Multiple signal transduction pathways activated through the T cell receptor for antigen.

The T cell receptor for antigen (TCR) is a multichain complex on the surface of T lymphocytes which binds peptide antigen and transduces a transmembrane signal leading to IL-2 secretion. Engagement of the TCR leads to activation of a tyrosine phosphorylation pathway and a phospholipase C (PLC) pathway leading to activation of protein kinase C (PCK). Currently available data suggest that the primary event in signal transduction is tyrosine kinase activation, since when this pathway is inhibited, PLC activation is blocked and there is no production of IL-2. The nature of the tyrosine kinase which initiates the signaling cascade is currently unknown. The CD4/CD8 associated kinase p56lck clearly plays a role in tyrosine phosphorylation, but it is clearly not the only tyrosine kinase involved. Studies demonstrating physical association of p59lyn with the TCR implicate fyn as an important candidate for the TCR tyrosine kinase. The protein tyrosine phosphatase CD45 also plays a critical early role in signal transduction since in cells where it is deficient, neither tyrosine kinase activation nor later signaling events are seen. The importance of the PLC/PKC pathway is illustrated by the fact that activation of this pathway alone may lead to IL-2 production. However, there may also be other mechanisms which can generate an IL-2 response. Two proteins known to be involved in growth regulation--p21ras and c-raf--have now been shown to be downstream targets of the PLC/PKC pathway.     

Signal transduction by the TCR for antigen

The past several years have seen the beginning of a shift in the way that TCR signal transduction is studied. Although many investigators continue to identify new molecules, particularly adaptor proteins, others have attempted to look at signaling events in a larger cellular context. Thus the identification of distinct formations of signaling molecules at junctions between T cells and antigen-presenting cells, the role of the cytoskeleton and the partitioning of molecules into specialized lipid subdomains have been the subjects of many publications. Such concepts are helping to assemble a blueprint of how the myriad adaptors and kinases fit together to effect T cell activation.     

Signal transduction elements of the B cell antigen receptor and their role in immunodeficiencies

The primary function of B lymphocytes is to contribute to the elimination of foreign antigens by producing large amounts of soluble antibodies. The activation of B cells through their antigen receptor triggers a dynamic network of intracellular signaling proteins. The recent identification of the cytoplasmic adaptor protein SLP-65 (also called BLNK or BASH) provided insight in how the antigen receptor-regulated protein tyrosine kinases couple to downstream signaling cascades, including the mobilization of Ca2+ ions, activation of mitogen-activated kinases and reorganization of the cytoskeleton architecture. While these events have been mostly studied in mature B cells, it is now clear that the components of the antigen receptor and its downstream effector elements play also a central role during early and late B cell development, and in the apoptotic elimination of B cells with reactivity to self-antigens. Thus, genetic defects affecting the expression of antigen receptor subunits or its intracellular signaling proteins can interfere with B cell development and activation, and can cause severe antibody deficiencies in mouse and man. In this article I summarize our current picture of the B cell antigen receptor, how the extracellular signal is transported into the cell interior, and how dysregulation of these processes contribute to immune defects.     

Signal transduction in T cells.

Rapid progress was made during the past year in the delineation of the nature of the initial biochemical events triggered by the T-cell antigen receptor. Antigen-mediated activation of phospholipase C was demonstrated to require protein tyrosine phosphorylation and, most surprising, activation of the Ras family of signal transduction molecules was shown to closely follow stimulation of the T-cell antigen receptor. Major controversy continues over which events are relevant to the various effector functions of T cells.     

T cell long-term hyporesponsiveness follows antigen receptor engagement and results from defective signal transduction

