Extended Data Figure 10. L-Myc-deficient dendritic cells process and present soluble antigens efficiently.

a, b, CD8α⁺ DCs, CD8α⁻ DCs, and pDCs were isolated by cell sorting from the spleen of WT mice (top panels) and Mycl1^gfp/gfp mice (bottom panels). OT-I CD8⁺ T cells and OT-II CD4⁺ T cells were isolated by cell sorting from the spleen of CD45.1⁺ OT-I transgenic and CD45.1⁺ OT-II transgenic mice, respectively, and T cells were then labeled with CFSE. DCs were pulsed with whole ovalbumin protein (Ova) for 2 h at 37°C, then washed extensively before co-culture with OT-I CD8⁺ T cells (a) or OT-II CD4⁺ T cells (b) at a DC:T cell ratio of 1:5. Ova_257-264 (OT-I CD8⁺ T cell epitope) and Ova_323-339 (OT-II CD4⁺ T cell epitope) were used as positive controls (panels not shown). Cells were analyzed 3 days later for CFSE dilution. Shown are single-color histograms of CFSE for pre-gated live T cells. Data are representative of 2 independent experiments. c, WT mice (black dots) and Mycl1^gfp/gfp mice (green dots) were infected with 10⁵ L. monocytogenes expressing ovalbumin (LM-OVA). After 24 h, CD8α⁺ DCs and CD8α⁻ DCs were purified from infected spleens and co-cultured with CFSE-labeled OT-I CD45.1⁺ CD8⁺ T cells. Cells were analyzed 60 h later for CFSE dilution. Shown are single-color histograms of CFSE for live OT-I T cells gated as CD45.1⁺CD8α⁺. d, Shown is the quantitation (described in Methods) of live OT-I CD8⁺ T cells from (c) after co-culture with the indicated DCs from WT (black dots) and Mycl1^gfp/gfp (green dots). Data are from 2 independent experiments (bar, SD; n = 3 biological replicates, Student's t-test). e, Histopathology (H&E) of spleens (top panels) and livers (bottom panels) from WT (left panels), Mycl1^gfp/gfp (middle panels), and Batf3^−/− (right panels) mice 3 days after infection (10⁵ L. monocytogenes i.v.). Scale bars 200 μm. *, p<0.01; ns, p>0.05.