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Fig. 3 | Genome Biology

Fig. 3

From: HDAC4 degradation during senescence unleashes an epigenetic program driven by AP-1/p300 at selected enhancers and super-enhancers

Fig. 3

Depletion of HDAC4 triggers senescence in cancer cells. a Immunoblot analysis in SK-LMS-1/HDAC4+/+ or HDAC4−/− cells as indicated. Actin was used as loading control. b Cell-proliferation curve of the indicated HDAC4+/+ and HDAC4 −/− SK-LMS-1 cells. Mean ± SD; n = 4. c Analysis of the senescent cells as scored after SA-β-gal staining. Mean ± SD; n = 4. d Venn diagrams showing the number of transcripts differently regulated in SK-LMS-1/HDAC4−/− cells generated by using two different sgRNAs (green and red). Differentially expressed genes (DEGs) were selected based on |fold change| > 2 and p < 0.05. e Heat-map of the absolute expression levels of the DEGs in the indicated clones and biological replicates hierarchically clustered accordingly to average linkage. Blue shades intensity is proportional to transcripts abundance. f GSEA plots displaying the NES obtained by interrogating the transcriptome of HDAC4 WT and KO cells with the indicated gene sets. g Immunoblot analysis in SK-LMS-1 HDAC4+/+ or HDAC4−/−, stably re-expressing HygroR (clone 635) or HDAC4 (clones 275 and 279), as indicated. h Analysis of SA-β-gal positivity in WT or HDAC4−/− SK-LMS-1 cells expressing HYGROR (clone 635) or HDAC4 (clones 273–279). Mean ± SD; n = 4. The significance is relative to the KO clone 635. i. Analysis of BrdU incorporation in WT or HDAC4−/− SK-LMS-1 cells expressing HYGROR (clone 635) or HDAC4 (clones 273–279). Mean ± SD; n = 4. The significance is relative to the KO clone 635. j Analysis of SA-β-gal positivity in SK-LMS-1 WT or HDAC4 KO cells expressing HYGROR (clone 635) or GFP-NFKBIA (clones 55 and 70). Mean ± SD; n = 4. k Analysis of BrdU incorporation in SK-LMS-1 WT or HDAC4 KO cells expressing HYGROR (clone 635) or GFP-NFKBIA (clones 55 and 70). Mean ± SD; n = 4. l mRNA expression levels of the indicated genes in the indicated SK-LMS-1 WT or HDAC4−/− clones re-expressing HYGROR (clone 635), HDAC4 (clones 275–279) or GFP-NFKBIA (clones 55–70). Mean ± SD; n = 3. The significance is relative to the KO clone 635

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