The T cell receptor repertoire of CD4-8+ thymocytes is altered by overexpression of the BCL-2 protooncogene in the thymus.

The bcl-2 gene encodes an intracellular, membrane-associated protein that protects immature cortical thymocytes from a wide variety of apoptotic stimuli, including glucocorticoids, radiation, and anti-CD3 treatment. Since cortical thymocytes are the primary target cells for thymic positive and negative selection processes, and since these processes are associated with cell death, we evaluated the role of bcl-2 in T cell development in two ways. In the first approach, transgenic mice expressing high levels of Bcl-2 in cortical thymocytes were mated with H-Y T cell receptor (TCR) transgenic mice, the latter being a well-defined system for the study of positive and negative selection of T cells. We found that the bcl-2 transgene had a dramatic effect on positive selection. This was manifested by a greatly increased production of mature thymocytes that were highly skewed towards the CD4-8+ lineage. The change involving CD4-8+ thymocytes occurred not only in bcl-2 transgenic mice, but was also observed in H-Y TCR/bcl-2 doubly transgenic mice, regardless of whether the H-Y TCR was expressed in the selecting (H-2b) or nonselecting (H-2d) environments. Furthermore, a large proportion of CD4-8+ thymocytes produced in H-2b H-Y TCR/bcl-2 doubly transgenic female mice expressed endogenous TCR alpha chains rather than the transgenic TCR alpha chain. These observations are consistent with the model that high expression of Bcl-2 in cortical thymocytes overrides the normal apoptotic pathway. This then allows the selection of CD4-8+ thymocytes expressing TCRs that are otherwise nonselectable. However, the bcl-2 transgene did not protect CD4+8+ thymocytes expressing the male-specific TCR from deletion in male doubly transgenic mice. In the second approach, we determined the level of bcl-2 mRNA expression in populations of thymocytes defined by their CD4/CD8 phenotypes using quantitative reversed transcriptase PCR techniques. Our results indicate that bcl-2 mRNA was expressed at a high level in immature CD4-8- thymocytes and in mature CD4+8- thymocytes. There is a dramatic downregulation of bcl-2 mRNA in CD4+8+ thymocytes, particularly those expressing a low level of TCR. CD4+8+ thymocytes that upregulated their TCR, likely as a result of receiving positive selection signals, also upregulated bcl-2 mRNA. This observation suggests that rescue of immature thymocytes from the programmed cell death pathway by positive selection signals is accompanied by the upregulation of bcl-2 mRNA.

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Positive and negative selection of T cells in T-cell receptor transgenic mice expressing a bcl-2 transgene.

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.91.4.1376 ◽

1994 ◽

Vol 91 (4) ◽

pp. 1376-1380 ◽

Cited By ~ 99

Author(s):

A. Strasser ◽

A. W. Harris ◽

H. von Boehmer ◽

S. Cory

Keyword(s):

T Cells ◽

T Cell ◽

Transgenic Mice ◽

T Cell Receptor ◽

Negative Selection ◽

Cell Receptor ◽

Selection Of

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Early Deletion and Late Positive Selection of T-Cells Expressing a Male-Specific Receptor in T-Cell Receptor Transgenic Mice

Developmental Immunology ◽

10.1155/1990/18208 ◽

1990 ◽

Vol 1 (1) ◽

pp. 1-10 ◽

Cited By ~ 52

Author(s):

Hung Sia Teh ◽

Hiroyuki Kishi ◽

Bernadette Scott ◽

Peter Borgulya ◽

Harald Von Boehmer ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Transgenic Mice ◽

