Extended Data Fig. 4: Expression of I-A^g7-PD induces negative selection of tetramer^bright IA^g7-ChgA_HIP reactive thymocytes at the DP to SP transition.

(a) Flow cytometric analysis and quantification of IA^g7-ChgA_HIP tetramer staining of CD4SP thymocytes following tetramer-based enrichment, isolated from I-A^g7-WT/WT (n = 6), I-A^g7-PD/WT (n = 6) mice or BDC2.5 TCR Tg on an IA^g7-WT/WT genetic background (n = 7), or polyclonal CD4SP thymocyte without enrichment. (b) Mean fluorescence intensity (MFI) of tetramer^bright and tetramer^dim populations of CD4SP thymocytes for each mouse strain. (c–h) BDC2.5 TCR Tg thymocytes undergo negative selection at the DP to CD4SP transition. (c–h) Flow cytometric analysis of CD4 and CD8⁻expressing thymocyte subsets from 6 weeks old BDC2.5 TCR Tg mice on a I-A^g7-WT/WT (n = 7), I-A^g7-PD/WT (n = 7) or I-A^g7-PD/PD (n = 7) genetic backgrounds, and (d) quantification of CD4SP thymocyte frequency. (e) Representative examples and (f) quantification of CD4⁺CD8⁺ thymocytes expressing CD69 and high levels of TCRβ. (g) Representative examples and (h) quantification of frequency at which BDC2.5 TCR Tg mice matured thymocytes into T cells, based on expression of MHCI and high levels of TCRb. P values are from a one-way ANOVA with Tukey’s multiple comparisons test. n = biologically independent animals.