Expression and function of a variant T cell receptor complex lacking CD3-gamma.

Pérez-Aciego, Paloma; Alarcón, Balbino; Arnaiz‐Villena, Antonio; Terhorst, Cox; Timón, Marcos; Segurado, Oscar G.; Regueiro, José R.

doi:10.1084/jem.174.2.319

Cited by 69 publications

(40 citation statements)

References 27 publications

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“…WT31, Ref. 21), and we have reported previously that the lack of CD3␥ affects TCR⅐CD3 assembly and glycosylation in CD4 ϩ T cells (12,22). Therefore, the impaired binding observed in Fig.…”

Section: Deficient Cd8 ϩ T Lymphocytes Was Conformationally Differentmentioning

confidence: 54%

“…Flow Cytometry Analyses-The expression of different surface markers was studied by flow cytometry according to a standard procedure (12). The antibodies used in these studies are listed in Table I.…”

Section: Methodsmentioning

confidence: 99%

“…Calcium Flux Measurement-Intracellular Ca 2ϩ release was induced in PBLs loaded with the fluorescent dye Fluo3AM (Sigma) according to a standard procedure (12).…”

Section: Methodsmentioning

confidence: 99%

“…Then cells were 125 I-labeled using the lactoperoxidase method by adding 2 mCi of Na 125 I (Amersham Pharmacia Biotech), 30 l of a 140 (12), which can be then resolved by deglycosylation. Samples were then divided in two, and after five washings with lysis buffer, one part was digested overnight with N-glycosidase F (N-Gly ϩ , Roche Molecular Biochemicals), whereas the other was left undigested (N-Gly Ϫ ).…”

Section: Methodsmentioning

confidence: 99%

“…ϩ cells had previously been analyzed by immunoprecipitation (12,22) and immunohistochemistry (23) and had been found to contain all TCR␣␤ and CD3 chains except CD3␥. Thus, the impaired expression of their mutant TCR⅐CD3 complex could be explained as a simple consequence of the lack of CD3␥ on several CD3␥-dependent TCR⅐CD3 epitopes.…”

Section: Deficient Cd8 ϩ T Lymphocytes Was Conformationally Differentmentioning

confidence: 99%

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Conformational and Biochemical Differences in the TCR·CD3 Complex of CD8+ Versus CD4+ Mature Lymphocytes Revealed in the Absence of CD3γ

Zapata¹,

Pacheco‐Castro²,

Torres³

et al. 1999

Journal of Biological Chemistry

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Mature CD4؉ and CD8 ؉ T lymphocytes are believed to build and express essentially identical surface ␣␤ T-cell receptor-CD3 (TCR⅐CD3) complexes. However, TCR⅐CD3 expression has been shown to be more impaired in CD8 ؉ cells than in CD4؉ cells when CD3␥ is absent in humans or mice. We have addressed this paradox by performing a detailed phenotypical and biochemical analysis of the TCR⅐CD3 complex in human CD3␥-deficient CD8؉ and CD4 ؉ T cells. The results indicated that the membrane TCR⅐CD3 complex of CD8 ؉ T lymphocytes was conformationally different from that of CD4 ؉ lymphocytes in the absence of CD3␥. In addition, CD8؉ , but not CD4 ؉ , CD3␥-deficient T lymphocytes were shown to contain abnormally glycosylated TCR␤ proteins, together with a smaller, abnormal TCR chain (probably incompletely processed TCR␣). These results suggest the existence of hitherto unrecognized biochemical differences between mature CD4؉ and CD8 ؉ T lymphocytes in the intracellular control of ␣␤TCR⅐CD3 assembly, maturation, or transport that are revealed when CD3␥ is absent. Such lineage-specific differences may be important in receptor-coreceptor interactions during antigen recognition.Mature ␣␤ T lymphocytes recognize pathogen-derived peptides on antigen-presenting cells by means of the multimeric membrane protein ensemble termed the T-cell receptor (TCR) 1 ⅐CD3 complex. This TCR⅐CD3 complex includes two clonally distributed variable chains that directly interact with antigens (TCR␣ and TCR␤) and four invariant polypeptides that regulate assembly and signal transduction (CD3␥, CD3␦, CD3⑀, and ) (1). The assembly of complete TCR⅐CD3⅐ complexes takes place in a highly ordered manner within the endoplasmic reticulum: first CD3 chains, then TCR chains, and finally chains. Further conformational maturation, including carbohydrate processing, occurs in the Golgi apparatus before exportation of mature complexes to the T cell surface. The biochemical machinery involved in the assembly, processing, and exportation of TCR⅐CD3 complexes is assumed to be shared by all ␣␤ T-lineage cells. Thus, CD4ϩ and CD8 ϩ are believed to build biochemically and conformationally identical antigen receptors, although differences in the numbers that reach or remain at the cell surface have been noted (2). Therefore, the lack of any CD3 chain would be expected to affect to a similar extent the assembly and exportation of TCR⅐CD3 complexes by mature CD4 ϩ and CD8 ϩ T cells. However, this was not the case in several murine and human CD3 deficiencies (reviewed in Ref.3). In particular, it has been consistently shown that in the absence of CD3␥ or CD3␦, TCR⅐CD3 expression (or conformation) is more impaired in mature peripheral CD8 ϩ cells than in their CD4 ϩ counterparts, both in human and in murine deficiencies (3-7). Three other observations suggested the existence of CD8 ϩ cell-specific defects in human CD3␥ deficiency: first, the proband died after a viral infection (a cytolytic T-celldependent function) despite normal antibody responses (helper T-cell-dependent) (8...