Positive Selection ◽

T Cell Receptor ◽

Interleukin 2 ◽

Cell Receptor ◽

Early Expression ◽

Single Positive ◽

Selection Of

The ontogeny of T cells in T-cell receptor (TCR) transgenic mice, which express a transgenicαβheterodimer, specific for the male (H-Y) antigen in association with H-2Db, was determined. The transgenicαchain was expressed on about 10% of the fetal thymocytes on day 14 of gestation. About 50% of day-15 fetal thymocytes expressed bothαandβtranschains and virtually all fetal thymocytes expressed the transgenicαβheterodimer by day 17. The early expression of the transgenic TCR on CD4-8-thymocytes prevented the development ofγδcells, and led to accelerated growth of thymocytes and an earlier expression of CD4 and CD8 molecules. Up to day 17, no significant differences in T-cell development could be detected between female and male thymuses. By day 18 of gestation, the male transgenic thymus contained more CD4-8-thymocytes than the female transgenic thymus. The preponderance of CD4-8-thymocytes in the male transgenic thymus increased until birth and was a consequence of the deletion of the CD4+8+thymocytes and their CD4-8+precursors. By the time of birth, the male transgenic thymus contained half the number of cells as the female transgenic thymus. The deletion of autospecific precursor cells in the male transgenic mouse began only at day 18 of gestation, despite the fact that the ligand could already be detected by day 16.The preferential accumulation of CD4-8+T cells, which expressed a high density of the transgenic TCR, occurred only after birth and was .obvious in 6-week-old female thymus. These data support the hypothesis that the positive selection of T cells expressing this transgenic heterodimer may involve two steps, i.e., the commitment of CD4+8+thymocytes to the CD4-8+lineage following the interaction of the transgenic TCR with restricting major histocompatibility molecules, followed by a slow conversion of CD4+8+thymocytes into CD4-8+T cells.In normal mice, the precursors of CD+4+8 and single positive thymocytes have the CD4-8-CD3-J11d+(or M1/69+) phenotype. Because of the early expression of the transgenicαβheterodimer, this population was not detected in adult transgenic mice. All CD4-8-M1/ 69+cells expressed the transgenic receptor associated with CD3 and could be readily grown in media containing T-cell lectins and interleukin 2.

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Increased p300 Expression Inhibits Glucocorticoid Receptor-T-Cell Receptor Antagonism but Does Not Affect Thymocyte Positive Selection

Molecular and Cellular Biology ◽

10.1128/mcb.22.13.4556-4566.2002 ◽

2002 ◽

Vol 22 (13) ◽

pp. 4556-4566 ◽

Cited By ~ 7

Author(s):

Cheng-Tai Yu ◽

Ming-Hsien Lin Feng ◽

Hsiu-ming Shih ◽

Ming-Zong Lai

Keyword(s):

T Cells ◽

T Cell ◽

Transgenic Mice ◽

Glucocorticoid Receptor ◽

Positive Selection ◽

T Cell Receptor ◽

Critical Role ◽

Interleukin 2 ◽

Cell Receptor ◽

Selection Of

ABSTRACT Positive selection of T cells is postulated to be dependent on the counterinteraction between glucocorticoid receptor (GR)- and T-cell-receptor (TCR)-induced death signals. In this study we used T-cell-specific expression of p300 to investigate whether GR-TCR cross talk between thymocytes was affected. Activation of the p300-transgenic T cells led to enhanced thymocyte proliferation and increased interleukin 2 production. Thymocyte death, induced by TCR engagement, was no longer prevented by dexamethasone in p300-transgenic mice, indicating an absence of GR-TCR cross-inhibition. This was accompanied by a 50% reduction in the number of thymocytes in p300-transgenic mice. However, the CD4/CD8 profile of thymocytes remained unchanged in p300-transgenic mice. There was no effect on positive selection of the bulk thymocytes or thymocytes with transgenic TCR in p300-transgenic mice. In addition, there was no apparent TCR repertoire “hole” in the selected antigens examined. Our results illustrate a critical role of CBP/p300 in thymic GR-TCR counterinteraction yet do not support the involvement of GR-TCR antagonism in thymocyte positive selection.

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T cell receptor-mediated signaling events in CD4+CD8+ thymocytes undergoing thymic selection: requirement of calcineurin activation for thymic positive selection but not negative selection.

Journal of Experimental Medicine ◽

10.1084/jem.181.3.927 ◽

1995 ◽

Vol 181 (3) ◽

pp. 927-941 ◽

Cited By ~ 70

Author(s):

C R Wang ◽

K Hashimoto ◽

S Kubo ◽

T Yokochi ◽

M Kubo ◽

...

