide Ag to stimulate a T cell response. Although some of these changes apparently lead to a complete loss of the ability to interact with the TCR, others result in partial agonist activity (e.g., cytokine production without proliferation) or antagonist activity (i.e., the capacity to inhibit the engagement to the TCR by Ag). We show MHC class II-restricted antagonist activity was associated with a differential pattern of early tyrosine phosphorylation events that was characterized by a preponderance of phosphorylation of low molecular mass TCRzeta and the failure to phosphorylate Zap-70. These early tyrosine phosphorylation patterns are the same as those previously described for partial agonists. Thus, a partial agonist phenotype such as anergy induction cannot be ascribed in a causal manner to this pattern of tyrosine phosphorylation. We further extend the studies of signal transduction elicited by agonist and antagonist peptides by characterizing differential recruitment of Zap-70 associated with TCRzeta isoforms and differential phosphorylation of p120 proto-oncogene c-Cbl. Another early event following TCR engagement by Ag, down-modulation of the TCR, was studied with antagonist peptides. We show that antagonist peptides do not cause TCR down-modulation. This failure may represent a mechanism by which antagonists inhibit antigen-mediated stimulation of T cells.
    The Journal of Immunology 04/1997; 158(5):2057-64. · 5.52 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Activation of resting T lymphocytes is initiated by rapid but transient tyrosine phosphorylation of a number of cellular proteins. Several protein tyrosine kinases and protein tyrosine phosphatases are known to be important for this response. Here we report that normal T lymphocytes express the B isoform of low molecular weight protein tyrosine phosphatase B (LMPTP-B). The cDNA was cloned from Jurkat T cells, and an antiserum was raised against it. LMPTP immunoprecipitated from resting Jurkat T cells was found to be tyrosine phosphorylated. On stimulation of the cells through their T cell antigen receptor, the phosphotyrosine content of LMPTP-B declined rapidly. In co-transfected COS cells, Lck and Fyn caused phosphorylation of LMPTP, whereas Csk, Zap, and Jak2 did not. Most of the phosphate was located at Tyr-131, and some was also located at Tyr-132. Incubation of wild-type LMPTP with Lck and adenosine 5'-O-(thiotriphosphate) caused a 2-fold increase in the activity of LMPTP. Site-directed mutagenesis showed that Tyr-131 is important for the catalytic activity of LMPTP, and that thiophosphorylation of Tyr-131, and to a lesser degree Tyr-132, is responsible for the activation.
    Journal of Biological Chemistry 03/1997; 272(9):5371-4. · 4.65 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Syk family kinases are essential for lymphocyte development and activation. Therefore the identification of their direct effectors is of critical importance. Here, we report that Syk interacts in the yeast two-hybrid system with Vav, a proto-oncogene product exclusively expressed in hematopoietic cells. This interaction was direct, required the catalytic activity of Syk, the SH2 domain of Vav, and tyrosine residues in the linker domain of Syk. Vav also associated with Syk and Zap in antigen receptor-stimulated B or T cells, respectively. Functionally, Vav was phosphorylated by Syk family kinases both in vivo and in vitro. Furthermore, Syk and Vav cooperated to activate NF-AT synergistically. These results indicate that the interaction between Syk family kinases and Vav plays an important role in coupling immune recognition receptors to signaling pathways involved in lymphokine production.
    Immunity 01/1997; 5(6):591-604. · 19.80 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Src homology 2 (SH2) domains bind to phosphotyrosine (Tyr(P)) residues in specific sequence contexts in other proteins and thereby mediate tyrosine phosphorylationdependent protein-protein interactions. The SH2 domain of the Src family kinase Lck is phosphorylated at tyrosine 192 in T cells upon T cell antigen receptor triggering. We have studied the consequences of this phosphorylation on the properties of the SH2 domain and on the function of Lck in T cell activation. We report that phosphorylation at Tyr192 reduced the capacity of the isolated SH2 domain to bind a high affinity peptide ligand and Tyr(P)-containing cellular proteins. This effect was mimicked by mutation of Tyr192 to an acidic residue. In intact T cells, where Lck participates in T cell antigen receptor signal transduction in an SH2 domain-dependent manner, phosphorylation of Tyr192 correlated with reduced downstream signaling. Our results indicate that tyrosine phosphorylation of the SH2 domain of Lck terminates its high affinity binding to ligands, thereby negatively regulating its participation in T cell antigen receptor signaling. This represents a novel mechanism for the regulation of the function of SH2 domains.
