Fig. 2: The O-galactosyltransferase C1GALT1 is highly expressed in ES cells, and its genetic or pharmacological inhibition reduces EWSR1::FLI1 levels and function.

a Representative immunoblot showing EWSR1::FLI1 levels (monitored using an anti-FLI1 antibody) in two independent C1GALT1 knockout (KO) A673 clones. b qRT-PCR analysis monitoring relative mRNA levels of EWSR1::FLI1 target genes in A673 cells stably expressing an NS, EWSR1, or C1GALT1 shRNA. Data are presented as mean ± SEM (n = 3 biologically independent experiments). P-values were calculated using one-way ANOVA with post-hoc Dunnett’s multiple comparisons test. c Representative IHC images showing H&E and C1GALT1 staining in n = 3 independent ES patient tumor samples, as well as normal bone and cartilage tissues. FLI1 staining is shown in the ES tumor samples. Scale bars, 50 µm. d (Left) Representative immunoblot showing levels of EWSR1::FLI1 (monitored using an anti-FLI1 antibody) and C1GALT1 in A673 cells treated with DMSO or itraconazole (ITZ) for 72 h. (Right) qRT-PCR analysis monitoring relative EWSR1::FLI1 mRNA levels in A673 cells treated with DMSO or ITZ for 72 h. The results were normalized to that obtained with DMSO, which was set to 1. Data are presented as mean ± SEM (n = 3 biologically independent experiments). P-values were calculated using one-way ANOVA with post-hoc Dunnett’s multiple comparisons test. e qRT-PCR analysis monitoring relative mRNA levels of EWSR1::FLI1 target genes in A673 cells following treatment with DMSO or ITZ (100 nM for 72 h). Data are presented as mean ± SEM (n = 4 biologically independent experiments). P-values were calculated using a two-tailed Mann–Whitney test. Source data are provided as a Source Data file.