Disease relevance of Zap70

• Tissue hyperplasia and enhanced T-cell signalling via ZAP-70 in c-Cbl-deficient mice [1].
• Restoration of thymic development in an Lck(-/-) thymoma overexpressing ZAP-70 [2].
• Defective recruitment and activation of ZAP-70 in common variable immunodeficiency patients with T cell defects [3].
• Moreover, we demonstrate that this inhibitor prevented tetanus toxoid antigen-specific T cell proliferation and expansion but failed to block activation of Zap70 or p56Lck after anti-CD3 stimulation of human T cells [4].
• As human tumors are unlikely to grow to such large sizes (e.g., > 20% of the total body weight), the significance of the alterations in T cell expression of lck, fyn, ZAP-70, or CD3-zeta in the immune status of cancer patients is unclear [5].

High impact information on Zap70

• Data from recent knockout experiments in cell lines and mice contradict the assumption that Src-family kinase and tyrosine kinases of the Syk/Zap-70 family are the transducer elements that set signaling from these receptors in motion [6].
• This results in the phosphorylation of CD3epsilon and Zap-70 signal transducing molecules [7].
• Here we show that mice lacking ZAP-70 had neither CD4 nor CD8 single-positive T cells, but human ZAP-70 reconstituted both CD4 and CD8 single-positive populations [8].
• Natural killer cell function was intact in the absence of ZAP-70 [8].
• Zeta phosphorylation without ZAP-70 activation induced by TCR antagonists or partial agonists [9].

Chemical compound and disease context of Zap70

• Mutations in the ZAP-70 protein tyrosine kinase gene result in a severe combined immunodeficiency (SCID) characterized by a selective inability to produce CD8(+) T cells and a signal transduction defect in peripheral CD4(+) cells [10].

Biological context of Zap70

• In such mice, termination of Zap70 expression impaired TCR signal transduction and arrested thymocyte development after the initiation, but before the completion, of positive selection [11].
• Phosphorylations of the Zap70 substrates (linker of activated T cells and phospholipase Cgamma1 were also profoundly diminished [12].
• This state was contrasted with T cell clonal anergy in which the RAS/MAPK pathway was preferentially impaired and there was much less inhibition of Zap70 kinase activity [12].
• The phenotype is accompanied by increased Zap-70 phosphorylation and activation, including its ubiquitinylated forms [13].
• Furthermore, the association of Zap-70 with c-Cbl was shown to be induced by T cell receptor stimulation, implying that it required posttranslational modification of one or both of these products [14].

Anatomical context of Zap70

• T cells have an alternative pathway in which T cell receptor-activated tyrosine kinase Zap70 phosphorylates p38 on Tyr323 [15].
• In contrast to ZAP-70, expression of Syk is 12- to 15-fold higher in peripheral B cells when compared with peripheral T cells [16].
• Neither cyclosporin A nor inhibitors of PKC produced detectable inhibition of ZAP-70 phosphorylation or MAP kinase activation in these Th2 cells [17].
• Transduction of TCR signals in the thymus involves tyrosine phosphorylation of the protein tyrosine kinase ZAP-70 by p56(lck) and results in induction of ZAP-70 enzymatic activity [18].
• Syk, but not Zap-70 alone, could also stimulate tyrosine phosphorylation of a zeta-bearing chimera in transiently transfected Cos-1 cells [19].

Associations of Zap70 with chemical compounds

• This required the TCR-proximal tyrosine kinase Zap70 but not the adaptor protein LAT, which was required for activation of extracellular signal-regulated protein kinase MAPKs [20].
• This finding contradicts the accepted mechanism of c-Cbl-mediated negative regulation, which requires TKB domain targeting of phosphotyrosine 292 in ZAP-70 [21].
• Furthermore, DP thymocytes with mutant receptors can respond to agonist ligand with normal antigen sensitivity and to normal levels, as shown by their ability to induce CD69 up-regulation, TCR down-regulation, negative selection, and ZAP70 and c-Jun NH2-terminal kinase activation [22].
• We show that ZAP-70 is also required for migration of T cells that is dependent on the integrin LFA-1 [23].
• LFA-1 cross-linking induced tyrosine phosphorylation, which was blocked by dominant-negative ZAP-70 and piceatannol [23].