    Journal of Biological Chemistry 11/1996; 271(40):24880-4. · 4.65 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: The Syk protein tyrosine kinase (PTK) is expressed in many hematopoietic cells and is involved in signaling from various receptors for antigen and Fc portions of IgG and IgE. Upon cross-linking of these receptors, Syk is rapidly phosphorylated on tyrosine residues and enzymatically activated. We and others have found that the Lck kinase, a member of the Src family of PTKs, binds through its Src homology (SH) 2 domain to tyrosine phosphorylated Syk and to the related Zap kinase. Here we report that this interaction is direct and identify the two tandem tyrosines at the autophosphorylation site of Syk, Tyr518, and Tyr519, as the binding site for the SH2 domain of Lck. Mutation of either or both tyrosines to phenylalanines abrogated binding, while mutation of a second repetition of the motif at Tyr539 and Tyr540, or of the three tyrosines in the C terminus of Syk, did not. The SH2 domain of Lck bound the autophosphorylation site only when both Tyr518 and Tyr519 were phosphorylated. In intact cells the binding of the SH2 domain of Lck correlated with the ability of Syk to induce tyrosine phosphorylation of cellular proteins.
    Journal of Biological Chemistry 10/1996; 271(39):24294-9. · 4.65 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Activation of resting T lymphocytes by ligands to the complex of T cell antigen receptor (TCR) and CD3 is initiated by a series of critical tyrosine phosphorylation and dephosphorylation events. Protein-tyrosine kinases of the Syk, Src and Csk families and the CD45 protein-tyrosine phosphatase (PTPase) are known to be involved in these early biochemical reactions. We have found that one of the two T-cell-expressed SH2-domain-containing PTPases, SHPTP2, is rapidly phosphorylated on tyrosine upon addition of anti-CD3 mAbs. This response was absent in cells lacking the Src family kinase Lck. Concomitantly with tyrosine phosphorylation, SHPTP2 co-immunoprecipitated with two unphosphorylated cellular proteins; phosphatidylinositol 3-kinase p85 and Grb2. Binding of SHPTP2 to Grb2 occurred through the SH2 domain of Grb2, while the association between SHPTP2 and p85 seemed to be mediated through Grb2 as an intermediate. In addition, many other molecules associate with Grb2 and may thereby become juxtaposed to SHPTP2. Our results indicate that SHPTP2 participates actively at an early stage in TCR signaling and that its phosphorylation on tyrosine may direct a Grb2-dependent association with selected substrates.
    European Journal of Biochemistry 06/1996; 237(3):736-42. · 3.58 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Vav, a 95 kDa proto-oncogene product expressed specifically in hematopoietic cells, was originally isolated as a transforming human oncogene. Vav contains an array of functional domains that are involved in interactions with other proteins and, possibly, with lipids. These include, among others, a putative guanine nucleotide exchange domain, a cysteine-rich region similar to the phorbol ester/diacylglycerol-binding domain of protein kinase C, a pleckstrin-homology domain, and Src-homology 2 and 3 (SH2 and SH3, respectively) domains. The presence of these domains, the transforming activity of the vav oncogene, and the rapid increase in tyrosine phosphorylation of Vav induced by triggering of diverse receptors indicate that it plays an important role in hematopoietic cell signaling pathways. Such a role is supported by recent studies using "knockout" mice and transiently transfected T cells, in which Vav deletion or overexpression, respectively, had marked effects on lymphocyte development or activation. The presence of a putative guanine nucleotide exchange domain, the prototype of which is found in the dbl oncogene product, implies that Vav functions as a guanine nucleotide exchange factor (GEF) for one (or more) members of the Ras-like family of small GTP-binding proteins. In support of such a role, Vav preparations were found in some (but not other) studies to mediate in vitro-specific GEF activity for Ras. Additional studies are required to identify the physiological regulators and targets of Vav, and its exact role in hematopoietic cell development and signaling.
