Figure 6: Ezh2 ablation and GSK503 treatment prevents murine melanoma growth.

(a) Mouse genotypes and strategy as in (Fig. 5b) used to analyze the effect of conditional Ezh2 ablation on melanoma proliferation in Tyr::N-Ras^Q61K Ink4a^−/− mice. (b) Immunofluorescent staining on skin melanoma sections for Sox10 (control) or β-Gal (cKO) and Ki67 4 weeks after conditional Ezh2 ablation to quantify a proliferation rate. White arrowheads, Sox10- or β-Gal-positive/Ki67-positive cells. (c) Mouse genotypes and strategy as in (Fig. 5f) used to analyze the effect of temporary GSK503 treatment on melanoma proliferation in Tyr::N-Ras^Q61K Ink4a^−/− mice. (d) Immunofluorescent staining on skin melanoma sections for Sox10 and Ki67 4 weeks after treatment start with vehicle or GSK503 to quantify a proliferation rate. White arrowheads, Sox10-positive/Ki67-positive cells. (e) Mouse genotypes and strategy used to s.c. engraft and expand Tyr::N-Ras^Q61K Ink4a^−/− mice-derived melanoma cells in Foxn1^nu/nu animals to analyze the effect of conditional Ezh2 ablation and GSK503 treatment on melanoma growth. (f) Representative macroscopic pictures of control, cKO and GSK503-treated RIM-1 allografts. (g–i) Western blot for Ezh2 protein and H3K27me3 on lysed tumours from (f) to quantify loss of Ezh2 (h) and H3K27me3 (i). (j,k) Growth of control, cKO and GSK503-treated RIM-1 allografts (j) and relative tumour volume of all RIM allografts (j; Supplementary Fig. 8k,n) at endpoints (k). Black arrow, time point of TM application/start of GSK503 treatment. E, epidermis; s.c., subcutaneous. Data are represented as mean±s.e.m. of n=5 (b), mean±s.e.m. of n=6 (vehicle), n=5 (GSK503) (d), mean±s.e.m. of n=3 (control), n=4 (cKO, GSK503) (g–j), mean±s.e.m. of n=9 (control, GSK503), n=6 (cKO) (k). P values calculated with unpaired Student’s t-test (b,d), analysis of variance and Fisher’s least significant difference test (h–k). Scale bars, 50 μm (b,d), 1mm (f).