Fig. 1. Nur77-GFP^HI and Nur77-GFP^LO CD4 T cells are phenotypically and functionally distinct.

a Frequency of Nur77-GFP^HI CD4 T cells in lungs of Nur77-GFP or wildtype mice at four weeks post-infection or uninfected. Gated on live CD4⁺ CD44⁺ Nur77-GFP^HI cells. P values calculated with an unpaired two-tailed t-test, 3 mice per group. b Flow cytometry of Ag85B- or ESAT-6-specific CD4 T cells from Nur77-GFP lungs harvested at four weeks post-infection. Mice indicated received Ag85B peptide injected six hours before harvest. Gated on live CD3⁺ CD4⁺ CD44⁺ Tetramer⁺ cells. Tetramer⁺ cells represent either Ag85B or ESAT-6 specificity. Representative plots from experiment with 4 mice per group. c Flow cytometry of Nur77-GFP^LO and Nur77-GFP^HI CD4 T cells from Nur77-GFP lungs harvested at four weeks post-infection. CD45.2 fluorescent antibody injected intravenously immediately before harvest identifies vascular cells. Gated on live CD3⁺ CD4⁺ CD44⁺ and either lowest or highest 1/3 of Nur77-GFP expression. Further gated on IV^-, CX3CR1⁺ KLRG1⁺, or Tetramer⁺ cells. Tetramer⁺ cells represent either Ag85B or ESAT-6 specificity. P values calculated with a paired two tailed t-test, 4 or 5 mice per group. d Nur77-GFP^LO and Nur77-GFP^HI CD4 T cells sorted from Nur77-GFP lungs at four weeks post-infection and injected into one week post-infection TCRα^-/- mice. Donor cells sorted for live CD3⁺ CD4⁺ CD44⁺ Nur77-GFP^LO, Nur77-GFP^HI, or total bulk CD4 T cells. e Curves show survival of recipient TCRα-/- mice. P value calculated with a two-sided Mantel-Cox test, 4-6 mice per group. f–h Nur77-GFP^LO and Nur77-GFP^HI CD4 T cells sorted from Nur77-GFP lungs at four weeks post-infection and injected into infection-matched wildtype CD45.1 mice. Donor cells sorted on live CD4⁺ CD44⁺ Nur77-GFP^LO or Nur77-GFP^HI cells. Recipient and no transfer wildtype CD45.1 lungs harvested at one week post-transfer for flow cytometry and lung bacterial load. g Number and lung localization of donor and recipient cells at one-week post-transfer. CD45.2 fluorescent antibody injected immediately before harvest identifies vascular cells. Gated on live CD4⁺ CD44⁺ cells. Cell numbers assessed using counting beads. P value calculated with an unpaired two tailed t-test. Mice per group: 4 for Nur77-GFP^LO recipients or 3 for Nur77-GFP^HI. h Lung bacterial load at one-week post-transfer or without transfer. Pre-transfer wildtype CD45.1 lungs harvested at four weeks post-infection. P values calculated with an unpaired two-tailed t-test. Mice per group: 5 for pre-transfer, 4 for no transfer, 4 for Nur77-GFP^LO recipients or 3 for Nur77-GFP^HI. Error bars indicate standard deviation.