    Stem Cells 06/1996; 14(3):250-68. · 7.70 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Syk family kinases are essential for lymphocyte development and activation. Therefore the identification of their direct effectors is of critical importance. Here, we report that Syk interacts in the yeast two-hybrid system with Vav, a proto-oncogene product exclusively expressed in hematopoietic cells. This interaction was direct, required the catalytic activity of Syk, the SH2 domain of Vav, and tyrosine residues in the linker domain of Syk. Vav also associated with Syk and Zap in antigen receptor-stimulated B or T cells, respectively. Functionally, Vav was phosphorylated by Syk family kinases both in vivo and in vitro. Furthermore, Syk and Vav cooperated to activate NF-AT synergistically. These results indicate that the interaction between Syk family kinases and Vav plays an important role in coupling immune recognition receptors to signaling pathways involved in lymphokine production.
    Immunity 01/1996; 5(6):591-604. · 19.80 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Two classes of protein tyrosine kinases (PTK) are utilized by the T cell antigen receptor (TcR)/CD3 complex for initiation of the signaling cascade, the Src-family PTK p56lck and p59fyn, and the Syk-family PTK p70zap and p72syk. In addition, the CD45 phosphotyrosine phosphatase (PTPase) is required for the induction of tyrosine phosphorylation by the TcR/CD3, presumably by positively regulating Src-family PTK. Here we report that CD45 also regulates the Syk-family PTK p70zap (or ZAP-70). In CD45-negative T cells, p70zap was constitutively phosphorylated on tyrosine and co-immunoprecipitated with the TcR-zeta chain. In resting wild-type CD45-positive cells, p70zap was mainly unphosphorylated, but it was rapidly phosphorylated on tyrosine upon treatment of the cells with anti-CD3 or PTPase inhibitors. Finally, p70zap co-distributed with CD45 in intact T cells, and tyrosine phosphorylated p70zap was dephosphorylated by CD45 in vitro. These findings suggest that CD45 plays an important role, direct or indirect, in the regulation of p70zap and its function in TcR/CD3 signaling.
    European Journal of Immunology 05/1995; 25(4):942-6. · 4.97 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Activation of protein tyrosine kinases (PTK) is an initial and obligatory event for the triggering of human natural killer (NK) cells to cytotoxicity. Of the different PTK detected in NK cells, only p56lck has previously been shown to participate in NK cell activation. Here we present evidence that another PTK, p72syk, is involved in activation of NK cells. Stimulation with a monoclonal antibody to to the Fc gamma RIII receptor (CD16) induced an increased tyrosine phosphorylation of p72syk. This phosphorylation correlated with an increased tyrosine kinase activity of p72syk towards a synthetic peptide substrate. A severalfold increase in the catalytic activity of p72syk was also seen after treatment of NK cells with an inhibitor of phosphotyrosine phosphatases, pervanadate. We conclude that triggering of the cytotoxic response in NK cells is associated with activation of p72syk.
    European Journal of Immunology 11/1994; 24(10):2491-6. · 4.97 Impact Factor

Publication Stats

912 Citations
165.71 Total Impact Points

Institutions

  • 1997–2003
    • McGill University
      • • Division of Hematology
      • • Department of Medicine
      Montréal, Quebec, Canada
  • 1999–2000
    • Lady Davis Institute for Medical Research
      Montréal, Quebec, Canada
  • 1994–1999
    • La Jolla Institute for Allergy & Immunology
      • Division of Cell Biology
      La Jolla, CA, United States