Calcineurin is required for positive but not negative selection of developing T cells in the thymus.
According to new research published in Immunity, calcineurin is required for positive but not negative selection of developing T cells in the thymus.
Before T cells emigrate from the thymus, they undergo a selection process, which occurs at the CD4+CD8+ double-positive (DP) stage of thymocyte development (before the downregulation of either CD4 or CD8 expression). It is thought that only those T cells able to engage in transient interactions with self-MHC molecules are maintained (positive selection) and that those T cells displaying strong reactivity are eliminated (negative selection). But what is the mechanism by which low-intensity interactions result in cell survival and proliferation and high-intensity interactions result in cell death?
Neilson and colleagues set out to explore this by examining the role of calcineurin (which activates Nfatc subunits) in thymic selection. The authors generated mice with a targeted deletion of the regulatory subunit of calcineurin (Cnb1), rendering all isoforms of calcineurin inactive in thymocytes but not non-lymphoid components of the thymus. The thymi of these mice were found to have essentially no single-positive (SP) T cells. This complete failure to progress from the DP to the SP stage indicates that calcineurin is required for positive selection. DP thymocytes were also unable to upregulate the expression of cell-surface markers associated with positive selection, such as the -chain of the T-cell receptor (TCR-), CD69 and CD5. Furthermore, positive selection of cells expressing a clonotypic TCR was blocked in the absence of calcineurin.
Next, the authors investigated the impact of calcineurin on negative selection. Negative selection of H-Y TCR-transgenic, Cnb1-deficient thymocytes was shown to occur normally in male mice, which express the male-specific H-Y antigen. This lack of requirement for calcineurin was confirmed using several in vitro assays. When Cnb1-deficient thymocytes were co-cultured with control thymocytes (from wild-type littermates) in the presence of antibodies specific for both CD3 and CD28, both cell populations displayed similar levels of DP cell death. Similar results were obtained (using a more physiological model) when moth cytochrome C peptide–I-Ek complexes were added to co-cultures of Cnb1-deficient and control thymocytes transgenic for the cognate TCR, 5C.C7. Moreover, Cnb1-deficient thymocytes had the same threshold for negative selection as wild-type thymocytes and expressed similar levels of the pro-apoptotic protein Bim (which is required for negative selection) and the anti-apoptotic protein Bcl-XL.
Calcineurin and another signal transducer, extracellular-signal-related kinase (Erk), are known to be activated by both positively and negatively selecting signals, although neither of these is required for negative selection. So the authors suggest that the lowest molecule which is common to the signalling pathways of both positive and negative selection must be upstream of calcineurin and Erk (and also Bim) and downstream of TCR proximal signalling molecules, which are clearly required for TCR signalling. They propose a model whereby both low- and high-intensity signals activate the calcineurin and Erk pathways, but high-intensity signals cause negative selection by activating a dominant pathway, which induces apoptosis through Bcl-2 family members (including Bim).
Davina Dadley-Moore
References
Neilson, J. R. et al. Calcineurin B1 is essential for positive but not negative selection during thymic development. Immunity20, 255–266 (2004) | Article | PubMed |
Palmer, E. Negative selection — clearing out the bad apples from the T-cell repertoire. Nature Rev. Immunol.3, 383–391 (2003) | Article | PubMed |
-chain of the T-cell receptor (TCR-