Targeted Expression of Human CD1d in Transgenic Mice Reveals Independent Roles for Thymocytes and Thymic APCs in Positive and Negative Selection of Vα14i NKT Cells

Mouse natural killer T (NKT) cells with an invariant Vα14-Jα18 rearrangement (Vα14 invariant [Vα14i] NKT cells) are either CD4+CD8− or CD4−CD8−. Because transgenic mice with forced CD8 expression in all T cells exhibited a profound NKT cell deficit, the absence of CD8 has been attributed to negative selection. We now present evidence that CD8 does not serve as a coreceptor for CD1d recognition and that the defect in development in CD8 transgene homozygous mice is the result of a reduction in secondary T cell receptor α rearrangements. Thymocytes from mice hemizygous for the CD8 transgene have a less severe rearrangement defect and have functional CD8+ Vα14i NKT cells. Furthermore, we demonstrate that the transcription factor Th, Poxviruses and Zinc finger, and Krüppel family (Th-POK) is expressed by Vα14i NKT cells throughout their differentiation and is necessary both to silence CD8 expression and for the functional maturity of Vα14i NKT cells. We therefore suggest that Th-POK expression is required for the normal development of Vα14i NKT cells and that the absence of CD8 expression by these cells is a by-product of such expression, as opposed to the result of negative selection of CD8-expressing Vα14i NKT cells.

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CD1d-expressing Dendritic Cells but Not Thymic Epithelial Cells Can Mediate Negative Selection of NKT Cells

Journal of Experimental Medicine ◽

10.1084/jem.20021366 ◽

2003 ◽

Vol 197 (7) ◽

pp. 907-918 ◽

Cited By ~ 107

Author(s):

Taehoon Chun ◽

Michael J. Page ◽

Laurent Gapin ◽

Jennifer L. Matsuda ◽

Honglin Xu ◽

...

Keyword(s):

Dendritic Cells ◽

Epithelial Cells ◽

Negative Selection ◽

Nkt Cells ◽

Thymic Epithelial Cells ◽

High Avidity ◽

Fetal Thymic Organ Culture ◽

Self Antigen ◽

Dose Dependent ◽

Selection Of

Natural killer T (NKT) cells are a unique immunoregulatory T cell population that is positively selected by CD1d-expressing thymocytes. Previous studies have shown that NKT cells exhibit autoreactivity, which raises the question of whether they are subject to negative selection. Here, we report that the addition of agonist glycolipid α-galactosylceramide (α-GalCer) to a fetal thymic organ culture (FTOC) induces a dose-dependent disappearance of NKT cells, suggesting that NKT cells are susceptible to negative selection. Overexpression of CD1d in transgenic (Tg) mice results in reduced numbers of NKT cells, and the residual NKT cells in CD1d-Tg mice exhibit both an altered Vβ usage and a reduced sensitivity to antigen. Furthermore, bone marrow (BM) chimeras between Tg and WT mice reveal that CD1d-expressing BM-derived dendritic cells, but not thymic epithelial cells, mediate the efficient negative selection of NKT cells. Thus, our data suggest that NKT cells developmentally undergo negative selection when engaged by high-avidity antigen or abundant self-antigen.

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

Journal of Experimental Medicine ◽

10.1084/jem.179.1.145 ◽

1994 ◽

Vol 179 (1) ◽

pp. 145-153 ◽

Cited By ~ 59

Author(s):

W Tao ◽

S J Teh ◽

I Melhado ◽

F Jirik ◽

S J Korsmeyer ◽

...

Keyword(s):

Cell Death ◽

T Cell ◽

Transgenic Mice ◽

Positive Selection ◽

T Cell Receptor ◽

Negative Selection ◽

Cell Receptor ◽

Target Cells ◽

Cell Death Pathway ◽

Selection Of

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 an antigen receptor on T cells in transgenic mice

Nature ◽

10.1038/336073a0 ◽

1988 ◽

Vol 336 (6194) ◽

pp. 73-76 ◽

Cited By ~ 487

Author(s):

William C. Sha ◽

Christopher A. Nelson ◽

Rodney D. Newberry ◽

David M. Kranz ◽

John H. Russell ◽

...

Keyword(s):

T Cells ◽

Transgenic Mice ◽

Negative Selection ◽

Antigen Receptor ◽

Selection Of

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In vitro negative selection of viral superantigen-reactive TCR transgenic thymocytes by thymic dendritic cells

Immunology Letters ◽

10.1016/s0165-2478(97)87831-5 ◽

1997 ◽

Vol 56 (1-3) ◽

pp. 249

Author(s):

I Ferrero

Keyword(s):

Dendritic Cells ◽

Negative Selection ◽

Selection Of

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A Role for Fas in Negative Selection of Thymocytes In Vivo

Journal of Experimental Medicine ◽

10.1084/jem.187.9.1427 ◽

1998 ◽

Vol 187 (9) ◽

pp. 1427-1438 ◽

Cited By ~ 112

Author(s):

Hidehiro Kishimoto ◽

Charles D. Surh ◽

Jonathan Sprent

Keyword(s):

Negative Selection ◽

Cell Receptor ◽

Staphylococcal Enterotoxin B ◽

Thymic Medulla ◽

Enterotoxin B ◽

High Doses ◽

Specific Peptide ◽

Selection Of

To seek information on the role of Fas in negative selection, we examined subsets of thymocytes from normal neonatal mice versus Fas-deficient lpr/lpr mice injected with graded doses of antigen. In normal mice, injection of 1–100 μg of staphylococcal enterotoxin B (SEB) induced clonal elimination of SEB-reactive Vβ8+ cells at the level of the semi-mature population of HSAhi CD4+ 8− cells found in the thymic medulla; deletion of CD4+ 8+ cells was minimal. SEB injection also caused marked elimination of Vβ8+ HSAhi CD4+ 8− thymocytes in lpr/lpr mice. Paradoxically, however, elimination of these cells in lpr/lpr mice was induced by low-to-moderate doses of SEB (≤1 μg) but not by high doses (100 μg). Similar findings applied when T cell receptor transgenic mice were injected with specific peptide. These findings suggest that clonal elimination of semi-mature medullary T cells is Fas independent at low doses of antigen but Fas dependent at high doses. Previous reports documenting that negative selection is not obviously impaired in lpr/lpr mice could thus reflect that the antigens studied were expressed at only a low level.

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