Fig. 6: RCOR1 globally represses transcription of newly synthesized transcripts.

a Workflow depicting the followed steps for imaging of newly synthesized RNAs under RCOR1 overexpression or knock down in HeLa cells. b Pseudocolored images of labeled transcripts under HA-RCOR1 overexpression. Scale bar represents 40 µm. c Box plots showing the quantitation of the relative fluorescent intensities per cell on each condition. Red asterisks indicate p < 0.05 respect to Mock-DMSO control. Green asterisks indicate p < 0.05 as significant rescue of the decreased transcription produced by RCOR1 overexpression. Non parametric, unpaired Kruskal Wallis tests were performed to evaluate statistical significance. d Pseudocolored images of labeled transcripts under HA-RCOR1 overexpression and DMSO or GSK-LSD1 treatments. Scale bar represents 40 µm. Right box plot shows the quantitation of the relative fluorescent intensities per cell on each condition. Asterisks indicate statistical significance as in (c). e Pseudocolored images of labeled transcripts under RCOR1 KD. Scale bar represents 40 µm. Right box plot shows quantitation of the relative fluorescent intensities per cell under RCOR1 KD conditions. Red asterisk indicates p < 0.05 respect to siControl according to non parametric, two tailed Mann Whitney test. f Pseudocolored images of labeled nascent transcripts when RCOR1 KD cells were subjected to recovery after washing out flavopiridol at 0, 20 and 40 min. Scale bar represents 40 µm. Box plots to the right show the quantitation of the relative fluorescent intensities per cell after each time point. Red asterisks represent p < 0.05 with respect to siControl cells at time 0 min according to non parametric, unpaired Kruskal Wallis tests were performed to evaluate statistical significance. Green asterisks represent p < 0.05 significantly different nascent transcription between the two groups at 20 min according to non parametric, two tailed Mann–Whitney test. g In vitro transcription using chromatin-free nuclear extracts from HeLa cells treated with Corin. Transcripts were labeled with 32P-α-ATP and the DNA template used was stained with EtBr as a parallel experiment. Right plots show the quantitation of the main or both transcript(s) obtained in this assay at each time point. *** indicates p < 0.001 whereas ** indicates p < 0.05 significantly different activity of Corin versus DMSO treated extracts (n = 3). *Additional representative images of EU-incorporation experiments are available in Supplementary Fig. 7. Statistical tests used for Fig. 6c–f were two-sided, unpaired T-tests.