Germ-line elimination of electric charge on pre-T-cell receptor (TCR) impairs autonomous signaling for β-selection and TCR repertoire formation

摘要

The pre-T-cell receptor (TCR) is crucial for the early T-cell development, but the ligand for pre-TCR remains unidentified. We recently proposed a model that pre-TCR complexes oligomerize spontaneously through interactions of the pre-TCR_α chain. To investigate the mechanism underlying this ligand-independent signaling in vivo, we established knock-in mice that express a pre-TCR_α mutant lacking charged amino acids (D~(22)R~(24)R~(102)R~(117) to A~(22)A~(24)A~(102)A~(117); 4A). CD4~+CD8~+ thymocyte number was significantly reduced in invariant pre-TCRa (pT_α~(4A/4A)) mice, whereas CD4~-CD8~- thymocytes were unaffected. The percentages of double-negative 3 (DN3) cells and γδ T cells were increased in the pTa~(4A/4A) thymus, indicating that p-selection is impaired in pTa~(4A/4A) mice. Pre-TCR-mediated tyro-sine phosphorylation and clonal expansion into double-positive thymocytes were also defective in the knock-in mice. Pre-TCR was expressed at higher levels on pToα~(4A/4A) cell surfaces than on those of the wild type, suggesting that the charged residues in pTa are critical - for autonomous engagement and subsequent internalization of pre-TCR. Pre-TCR-mediated allelic exclusion of the TCRβ gene was also inhibited in pTa~(4A/4A) mice, and thereby, dual TCRβs were expressed on pT_α~(4A/4A)T cells. Furthermore, the TCRp chain variable region (Vβ) repertoire of mature T cells was significantly altered in pTa~(4A/4A) mice. These results suggest that charged residues of pTα are critical for p-selection, allelic exclusion, and TCRp repertoire formation.