Reduced Generation but Efficient TCRβ-Chain Selection of CD4+8+ Double-Positive Thymocytes in Mice with Compromised CD3 Complex Signaling

Abstract

Abstract Maturation to the CD4+8+ double-positive (DP) stage of thymocyte development is restricted to cells that have passed TCRβ selection, an important checkpoint at which immature CD4−8− double-negative (DN) cells that express TCRβ polypeptide chains are selected for further maturation. The generation of DP thymocytes following TCRβ selection is dependent on cellular survival, differentiation, and proliferation, and the entire process appears to be mediated by the pre-TCR/CD3 complex. In this study, we investigate the signaling requirements for TCRβ selection using mice single deficient and double deficient for CD3ζ/η and/or p56lck. While the numbers of DP cells are strongly reduced in the single-deficient mice, a further drastic reduction in the generation of DP thymocytes is seen in the double-deficient mice. The poor generation of DP cells in the mutant mice is primarily due to an impaired ability of CD25+ DN thymocytes to proliferate following expression of a TCRβ-chain. Nevertheless, the residual DP cells in all mutant mice are strictly selected for expression of TCRβ polypeptide chains. DN thymocytes of mutant mice expressed TCRβ and CD3ε at the cell surface and contained mRNA for pre-Tα, but not for clonotypic TCRα-chains, together suggesting that TCRβ selection is mediated by pre-TCR signaling in all cases. The data suggest differential requirements of pre-TCR signaling for cell survival on the one hand, and for the proliferative burst associated with TCRβ selection on the other.