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CD8 beta increases CD8 coreceptor function and participation in TCR-ligand binding.

Renard V, Romero P, Vivier E, Malissen B, Luescher IF - J. Exp. Med. (1996)

Bottom Line: T cell receptor (TCR) photoaffinity labeling indicated that TCR-ligand binding on CD8 alpha/beta cells was approximately 5- and 20-fold more avid than on CD8 alpha/a and CD8-/- cells, respectively.SF1-1.1.1 Fab' or Kd mutation D227K reduced the TCR photoaffinity labeling on CD8 alpha/beta cells to approximately the same low levels observed on CD8-/- cells.These results indicate that CD8 alpha/beta is a more efficient coreceptor than CD8alpha/alpha, because it more avidly strengthens TCR-ligand binding.

View Article: PubMed Central - PubMed

Affiliation: Centre d'Immunologie Institut National de la Santé et de la Recherche Médicale, Centre National de la Recherche Scientifique de Marseille-Luminy, France.

ABSTRACT
To study the role of CD8 beta in T cell function, we derived a CD8 alpha/beta-(CD8-/-) T cell hybridoma of the H-2Kd-restricted N9 cytotoxic T lymphocyte clone specific for a photoreactive derivative of the Plasmodium berghei circumsporozoite peptide PbCS 252-260. This hybridoma was transfected either with CD8 alpha alone or together with CD8 beta. All three hybridomas released interleukin 2 upon incubation with L cells expressing Kd-peptide derivative complexes, though CD8 alpha/beta cells did so more efficiently than CD8 alpha/alpha and especially CD8-/- cells. More strikingly, only CD8 alpha/beta cells were able to recognize a weak agonist peptide derivative variant. This recognition was abolished by Fab' fragments of the anti-Kd alpha 3 monoclonal antibody SF1-1.1.1 or substitution of Kd D-227 with K, both conditions known to impair CD8 coreceptor function. T cell receptor (TCR) photoaffinity labeling indicated that TCR-ligand binding on CD8 alpha/beta cells was approximately 5- and 20-fold more avid than on CD8 alpha/a and CD8-/- cells, respectively. SF1-1.1.1 Fab' or Kd mutation D227K reduced the TCR photoaffinity labeling on CD8 alpha/beta cells to approximately the same low levels observed on CD8-/- cells. These results indicate that CD8 alpha/beta is a more efficient coreceptor than CD8alpha/alpha, because it more avidly strengthens TCR-ligand binding.

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TCR photoaffinity labeling with soluble ligand Kd–peptide  derivative complexes. CD8α/β, CD8α/α or CD8−/− cells were incubated with Kd-125IASA-YIPSAEK(ABA)I in absence (black bars) or presence  of SF1-1.1.1 Fab′ (20 μg/ml) (white bars) or 20-8-4S mAb (10 μg/ml)  (hatched bars), and TCR photoaffinity labeling was evaluated by SDSPAGE and autoradiography (A) and densitometry (B). Alternatively,  CD8α/β and CD8−/− cells, respectively, were tested likewise, either with  Kd-IASA-YIPSAEK(ABA)I (black bars) or mutant KdD227K-IASA-YIPSAEK(BA)I (white bars) (C). In both experiments, 100% refers to the  TCR labeling observed on CD8α/β cells with the wild type ligand. The  TCR binding of Kd-IASA-YIPSAEK(BA)I was assessed by its ability to  inhibit the TCR photoaffinity labeling by Kd-125IASA-YIPSAEK (ABA)I  on N9 CTL (D). Mean values and standard deviations were calculated  from at least three independent experiments.
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Figure 3: TCR photoaffinity labeling with soluble ligand Kd–peptide derivative complexes. CD8α/β, CD8α/α or CD8−/− cells were incubated with Kd-125IASA-YIPSAEK(ABA)I in absence (black bars) or presence of SF1-1.1.1 Fab′ (20 μg/ml) (white bars) or 20-8-4S mAb (10 μg/ml) (hatched bars), and TCR photoaffinity labeling was evaluated by SDSPAGE and autoradiography (A) and densitometry (B). Alternatively, CD8α/β and CD8−/− cells, respectively, were tested likewise, either with Kd-IASA-YIPSAEK(ABA)I (black bars) or mutant KdD227K-IASA-YIPSAEK(BA)I (white bars) (C). In both experiments, 100% refers to the TCR labeling observed on CD8α/β cells with the wild type ligand. The TCR binding of Kd-IASA-YIPSAEK(BA)I was assessed by its ability to inhibit the TCR photoaffinity labeling by Kd-125IASA-YIPSAEK (ABA)I on N9 CTL (D). Mean values and standard deviations were calculated from at least three independent experiments.

Mentions: More strikingly, the peptide derivative variant IASAYIPSAEK(BA)I, which lacks the azido function of the ABA group, was efficiently recognized only by CD8α/β cells (Fig. 1). Half-maximal IL-2 release was observed at >10-fold higher degree of sensitization than the wild-type conjugate. Substitution of the ABA group with BA reduced the efficiency of recognition by N9 CTL by ∼50-fold (18). As assessed by TCR photoaffinity labeling (see below), Kd complexes with IASA-YIPSAEK(BA)I bound to the N9 TCR approximately seven times less efficiently than those containing IASA-YIPSAEK(ABA)I (see Fig. 3 D). The observation that this weak agonist was significantly recognized only by CD8α/β cells is in accordance with reports indicating that CD8β broadens the range of antigen recognition by CD8+ T cells (10, 11) and in addition provides a quantitative correlation between antigen recognition and TCR–ligand binding.