Physical interactions of Zap70

• We show that ZAP-70 fails to bind the signaling-competent CD3zeta tyrosine phosphorylation isoform and to become activated following TCR engagement, suggesting that defective recruitment of ZAP-70 might underlie the TCR signaling dysfunction in these patients [3].
• The Cbl phosphotyrosine-binding domain selects a D(N/D)XpY motif and binds to the Tyr292 negative regulatory phosphorylation site of ZAP-70 [24].
• Co-immunoprecipitation experiments showed, moreover, that a small amount of Vav is engaged in the multimolecular complex that includes elements of the T cell receptor and the T cell specific ZAP-70 tyrosine kinase [25].

Enzymatic interactions of Zap70

• Thus, we propose that Fyn phosphorylates and activates ZAP-70 and that both kinases cooperate in TCR signaling [26].
• At this time, ZAP-70 from both normal and c-Cbl-/- thymocytes becomes hyperphosphorylated; however, only in normal thymocytes does this correlate with ZAP-70 down-regulation and a diminished ability to phosphorylate LAT and SLP-76 [27].
• Recruitment of ZAP-70 protein tyrosine kinase to the T cell antigen receptor (TCR) is mediated by the binding of the SH2 domains of this enzyme to phosphorylated ITAM motifs in the CD3 and TCRzeta subunits [28].

Regulatory relationships of Zap70

• Lck regulates the tyrosine phosphorylation of the T cell receptor subunits and ZAP-70 in murine thymocytes [29].
• First, Cbl deficiency markedly up-regulated the activity of ZAP-70 and mitogen-activated protein kinases [30].
• In contrast, ZAP-70 appeared to control TCR-directed Vav phosphorylation [31].
• SLP-76 is an adapter protein expressed in T cells and myeloid cells that is a substrate for ZAP-70 and Syk [32].
• In addition to supporting the notion that Src-related enzymes regulate the recruitment of Zap-70 in TCR signaling, these data added further complexity to previous models of regulation of Zap-70 [33].

Other interactions of Zap70

• The Src-family and Syk/ZAP-70 family of protein tyrosine kinases (PTK) are required for T cell receptor (TCR) functions [29].
• Moreover, ligation of the TCR expressed on thymocytes from Lck-deficient mice largely failed to induce the phosphorylation of TCR-zeta, CD3 epsilon, or ZAP-70 [29].
• Compared to Jurkat and HT-2 cells, fresh T cells show both higher baseline levels and lower induced levels of Zap-70 function in CD3zeta complexes [34].
• Cytochalasin D, an inhibitor of actin polymerization, inhibited ZAP-70, MAP kinase activation, or IL-4/IL-10 secretion [17].
• To address the role of these residues in T cell development and in the function of primary T cells, we have generated mice that express ZAP-70 molecules with Tyr to Phe substitution at position 292 (Y292F) or 315 (Y315F) [35].
• Our results establish a temporal separation of Syk family kinase function during early thymocyte development and a novel role for ZAP-70 [36].

Analytical, diagnostic and therapeutic context of Zap70

• Northern blot analysis indicated that the expression of ZAP-70 mRNA was most pronounced in the thymus and spleen and also slightly in the brain [37].
• Using a newly devised retrovirus gene transfer and an efficient green fluorescence protein detection technique in fetal thymus organ cultures, the present study shows that forced expression in developing thymocytes of a catalytically inactive mutant of ZAP-70, but not wild-type ZAP-70, inhibits T cell development at the earlier CD4-CD8- stage [38].
• An improved retroviral gene transfer technique demonstrates inhibition of CD4-CD8- thymocyte development by kinase-inactive ZAP-70 [38].
• Analysis of ZAP-70 and other signaling proteins of the pre-TCR/TCR was done by Western blot [39].
• Improved treatment efficacy was achieved in infant ZAP-70-/- mice, in which the thymus is proportionately larger and a higher percentage of prothymocytes are in cycle [40].

References