Extended Data Fig. 3: Iron depletion impairs mitochondrial function and switches OXPHOS to glycolysis in ILC3s (Related to Fig. 3).

a, PCA of RNA-seq analysis of MNK-3 cells treated with or without DFO for 16 h in vitro (n = 3 biologically independent replicates per group). b–g, Mitochondrial mass and activity in MNK-3 cells. MitoTracker Green FM (b), TMRE (d) and MitoSox (f) intensity. Representative FACS plots of three independent experiments. gMFI of MitoTracker Green FM (c), TMRE (e) and MitoSox (g) (n = 6 biologically independent replicates per group). Data are compiled from three independent experiments. h–j, Mitochondrial mass and activity of ILC3s from WT mice fed an ID or Ctrl diet. gMFI of MitoTracker Green FM (h), TMRE (i) and MitoSox (j) in ILC3s (control diet: n = 7; ID diet: n = 5). Data are compiled from three independent experiments. k, FACS analysis of 2-NBDG uptake in MNK-3 cells. Representative FACS plot of three independent experiments. l–n, Seahorse metabolic flux analysis. Real-time ECAR (l) are representative of two independent experiments, and compiled data on quantification of basal (m) and maximal (n) ECAR of one experiment are shown (n = 3 biologically independent replicates per group). o and p, FACS analysis of IL-22 and IL-17A expression in MNK-3 cells transduced with the indicated retroviral constructs. The cells were treated with FICZ on day 2. Representative FACS plot (o) and percentages of IL-22⁺ cells in MNK-3 cells from two independent experiments (n = 6 biologically independent replicates per group) (p). q and r, FACS analysis of IL-22 and IL-17A expression in MNK-3 cells transduced with the indicated retroviral constructs. Representative FACS plot (q) and percentages of IL-22⁺ cells in MNK-3 cells compiled from two independent experiments (n = 3 biologically independent replicates per group) (r).

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