Figure 5. R-spondin3 promotes MYC expression in NK cells in the TME.

(A) Quantitative RT-PCR results of Wnt target genes of flow-sorted CD11b⁺CD27⁻ NK cells from tumor tissues (n = 4 for each group). (B) Flow analysis of tumor-infiltrating NK cells from B16F10-EV or B16F10-Rspo3 tumors inoculated subcutaneously to Myc^G/G mice (n = 5 for each group). Median fluorescence intensities of GFP for the indicated NK cell subpopulations are shown. (C-D) Quantitative RT-PCR results of rRNA (C) or ribosomal protein mRNA (D) of flow-sorted tumor-infiltrating NK cells from B16F10-EV or B16F10-Rspo3 tumor tissues. Data are shown as mean ± s.d. Expression levels were normalized by cell numbers sorted. (E) Median FSC intensities of flow analyzed tumor-infiltrating NK cells (CD45⁺CD3⁻NK1.1⁺DX5⁺) from B16F10-EV and B16F10-Rspo3 tumors (n = 4-5 per group). (F) Tumor growth curves of B16F10-EV and B16F10-Rspo3 in Ncr1^Cre and Myc^Δ/Δ/Ncr1^Cre mice (n = 4-5 per group). (G) Tumor growth curves of B16F10-EV and B16F10-Rspo3 cells in Ncr1^Cre and Myc^Δ/Δ/Ncr1^Cre mice depleted with NK cells (left) or CD8⁺ T cells (right) (n = 4-5 per group). For panels A-D, and F-G, two-way ANOVA with or without Sidak’s multiple comparisons tests were performed. For panel E, student’s t-test was performed. Data represent at least two independent experiments. Data are shown as mean ± s.e.m. unless otherwise noted. P < 0.05 is considered as statistically significant. ns, not significant, *P < 0.05, **P < 0.01, ***P < 0.001 and ****P < 0.0001.