CD8 beta increases CD8 coreceptor function and participation in TCR-ligand binding.

Renard V, Romero P, Vivier E, Malissen B, Luescher IF - J. Exp. Med. (1996)

TCR photoaffinity labeling with soluble ligand Kd–peptide  derivative complexes. CD8α/β, CD8α/α or CD8−/− cells were incubated with Kd-125IASA-YIPSAEK(ABA)I in absence (black bars) or presence  of SF1-1.1.1 Fab′ (20 μg/ml) (white bars) or 20-8-4S mAb (10 μg/ml)  (hatched bars), and TCR photoaffinity labeling was evaluated by SDSPAGE and autoradiography (A) and densitometry (B). Alternatively,  CD8α/β and CD8−/− cells, respectively, were tested likewise, either with  Kd-IASA-YIPSAEK(ABA)I (black bars) or mutant KdD227K-IASA-YIPSAEK(BA)I (white bars) (C). In both experiments, 100% refers to the  TCR labeling observed on CD8α/β cells with the wild type ligand. The  TCR binding of Kd-IASA-YIPSAEK(BA)I was assessed by its ability to  inhibit the TCR photoaffinity labeling by Kd-125IASA-YIPSAEK (ABA)I  on N9 CTL (D). Mean values and standard deviations were calculated  from at least three independent experiments.
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Figure 3: TCR photoaffinity labeling with soluble ligand Kd–peptide derivative complexes. CD8α/β, CD8α/α or CD8−/− cells were incubated with Kd-125IASA-YIPSAEK(ABA)I in absence (black bars) or presence of SF1-1.1.1 Fab′ (20 μg/ml) (white bars) or 20-8-4S mAb (10 μg/ml) (hatched bars), and TCR photoaffinity labeling was evaluated by SDSPAGE and autoradiography (A) and densitometry (B). Alternatively, CD8α/β and CD8−/− cells, respectively, were tested likewise, either with Kd-IASA-YIPSAEK(ABA)I (black bars) or mutant KdD227K-IASA-YIPSAEK(BA)I (white bars) (C). In both experiments, 100% refers to the TCR labeling observed on CD8α/β cells with the wild type ligand. The TCR binding of Kd-IASA-YIPSAEK(BA)I was assessed by its ability to inhibit the TCR photoaffinity labeling by Kd-125IASA-YIPSAEK (ABA)I on N9 CTL (D). Mean values and standard deviations were calculated from at least three independent experiments.
Mentions: More strikingly, the peptide derivative variant IASAYIPSAEK(BA)I, which lacks the azido function of the ABA group, was efficiently recognized only by CD8α/β cells (Fig. 1). Half-maximal IL-2 release was observed at >10-fold higher degree of sensitization than the wild-type conjugate. Substitution of the ABA group with BA reduced the efficiency of recognition by N9 CTL by ∼50-fold (18). As assessed by TCR photoaffinity labeling (see below), Kd complexes with IASA-YIPSAEK(BA)I bound to the N9 TCR approximately seven times less efficiently than those containing IASA-YIPSAEK(ABA)I (see Fig. 3 D). The observation that this weak agonist was significantly recognized only by CD8α/β cells is in accordance with reports indicating that CD8β broadens the range of antigen recognition by CD8+ T cells (10, 11) and in addition provides a quantitative correlation between antigen recognition and TCR–ligand binding.

Bottom Line: T cell receptor (TCR) photoaffinity labeling indicated that TCR-ligand binding on CD8 alpha/beta cells was approximately 5- and 20-fold more avid than on CD8 alpha/a and CD8-/- cells, respectively.SF1-1.1.1 Fab' or Kd mutation D227K reduced the TCR photoaffinity labeling on CD8 alpha/beta cells to approximately the same low levels observed on CD8-/- cells.These results indicate that CD8 alpha/beta is a more efficient coreceptor than CD8alpha/alpha, because it more avidly strengthens TCR-ligand binding.

View Article: PubMed Central - PubMed

Affiliation: Centre d'Immunologie Institut National de la Santé et de la Recherche Médicale, Centre National de la Recherche Scientifique de Marseille-Luminy, France.

ABSTRACT
To study the role of CD8 beta in T cell function, we derived a CD8 alpha/beta-(CD8-/-) T cell hybridoma of the H-2Kd-restricted N9 cytotoxic T lymphocyte clone specific for a photoreactive derivative of the Plasmodium berghei circumsporozoite peptide PbCS 252-260. This hybridoma was transfected either with CD8 alpha alone or together with CD8 beta. All three hybridomas released interleukin 2 upon incubation with L cells expressing Kd-peptide derivative complexes, though CD8 alpha/beta cells did so more efficiently than CD8 alpha/alpha and especially CD8-/- cells. More strikingly, only CD8 alpha/beta cells were able to recognize a weak agonist peptide derivative variant. This recognition was abolished by Fab' fragments of the anti-Kd alpha 3 monoclonal antibody SF1-1.1.1 or substitution of Kd D-227 with K, both conditions known to impair CD8 coreceptor function. T cell receptor (TCR) photoaffinity labeling indicated that TCR-ligand binding on CD8 alpha/beta cells was approximately 5- and 20-fold more avid than on CD8 alpha/a and CD8-/- cells, respectively. SF1-1.1.1 Fab' or Kd mutation D227K reduced the TCR photoaffinity labeling on CD8 alpha/beta cells to approximately the same low levels observed on CD8-/- cells. These results indicate that CD8 alpha/beta is a more efficient coreceptor than CD8alpha/alpha, because it more avidly strengthens TCR-ligand binding.

Show MeSH
Related in: MedlinePlus