T cell receptor (TCR)-mediated stimulation of T hybridomas leads to cell activation and lymphokine production that is followed by a long-term hyporesponsiveness. To investigate the biochemical events involved in the induction and maintenance of this antigen receptor hyporesponsiveness or anergy, we have expressed a G protein/PLCβ1-coupled muscarinic subtype 1 acetylcholine receptor in a murine T cell hybrid. Transfected cells were capable of responding to both muscarinic agonists and TCR ligands by inducing interleukin-2 secretion that was sensitive to cyclosporin A and dexamethasone. Both receptors induced tyrosine kinase (TK) activity, but muscarinic stimulation did not affect tyrosine phosphorylation of PLCγ1, nor did the TK inhibitor, herbimycin, block muscarinic receptor-mediated calcium mobilization. These data indicate that in T cells, the muscarinic receptor mediates T cell effector functions by regulating a TK-independent proximal pathway which later converges with the TCR pathway. Using these cells, we have explored the long-term consequences of T cell stimulation via antigen or muscarinic receptors. Our results show that hyporesponsiveness specifically follows TCR engagement and appears to result from a defect in the early signal transduction initiated by TCR cross-linking. A study of TCR-mediated signaling supports this model by showing that tyrosine phosphorylation and calcium mobilization are deficient in hyporesponsive T cells.     

Structure and specificity of the T cell antigen receptor.

The antigen receptor on T lymphocytes is a multi-chain complex of which two chains determine its specificity for antigen. Although these receptor chains possess genetic and structural similarities with membrane-bound immunoglobulin, the B cell antigen receptor, they endow T cells with a distinct specificity. Unlike B cells, T cells recognize foreign antigenic properties only in the form of proteolytic fragments bound to molecules encoded by the major histocompatibility complex. In addition, one stage of the development of T cells in the thymus is dependent upon an antigen receptor-mediated event. Utilizing the response to a well defined antigen as a model system, we discuss the relationship between receptor structure and the specificity of these recognition events.     

T细胞抗原受体复合体信号转导及其与疾病关系的研究进展

 T 细胞抗原受体（TCR）是由 TCRαβ 或 TCRγδ 组成的异源二聚体，它与 CD3 分子组成跨膜蛋白复合体结构。抗原在诱导幼稚 T 细胞或记忆性T细胞进行增殖进而分化成效应细胞时，需要有两个信号刺激，第一信号来自TCR与抗原的特异性结合，第二信号来自抗原提呈细胞（antigen presenting cell，APC）表面的协同刺激因子与 T细胞表面相应受体的相互作用。TCR 通过胞外部分可变区（V 区）的互补决定区（CDR）特异性识别结合抗原；胞内部分在CD3、CD4/CD8 和 CD28 等分子的辅助下，将胞外刺激信号经磷脂酶 C（PLC）-γ活化途径和促分裂原活化蛋白激酶（mitogen activated protein kinaes，MAPK）活化途径传递至胞内，使转录因子活化，这一过程称为 T 细胞活化的信号转导。而这一过程可使 T细胞活化而发挥其生物学作用。TCR/CD3 复合体介导的信号转导是T细胞活化并发生抗原特异性免疫反应的重要途径，很多疾病的发生都与其信号转导异常有关，因此更深入地了解 T 细胞信号转导的分子机制显得尤为重要。本文对 TCR 介导的信号转导途径作了较为系统地阐述，并简要介绍了其异常与几种重要疾病的关系。......     

Defects in signal transduction caused by a T cell receptor beta chain substitution.

An antigen-specific T-T hybridoma was mutagenized with ethylmethane sulfonate and negatively selected by anti-Ly-6 antibody-induced growth inhibition. One of the mutants generated, M4/8, had lost surface expression of a T cell receptor (TcR) V beta 8 epitope detected on the surface of the parental cell line. However, the mutant cell line did express high levels of TcR heterodimer as detected with a pan-specific anti-TcR antibody. CD3 epsilon, Ly-6 and Thy-1 were expressed at levels similar to the wild-type parental cell line. Analysis of the surface TcR/CD3 complexes by immunoprecipitation and two-dimension gel electrophoresis confirmed that the major discernable difference between the wild-type and mutant TcR/CD3 complexes resided in the TcR beta chain. The parental cell line had the potential to express two TcR heterodimers, V alpha V beta 1 and V alpha V beta 8, as determined by Northern blot analysis. Co-modulation experiments suggested that both types of receptors were expressed. However, the V alpha V beta 8 receptor was the predominant form. In contrast, the mutant M4/8 cell line did not synthesize V beta 8 mRNA and, thus, only the V alpha V beta 1 TcR was synthesized. Despite the normal surface expression of TcR/CD3 complex, the M4/8 mutant cell line did not produce interleukin 2 (IL 2) in response to antigen or soluble anti-CD3 epsilon monoclonal antibody (mAb). Furthermore, it responded poorly to concanavalin A, phytohemagglutinin and anti-Ly-6 mAb. Cross-linking of the stimulatory antibodies partially restored the IL 2 response to anti-CD3 epsilon or anti-Ly-6 to wild-type levels. Phorbol ester and ionomycin stimulated a full IL 2 response in the M4/8 cell line, demonstrating that the defect in the decreased signaling in the mutant did not result from a defect in the IL 2 gene program. In conclusion, these data suggested that the pairing of alpha/beta heterodimer not only determined antigen/MHC specificity but also the signaling efficiency of the TcR/CD3 complex.     

