Apoptosis but Not Other Activation Events Is Inhibited by a Mutation in the Transmembrane Domain of T Cell Receptor β That Impairs CD3ζ Association

Abstract

The transmembrane domain of T cell receptor (TCR) ␤ contains a conserved immunoreceptor tyrosine-based activation-like motif consisting of a duplicated YXXL sequence. The motif is also present in TCR␥, the equivalent chain to TCR␤ in ␥␦ T lymphocytes but is absent in TCR␣ and TCR␦. To determine the putative role of this sequence in TCR⅐CD3 complex assembly and signal transduction, a TCR␤ chain cDNA was mutated in the C-terminal tyrosine of the motif, cloned in an expression vector, and transfected into TCR␤-negative Jurkat cells. Transfectants of the mutated chain (MUT) expressed, on average, much less TCR⅐CD3 complex on the membrane than wild type TCR␤ transfectants. Radiolabeling experiments suggested that the mutation caused a loose association of the CD3 chain resulting in a defective assembly. However, stimulation of high TCR⅐CD3 expressing wild type and MUT clones with monoclonal antibodies and Staphylococcus aureus enterotoxin B resulted in similar levels of CD25 and CD69 expression, interleukin-2 secretion, and TCR⅐CD3 complex downregulation. By contrast, MUT cells were clearly resistant to activation-induced cell death, and they did not express CD95-ligand upon activation. These results suggest a differentiated intracellular signaling pathway leading to apoptosis in which Tyr-TM11 of the immunoreceptor tyrosine-based activation motif-like motif and CD3 appear to be involved. The T cell antigen receptor⅐CD3 complex (TCR⅐CD3)1 is formed by a clonotypic heterodimer (␣␤ or ␥␦), which provides ligand specificity, non-covalently linked to at least four invariant chains (CD3⑀, -␥, -␦, and -) (for review see Refs. 1-3). Assembly occurs by pairwise interactions (4), and as a result, the TCR⅐CD3 complex is formed by four dimeric components as follows: (a) clonotypic TCR␣ and TCR␤ chains that are covalently linked via a single extracellular disulfide bond; (b) the non-covalent CD3␥⑀ dimer; (c) the non-covalent CD3␦⑀ dimer; and (d) a disulfide-linked family dimer consisting of any of the five defined members of this family, , , , , and the ␥ chain of the high affinity Fc⑀ receptor (5). In addition to these interactions, stable pairwise associations can also be observed between single clonotypic and CD3 chains (4, 6, 7). Nevertheless, the CD3 dimer will only assemble in the complex if all the other subunits are present, thus explaining why CD3 is the last component to be integrated into the TCR⅐CD3 complex during assembly (8 -10). The stoichiometry, as well as the possible formation of alternative TCR⅐CD3 complexes, is still a question of debate.The ability of antigen receptors to transduce signals to multiple biochemical cascades is the central event of immune cell activation (11). Engagement of the multicomponent TCR⅐CD3 complex with its antigen/MHC ligand, agonist mAbs, or superantigens results in several biochemical processes critical for the functional activation of T lymphocytes, including cellular proliferation, cell differentiation, and programmed cell death. Using chimeric molecules and reconstituted recept...

CD38 Signaling in T Cells Is Initiated within a Subset of Membrane Rafts Containing Lck and the CD3-ζ Subunit of the T Cell Antigen Receptor

“…Perhaps the best example is the CD3--CD3-⑀ complexes, which are detected in both raft and soluble fractions. Our data are consistent with the evidence that CD3-is loosely associated to the other TCR-CD3 subunits (66), and therefore its interaction with the other CD3 polypeptide chains is more sensitive to non-ionic detergents than the more tightly associated TCR-␣␤⅐CD3-␥⑀ or TCR-␣␤⅐CD3-␦⑀ subcomplexes (50,67,68).…”

Opposite Ability of Pre-TCR and αβTCR to Induce Apoptosis

“…Impaired apoptosis following ␣␤TCR cross-linking on mature T cells has previously been linked to defects in the association of ␣␤TCR with TCR (13)(14)(15)29). From these studies, it could be inferred that the failure of pre-TCR, as opposed to ␣␤TCR, to induce apoptosis in SL12 cells could be related to the constitutive weak physical association between pre-TCR and TCR (10 -12).…”

“…For instance, while IL-2 secretion is not affected in activated T cell hybridomas or peripheral T cells expressing "ITAM (immunoreceptor tyrosine-based activation motif)-less" TCR chains, the subsequent induction of apoptosis is abrogated (15,29). More importantly, a mutation in the TCR␤ chain that abolishes the strong physical association of TCR to the ␣␤ TCR complex in Jurkat cells prevents these cells from undergoing activation-induced cell death, without affecting other responses (13).…”

“…Third, while TCR chains are functionally linked to both TCRs, physical association of TCR is much stronger with the ␣␤TCR than with the pre-TCR (8 -12). Notably, TCR has been shown to play a major role in TCR-mediated apoptosis in mature T cells (13)(14)(15)(16).…”

Fc Receptor γ Chain Residues at the Interface of the Cytoplasmic and Transmembrane Domains Affect Association with FcαRI, Surface Expression, and Function

“…Transfection of TCR ␤-deficient Jurkat cells with a cDNA encoding the human TCR ␤ with a mutated transmembrane tyrosine reconstituted a TCR with mostly intracellular expression and weakly associated CD3 subunit (52). The weak association of CD3 resulted in defective recruitment of ZAP70 (53).…”