Keyword(s):

T Cell ◽

Positive Selection ◽

T Cell Receptor ◽

Intracellular Signaling ◽

Negative Selection ◽

Cell Receptor ◽

Clonal Deletion ◽

Signaling Events ◽

Selection Of

The goal of this study was to identify the differences of intracellular signals between the processes of thymic positive and negative selection. The activation of calcineurin, a calcium- and calmodulin-dependent phosphatase, is known to be an essential event in T cell activation via the T cell receptor (TCR). The effect of FK506, an inhibitor of calcineurin activation, on positive and negative selection in CD4+CD8+ double positive (DP) thymocytes was examined in normal mice and in a TCR transgenic mouse model. In vivo FK506 treatment blocked the generation of mature TCRhighCD4+CD8- and TCRhighCD4-CD8+ thymocytes, and the induction of CD69 expression on DP thymocytes. In addition, the shutdown of recombination activating gene 1 (RAG-1) transcription and the downregulation of CD4 and CD8 expression were inhibited by FK506 treatment suggesting that the activation of calcineurin is required for the first step (or the very early intracellular signaling events) of TCR-mediated positive selection of DP thymocytes. In contrast, FK506-sensitive calcineurin activation did not appear to be required for negative selection based on the observations that negative selection of TCR alpha beta T cells in the H-2b male thymus (a negative selecting environment) was not inhibited by in vivo treatment with FK506 and that there was no rescue of the endogenous superantigen-mediated clonal deletion of V beta 6 and V beta 11 thymocytes in FK506-treated CBA/J mice. DNA fragmentation induced by TCR activation of DP thymocytes in vitro was not affected by FK506. In addition, different effects of FK506 from Cyclosporin A on the T cell development in the thymus were demonstrated. The results of this study suggest that different signaling pathways work in positive and negative selection and that there is a differential dependence on calcineurin activation in the selection processes.

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Defective T-cell receptor signalling and positive selection of Vav-deficient CD4+CDS+thymocytes

Nature ◽

10.1038/374474a0 ◽

1995 ◽

Vol 374 (6521) ◽

pp. 474-476 ◽

Cited By ~ 232

Author(s):

Klaus-Dieter Fischer ◽

Antanina Zmuidzinas ◽

Sandra Gardner ◽

Mariano Barbacid ◽

Alan Bernstein ◽

...

Keyword(s):

T Cell ◽

Positive Selection ◽

T Cell Receptor ◽

Cell Receptor ◽

Receptor Signalling ◽

Selection Of

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A Kinetic Threshold between Negative and Positive Selection Based on the Longevity of the T Cell Receptor–Ligand Complex

Journal of Experimental Medicine ◽

10.1084/jem.189.10.1531 ◽

1999 ◽

Vol 189 (10) ◽

pp. 1531-1544 ◽

Cited By ~ 92

Author(s):

Calvin B. Williams ◽

Deborah L. Engle ◽

Gilbert J. Kersh ◽

J. Michael White ◽

Paul M. Allen

Keyword(s):

T Cells ◽

T Cell ◽

Positive Selection ◽

T Cell Receptor ◽

Cell Receptor ◽

Ligand Complex ◽

Altered Peptide Ligands ◽

Self Peptides ◽

Selection Of

We have developed a unique in vivo system to determine the relationship between endogenous altered peptide ligands and the development of major histocompatibility complex class II– restricted T cells. Our studies use the 3.L2 T cell receptor (TCR) transgenic mouse, in which T cells are specific for Hb(64–76)/I-Ek and positively selected on I-Ek plus self-peptides. To this endogenous peptide repertoire, we have individually added one of six well-characterized 3.L2 ligands. This transgenic approach expands rather than constrains the repertoire of self-peptides. We find that a broad range of ligands produce negative selection of thymocytes in vivo. When compared with the in vitro TCR–ligand binding kinetics, we find that these negatively selecting ligands all have a half-life of 2 s or greater. Additionally, one of two ligands examined with no detectable binding to the 3.L2 TCR and no activity on mature 3.L2 T cells (Q72) enhances the positive selection of transgenic thymocytes in vivo. Together, these data establish a kinetic threshold between negative and positive selection based on the longevity of TCR–ligand complexes.

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