Signal transduction from the B cell antigen-receptor.

Ligation of the B cell antigen-receptor triggers an intricate maze of intercalated biochemical events that ultimately affect B cell biological responses. Recent advances have helped to connect many loose ends by identifying key adaptor proteins, such as BLNK/SLP-65, defining crucial roles for phosphatidylinositol-3-kinase and mapping pathways controlling the mitogen-activated protein kinases (ERK, JNK and p38).     

The protein tyrosine kinase inhibitor herbimycin A, but not genistein, specifically inhibits signal transduction by the T cell antigen receptor.

Several lines of evidence implicate a regulatory tyrosine phosphorylation in the activation of phospholipase C (PLC) by the T cell antigen receptor (TCR). These include studies using inhibitors of protein tyrosine kinases (PTKs). In Jurkat T cells expressing the heterologous human muscarinic receptor (HM1), PLC activity can be induced by either the TCR or HM1. HM1 activates PLC via a guanine nucleotide binding protein. We have studied the selectivity of the effects of the PTK inhibitors, herbimycin A and genistein, in this system. The results indicate that these inhibitors have different mechanisms of action, and suggest that herbimycin A, but not genistein, is a specific inhibitor of PTKs in T cells. Herbimycin A markedly inhibited both the resting and induced levels of phosphotyrosine-containing proteins, including the gamma 1 isozyme of PLC and the zeta chain of the TCR, and prevented activation of PLC by anti-TCR mAb. Herbimycin A did not inhibit activation of PLC by HM1. Genistein had a much less pronounced effect than herbimycin A on the appearance of tyrosine phosphoproteins. Moreover, genistein inhibited activation of PLC by both the TCR and HM1, and inhibition was only partial. Genistein was cytotoxic and markedly inhibited protein synthesis in both Jurkat cells and human peripheral lymphocytes. Herbimycin A was not cytotoxic. These findings confirm the role of a regulatory tyrosine phosphorylation in activation of PLC by the TCR. Herbimycin A was a selective inhibitor of a subclass of PTKs in Jurkat cells. In contrast, inhibition of signal transduction and later events in T cells by genistein may be due to effects other than direct inhibition of PTK activity.     

Signal transduction events regulating integrin function and T cell migration

Integrin receptors facilitate T cell function by mediating adhesive events critical for T cell trafficking and recognition of foreign antigen, including interactions with vascular endothelium, extracellular matrix components, and antigen-presenting cells. Consequently, the functional activity of integrin receptors is acutely regulated by various intracellular signals delivered by other cell surface receptors, resulting in rapid changes in T cell adhesion and migration. This review highlights recent insights into our understanding of the signaling events by which the CD3/T cell receptor complex and chemokine receptors regulate integrin function and T cell migration. These studies highlight novel functions for several signaling molecules, including the tyrosine kinases Itk and ZAP-70, and the adapter protein SLAP-130/Fyb. In addition, analysis of the regulation of integrin function and chemokine-mediated migration has highlighted the critical role that spatial localization of signaling molecules plays in signal transduction, and the importance of the actin cytoskeleton in T cell function.     

Requirement of the T cell receptor for antigen presentation by T lymphocytes. Effect of envelope glycoproteins of HIV-1 on antigen presentation by T cells.

