Extended Data Fig. 3: IL-6-Myc signalling is crucial for the self-renewal of THY1+ CSCs.
a, FACS-sorted THY1− human and murine hepatoma cell lines were cultured for different durations in vitro. Percentage of THY1+ cells regenerated from THY1− cells over time (n = 5). b,c, Efficiency of shNANOG, shMYC (b) or MYC-OE (c) in Huh-7 cells (each n = 5). d, Percentage of THY1+ cells generated from unsorted, wild-type THY1− (cultured with control medium), or MYC-OE THY1− (cultured with doxycycline) Huh-7 cells over time (n = 5). The induced group was compared with their corresponding uninduced group. e, GSEA of NANOG or c-Myc signatures (right: M5926) on pseudotime-ordered developmental trajectory from CSC.c4 to CSC.c3 populations. f, Schematic illustration of c-Myc binding sites in the THY1 promoter region. TSS, transcription start site. g, Sequence analysis identifying c-Myc binding sites within the promoter regions of the THY1 gene. h, Schematic diagram illustrating the truncated 5′-flanking regions of the THY1 gene. i,j, Effect of signalling inhibitors treatment (i) or shIL6R (j) on c-Myc expression in Huh-7 cells (each n = 5). k, Effects of IL-6 family cytokines on c-Myc expression in Huh-7 cells. (n = 5). l–n, Wild-type or Il6ra-knockdown EpCAM+ Hepa1-6 cells were inoculated into mouse livers. The efficiency of Il6ra-knockdown was examined (l; n = 5). Gating strategy for FACS analysis of GFP+ tumour cells was shown (m). Tumour volume was measured (n; n = 5; scale bar, 1cm). o, GSEA of c-Myc signatures (left: M5926) or IL-6–STAT3 pathway (right: M5897) in THY1+ versus THY1− CSCs from pan-cancer scRNA datasets. Data are from five (b,c,i,j,k) independent experiments. Data are presented as mean ± s.e.m. of three (l,n) or five (a,d) independent experiments. Statistical significance was determined using two-way ANOVA with Tukey’s post hoc test (d), a one-sided, permutation-based test (e,o), or unpaired two-tailed Student’s t-test (l,n).
