Fig. 6

mTORC2 restrains NK cell effector functions by inhibiting mTORC1 activity. a, b The expression of perforin and granzyme B (a), CD71, and uptake of 2-NBDG (b) in total splenic NK cells (CD3⁻CD19⁻NK1.1⁺NKp46⁺) and NK cell subsets from mice of the indicated genotypes. c Flow cytometric analysis of CD107a⁺ cells among total splenic NK cells (CD3⁻CD19⁻NK1.1⁺NKp46⁺) or the NK cell subsets specified from mice of the indicated genotypes after coculture with YAC-1 targets. d The expression of phosphorylated (p-) S6 at ser235/236 (p-S6^ser235/236) in splenic NK cells (CD3⁻CD19⁻NK1.1⁺) from control versus Rictor^∆/∆ mice after stimulation with rapamycin of the indicated concentrations for 6 h. The MFI of p-S6 ^ser235/236 at each concentration was normalized to the baseline reading (i.e., 0 nm rapamycin-treated control NK cells in a littermate pair). The horizontal dashed line indicates the normalized baseline reading. e–f The expression of perforin, granzyme B, uptake of 2-NBDG (e), and CD71 and CD98 (f) (MFI relative to the littermates of the control + DMSO group) in splenic NK cells from control versus Rictor^∆/∆ mice treated with DMSO (set as the solvent control) or 0.1 nM rapamycin for 6 h ex vivo. g Flow cytometric analysis of CD107a⁺ splenic NK cells from control versus Rictor^∆/∆ mice following treatment with DMSO or 0.1 nM rapamycin, described in detail in the ONLINE METHODS. The control represents Rictor^fl/+/Ncr1-Cre⁺ mice; Rictor^∆NK represents Rictor^fl/fl/Ncr1-Cre⁺ mice; and Rictor^∆NKRaptor^fl/⁺ represents Rictor^fl/fl/Rptor^fl/+/Ncr1-Cre⁺ mice. The dashed line indicates the control group, and the solid line indicates the gene knockout group. The MFI was calculated relative to total NK cells or to the CD27SP subset of NK cells from control littermates. The subpopulations of NK cells are distinguished by surface expression of CD27 and CD11b, and CD27SP, DP, and CD11bSP represent the CD27⁺CD11b⁻, CD27⁺CD11b⁺, and CD27⁻CD11b⁺ NK cell subsets, respectively. Each dot represents one mouse, and all experiments were replicated 3 (NK subsets in a), 4 (c, e–g), 5 (b), or 7 (total NK cells in a) times; error bars represent SD; ^∗p < 0.05, ^∗∗p < 0.01, ^∗∗∗p < 0.001, and ^∗∗∗∗p < 0.0001; one-way ANOVA (a–c, e–g). Control, n = 3; Rictor^∆/∆, n = 5 in 3 independent experiments (d)