Fig. 5: RCOR1 establishes an interaction with RNA POL-II after initiation and before elongation.

a Western blot analyses of immunoprecipitation of RCOR1 and co-precipitation of RPB1 and its phosphorylated isoforms from whole cell extracts. II-O: Hyperphosphorylated isoforms. II-A: Hypophosphorylated isoforms. RPB1-NTD CoIP panel is representative of four independent experiments while others are representative of two independent experiments. b Western blot analyses of immunoprecipitation of LSD1 (top) or HDAC1 (bottom) and co-precipitation of RPB1. Experiment performed once. c Western blot analysis of immunoprecipitation of RCOR1 and RNA POL-II from MNased chromatin using 2 different RCOR1 antibody concentrations. Panels are representative of two independent experiments. d HA-Pulldown on extracts derived from empty-vector and HA-RCOR1 overexpressing HEK293 cells. Panels are representative of two independent experiments. e Scheme depicting different stages of eukaryotic RNA POL-II transcription. It highlights the steps that are inhibited by THZ1, Flavopiridol, Cordycepin, and Actinomycin D. CTD: Carboxy-terminal domain. S5Ph: Phosphorylated serine 5. S2Ph: Phosphorylated serine 2. f Western blot analyses of RCOR1 immunoprecipitation and RPB1 co-precipitation under inhibition of transcription at different steps. Panels are representative of three independent experiments. g Proximity ligation assay between RCOR1 and RPB1 under DMSO, THZ1 and Actinomycin D treatments in HT22 cells. Right graph shows the distribution of proximal-ligated spots per cell under each treatment. More than 128 cells from three independent experiments were analyzed per treatment. Statistical significance was checked on Graphpad Prism by confirming normal distribution of data by D’Agostino & Pearson, Shapiro-Wilk and KS normality tests. Data was subjected to an ordinary, unpaired, one-way ANOVA test with 99% confidence intervals, **** means p < 0.01. Source data are provided as a Source Data file.