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Section: Deficient Cd8 ϩ T Lymphocytes Was Conformationally Differentmentioning

confidence: 54%

Section: Methodsmentioning

confidence: 99%

“…Calcium Flux Measurement-Intracellular Ca 2ϩ release was induced in PBLs loaded with the fluorescent dye Fluo3AM (Sigma) according to a standard procedure (12).…”

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Deficient Cd8 ϩ T Lymphocytes Was Conformationally Differentmentioning

confidence: 99%

See 3 more Smart Citations

Conformational and Biochemical Differences in the TCR·CD3 Complex of CD8+ Versus CD4+ Mature Lymphocytes Revealed in the Absence of CD3γ

Zapata¹,

Pacheco‐Castro²,

Torres³

et al. 1999

Journal of Biological Chemistry

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Assembly of the TCR/CD3 complex: CD3ε/δ and CD3ε/γ dimers associate indistinctly with both TCRα and TCRβ chains. Evidence for a double TCR heterodimer model

et al. 1998

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The TCR/CD3 complex is composed of six subunits which are expressed on the cell surface in a coordinate fashion after assembly in the endoplasmic reticulum (ER). The TCR/CD3 complex is assembled after a series of pairwise interactions involving the formation of dimers of CD3 epsilon with either CD3 gamma or CD3 delta. These dimers assemble with TCR alpha and TCR beta chains, and finally, the CD3 zeta homodimer is added to allow export of the full complex from the ER. A model has been proposed suggesting that during assembly the CD3 epsilon/CD3 gamma dimer interacts exclusively with TCR beta and the CD3 epsilon/CD3 delta dimer with TCR alpha to form a complex with a single TCR alpha/beta heterodimer. We show in this study, by immunoprecipitation and two-dimensional gel electrophoresis, that in the human T cell line Jurkat as well as in total human thymocytes, this preferential interaction does not occur and instead, the CD3 epsilon/CD3 gamma and CD3 epsilon/CD3 delta dimers associate with both TCR chains simultaneously and indistinctly. These data are confirmed by the analysis of the TCR alpha-negative T cell line MOLT-4 in which TCR beta is found separately associated with CD3 epsilon/CD3 gamma and with CD3 epsilon/CD3 delta dimers. Indirectly, our results support a model of stoichiometry in which two TCR alpha/beta heterodimers are present in a TCR/CD3 complex. Furthermore, immunoprecipitation with anti-CD3 gamma and anti-CD3 delta antibodies from 1% NP40 and 1% Brij96 cell lysates showed that these subunits form independent partial complexes which are cross-linked through the CD3 zeta homodimer. This suggests that CD3 zeta mediates the interaction between both TCR alpha/beta heterodimers contained in the double TCR complex. Further proof for this hypothesis is obtained after analysis of a Jurkat cell transfectant containing a point mutation in the transmembrane domain of TCR beta that impairs the association of CD3 zeta. In this mutant cell line, unlike a control line with wild-type TCR beta, the CD3 gamma- and CD3 delta-containing complexes were found completely independent. Altogether, these results support a model of TCR/CD3 assembly and stoichiometry in which two TCR-alpha/beta heterodimers form two hemicomplexes containing either CD3 epsilon/gamma or CD3 epsilon/delta dimers which become associated via the CD3 zeta homodimer.

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The Immunocompromised Host

Fischer

2010

Topley &Amp; Wilson's Microbiology and Microbial Infections

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Expression and function of a variant T cell receptor complex lacking CD3-gamma.

Cited by 69 publications

References 27 publications

Conformational and Biochemical Differences in the TCR·CD3 Complex of CD8+ Versus CD4+ Mature Lymphocytes Revealed in the Absence of CD3γ

Conformational and Biochemical Differences in the TCR·CD3 Complex of CD8+ Versus CD4+ Mature Lymphocytes Revealed in the Absence of CD3γ

Assembly of the TCR/CD3 complex: CD3ε/δ and CD3ε/γ dimers associate indistinctly with both TCRα and TCRβ chains. Evidence for a double TCR heterodimer model

The Immunocompromised Host

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