We have developed CD4+, tetanus antigen-specific T cell clones that proliferate in the presence of tetanus antigen and autologous irradiated peripheral blood leucocytes (PBL) as antigen-presenting cells (APC). There have been several reports that T cells can present antigen themselves. We have used tetanus antigen-specific T cell clones to examine the effects of envelope glycoproteins of HIV-1 on processing and presentation of antigen to T cells. Cloned T cells were pre-incubated with soluble crude preparation of tetanus antigen for 4 h at 37 degrees C, irradiated, and used as APC (T-APC). These cells could present antigen, as assessed by the ability of the autologous cloned T cells to proliferate. Resting T cells and phytohaemagglutinin-activated T cell blasts from autologous PBL could not present tetanus antigen to the responder cloned T cells. Antigen presentation by T-APC was abrogated by treating cells with anti-HLA-DR but not by anti-HLA-DQ monoclonal antibodies; treatment of tetanus antigen-pulsed T-APC with anti-tetanus antibody also blocked the ability of these cells to induce proliferation in responder T cells. Antigen presentation by cloned T cells was by a chloroquine-resistant pathway. Pretreatment of T-APC with envelope glycoprotein of HIV-1, gp120, did not affect the proliferative responses of the responder T cells. These data suggest that gp120 does not inhibit the antigen-presenting function while suppressing antigen-specific responses.     

Requirement of the T cell antigen receptor occupancy for the target cell lysis by cytolytic T lymphocytes

We have constructed a bivalent bifunctional F(ab)2 fragment with binding specificity for a V beta 8 T cell antigen receptor and human tumor-associated antigen. Using the bifunctional antibody to focus cytolytic activity of mouse CTL to a human carcinoma cell line and anti-V beta 8 TCR Fab' as a competitor, we demonstrate that only a small percentage (-0.5%) of the TCR engaging on the target molecule is sufficient to deliver a lytic signal to the target cells.     

A human T cell tumor which expresses the putative T cell antigen receptor

A monoclonal antibody (mAb) called H1-2D4 which reacts only with the T cell line HPB-ALL and not with other T cell lines, normal or activated peripheral blood cells, B lymphoblasts (B-LCL), or thymocytes has been developed. This mAb cocaps the T3 antigen on HPB-ALL and anti-T3 mAb cocaps the antigen which reacts with H1-2D4. Purified H1-2D4 precipitates a heterodimer from HPB-ALL cells which has components with molecular weights of 51000 and 39000. The properties of the molecule recognized by H1-2D4 suggest that it is the HPB-ALL equivalent of the putative human T cell antigen receptor.     

T cell antigen-receptor signal transduction.

During the past year, major progress has been made in understanding proximal TCR signal-transduction events. Cbl has been identified as a negative regulator of kinases from the ZAP-70/Syk family. Studies on LAT, SLP-76, Itk and Vav have revealed their role in the activation of Ras and phospholipase-Cgamma1-Ca2+ signalling pathways. TCR-induced cytoskeletal changes involve signalling through SLP-76-Vav-Nck to activate effectors of the Rho-family of GTPases. Finally, glycolipid-enriched microdomains play a crucial role in T cell activation.     

A transfected human muscarinic receptor fails to substitute for the T cell antigen receptor complex in CD2-initiated signal transduction

Several T cell surface molecules can activate signal transduction pathways that lead to T cell activation. Like the T cell antigen receptor (TCR), several other molecules, including the sheep erythrocyte receptor CD2, are able to activate the phosphatidylinositol (PI) signal transduction pathway upon stimulation with appropriate agonists. However, CD2-initiated activation of this pathway is dependent on the functional expression of the TCR. Since the T cell does not express other known receptors that activate the PI pathway independent of the TCR, the specificity of the CD2 requirement for a functional TCR is not known. To evaluate the specificity of this requirement, we examined the functional capacity of CD2 to activate the PI pathway in a TCR-deficient cell which had been transfected with a heterologous receptor, the human muscarinic subtype 1 receptor (HM1). HM1 is a member of the cholinergic family of receptors and is known to activate the PI pathway. HM1 can function in the absence of the TCR in a Jurkat-derived T cell host. Here we demonstrate through calcium fluorimetry and PI metabolism assays that HM1 is unable to substitute functionally for the TCR in CD2-initiated signal transduction. These results suggest a specific functional interaction between CD2 and the TCR in CD2-mediated activation of the PI pathway in T cells.