Fig. 3: Cross-reactivity to an endogenous retrovirus epitope leads to negative selection of MolT-II CD4 T cells in the thymus.
From: Dysfunctional CD4 T cells in an oncovirus-specific TCR-transgenic in vivo model
A Expression of CD62L and CD44 on CD4 T cells from the spleens of C57BL/6, OT-II, and MolT-II mice. Representative dot plots showing the data from n = 5 per strain. B Representative dot plots from flow cytometric analysis of mouse thymocytes. Plots show the expression of CD4 (X axis) and CD8 (Y axis) on thymocytes from C57BL/6, OT-II and MolT-II mice (n = 5 per strain). C Comparison of the amino acid sequences of the exogenous Moloney MuLV-derived envH119-137 epitope and the envelope protein of the endogenous retrovirus AKV (epitope AKVenv138-156). D Expression of AKV transcripts encoding the AKVenv epitope in thymic stromal cell subpopulations relative to the expression of beta actin and normalized to mTECs, as determined by quantitative PCR. qPCR was performed for AKVenv and housekeeping gene beta actin on flow-sorted cortical thymic epithelial cells (cTECs), medullary thymic epithelial cells (mTECs), CD11c+F4/80- thymic dendritic cells (DCs) as well as the EL-4 thymoma cell line (positive control) and water (negative control). Columns show the relative expression (2-ΔΔCt) of AKVenv, ND = not detected, data pooled from n = 3 independent experiments. E Proliferation of MolT-II CD4 T cells without stimulation or after stimulation with peptides envH, AKVenv or ovaH. Proliferation is shown as percent CFSE-negative CD4 T cells (n = 4 for ‘unstimulated’, n = 5 for ‘envH’, ‘AKVenv’ and ‘ovaH’; unstimulated vs. AKVenv p = 0.0056, AKVenv vs. ovaH p = 0.0142, envH vs. AKVenv p = 0.3078). F Frequency of CD40L+TNF+ MolT-II CD4 T cells after co-incubation with mature D1 cells loaded with peptides envH, AKVenv, ovaH or without peptide (unstimulated, n = 9 per group, representative of 3 independent experiments; unstimulated vs. AKVenv p < 0.0001, envH vs. AKVenv p < 0.0001, AKVenv vs. ovaH p < 0.0001). Graphs show mean and SD, statistical significance (* p ≤ 0.05; ** p ≤ 0.01; *** p ≤ 0.001; **** p ≤ 0.0001; or ns for not significant) was determined by one-way ANOVA and Tukey’s multiple comparisons test (E, F).
