Fig. 2: Identifying the primary transcriptional targets of NONO-TFE3 in tRCC. | Nature Communications

Fig. 2: Identifying the primary transcriptional targets of NONO-TFE3 in tRCC.

From: RNA-mediated condensation of TFE3 oncofusions facilitates transcriptional hub formation to promote translocation renal cell carcinoma

Fig. 2

a Scheme of the experimental design for SLAM-seq in UOK109 KI cells. See details in the “Methods” section. b Principal component analysis (PCA) of SLAM-seq results at the indicated timepoints. n = 2 independent biological replicates. c Heatmap of differentially expressed genes (DEGs) at the indicated timepoints after dTAG-13 (500 nM) treatment as measured by SLAM-seq in UOK109 KI cells. d Volcano plot showing the genes differentially expressed after 4-h dTAG13 treatment (two-sided, unpaired Student’s t-test). e The Gene Ontology (GO) analysis of DEGs identified from the 4-h dTAG13 treatment group (Fisher’s exact test). f Venn diagram showing the overlapping genes identified from SLAM-seq and CUT&Tag analysis (comparison between before and 4-h after dTAG13 treatment). g Average metagene plots (left) and quantification (right) of CUT&Tag or RIP-seq signal intensities of defined NONO-TFE3 targeted (n = 115) or non-targeted genes (n = 4896) from −5 kb TSS to +5 kb TES. The box plots indicated the median (center line), the third and first quartiles (box limits) and 1.5x interquartile range (IQR) above and below the box (whiskers). (n = 2 independent biological replicates; two-sided Wilcoxon test). h Venn diagram showing the overlapping genes identified from SLAM-seq (4-h dTAG 13 treatment) and RIP-seq assays. i Scheme depicting the reporter system (top) where local RNA expression near a luciferase reporter gene can be induced by doxycycline (Dox). Transcriptional activity of NONO-TFE3 was monitored by luciferase intensity (normalized to 0 ng/mL Dox, bottom). n = 6 independent biological replicates, one-way ANOVA with Tukey’s post-hoc test. Data are shown as mean ± SD. j A tentative model showing how NONO-TFE3 regulates target gene expression. High promoter binding and cis binding of newly synthesized mRNAs at the same loci are both required to sustain the expression of NONO-TFE3 target genes. Source data are provided as a Source Data file. Created in BioRender. Suris, A. (https://BioRender.com/j0aoaxy).

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