Fig. 4: Terminal NK cell maturation block in Nr2f6-deficient mice and reduced NKp46 independent short-term effector responses.

A Nr2f6 expression values in splenic CD8 T cell subsets, naïve (n), effector (te), and central-memory (cm) on day 7 post lymphocytic choriomeningitis virus (LCMV) infection, and the different splenic natural killer (NK) cell subsets CD27⁺CD11b^-, CD27⁺CD11b⁺, and CD27^-CD11b⁺. Expression values are normalized by DESeq2 based on the Immgen consortium database [37]. B Representative dot-plots of CD27^-CD11b^-, CD27⁺CD11b^-, CD27⁺CD11b⁺ mature CD27^-CD11b⁺, and terminally matured (CD27^-CD11b⁺KLRG1⁺) splenic NK cell populations (out of CD45⁺CD3^-NK1.1⁺NKp46⁺) in wild-type (Nr2f6^+/+) or Nr2f6-deficient (Nr2f6^−/−) mice. C Quantification of frequency CD27⁺CD11b^-, CD27⁺CD11b⁺ and mature CD27^-CD11b⁺ splenic NK cells (CD45⁺CD3^-NK1.1⁺NKp46⁺) in wild-type (Nr2f6^+/+) or Nr2f6-deficient (Nr2f6^−/−) mice. D Quantification of terminally matured KLRG1⁺ NK cells (CD45⁺CD3^-NK1.1⁺NKp46⁺ CD27^-CD11b⁺) in the spleen of wild-type (Nr2f6^+/+) or Nr2f6-deficient (Nr2f6^−/−) mice. E Schematic overview of the experimental setup of LPS injection. Wild-type (Nr2f6^+/+) or Nr2f6 deficient (Nr2f6 ^−/−) mice were i.v. injected with 2 µg LPS or PBS as control and the animals were sacrificed 2 ½ hrs later. Splenocytes were isolated and incubated for another 2 ½ hrs in the presence of Brefeldin A. F Representative dot-plots of splenic NK cells (CD45⁺CD3^-NK1.1⁺NKp46⁺) IFNγ-producing NK cells in wild-type (Nr2f6^+/+) or Nr2f6-deficient (Nr2f6^−/−) mice after LPS injection. G Quantification of frequencies and total cell counts of IFNγ⁺ NK cells (CD45⁺CD3^-NK1.1⁺NKp46⁺) in wild-type (Nr2f6^+/+) or Nr2f6-deficient (Nr2f6^−/−) mice after LPS injection. H Quantification of NK1.1, NKp46, and IFNγ expression (MFI) in NK cells (CD45⁺CD3^-NK1.1⁺NKp46⁺) in wild-type (Nr2f6^+/+) or Nr2f6-deficient (Nr2f6^−/−) mice after LPS injection. B-D Representative data is shown as pooled experiments of at least two independent experiments n = 8-10. E, F For LPS injection, the representative data shown are from one independent out of two replicative experiments, with n = 4 per group and experiment. Each dot represents the data of an individual mouse. Results are shown as mean ± SD. The normality of data was evaluated by the Shapiro–Wilk test. An asterisk indicates statistically significant differences between genotypes calculated using Student’s t-test. A p value < 0.05 was considered statistically significant. *p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001.