Fig. 3. MYC-driven HCC cells depend on XPO1 activity.

a Correlation analysis of MYC-regulated gene expression and gene essentiality. Left: Differential gene expression (MYC^high/MYC^low, log2-fold change) is plotted against gene essentiality. Gene expression was determined in the context of MYC-induced HCC development (MYC^high HCC (n = 6 mice)) and MYC inactivation-induced tumor regression (MYC^low HCC (n = 3 mice)) by RNA sequencing. Right: Gene essentiality (Beta score) is plotted for genes that are either significantly up or down regulated in MYC^high compared to MYC^low HCC (adjusted p-value < 0.05). Box: 25–75 percentile, whiskers: 5–95 percentile, line: median, data points outside of 5–95 percentile are shown. The p-value was determined by two-tailed Mann-Whitney U test. b Results of KEGG pathway analysis of MYC-upregulated MYC-SL genes. Significantly enriched pathways (FDR < 0.05) are shown. RNA transport pathway genes identified as MYC-SL are highlighted in red box. c Gene expression changes of MYC-SL RNA transport genes in the context of MYC-induced HCC development (red: MYC^high HCC (n = 6 mice) vs. normal liver tissue (n = 2 mice)) and MYC inactivation-induced tumor regression (grey: MYC^low HCC (n = 3 mice) vs. MYC^high HCC). Gene expression was determined by RNA sequencing (bars: mean, error bars: SE). d Epifluorescence images of total RNA (heat map, and green) by EU labeling in MYC-driven HCC cells upon XPO1 inhibition by Selinexor (scale bar = 5 μm). Staining of alpha-tubulin (red) and DNA (blue) were used for segmentation of nucleus and cytoplasm. Quantification of RNA abundance in the nucleus versus cytoplasm per cell (n = 28 cells examined in one experiment). Lines are means +/− SEM. The p-value for the two-tailed Student’s t-test is shown. e Flow cytometric analysis of EC4 cells with high MYC expression (MYC^high) or MYC transgene expression shut off (MYC^low) by doxycycline treated with XPO1 inhibitor Selinexor. Survival was assessed by propidium iodide/AnnexinV staining. Graph shows quantification of three independent experiments (bars: mean, error bars: SEM). The p-value for the two-tailed Student’s t-test is shown. f Flow cytometric analysis of MYC-driven HCC cells expressing shRNAs targeting Xpo1 (two different Xpo1-targeting shRNAs) or GAPDH as a control in combination with GFP as a reporter. Live cells were detected by propidium iodide exclusion. Shown is the percentage of shRNA expressing (GFP+) live cells over time. Lines connect means of technical triplicates. For this figure, source data are provided as a Source Data file.