Supplementary Figure 9: EXOSC10 mediates interaction between C1D and EZH2

a, Nuclear protein extracts of MLE-12 cells were analyzed by immunoprecipitation (IP) using either immunoglobulin G (IgG, as control) or EXOSC10-specific antibodies to precipitate endogenous EXOSC10. Co-purifying proteins were analyzed by WB using the indicated antibodies. Input, 5% of material used for the IP prior to LC-MS/MS analysis. See Fig. 5a-b. b, Sequential ChIP (ChIP-reChIP) analysis of the Mafg- and Chmp2b-promoters using chromatin isolated from MLE-12 cells after transfection of control (Ctrl) plasmid or expression constructs for EXOSC10 fl, del1 or del4. See Fig. 5g. c, qRT-PCR-based analysis of ncRNA and cRNA of the indicated genes in cells after transfection of control (Ctrl) plasmid or expression constructs for EXOSC10 fl, del1 or del4. See Fig. 5g. d,e, Confocal microscopy after proximity ligation assay using EXOSC10- and EZH2-specific antibodies in MLg cells that were Ctrl or Mirlet7-sponge transfected. Squares show details at higher magnification. Scale bars, 20 µm. See Figure S6e. f, Nuclear protein extracts of MLE-12 cells that were transfected with control- (Ctrl), Mirlet7d-antagomir (anti), Mirlet7-sponge (spng) or C1d-specific shRNA construct were analyzed by immunoprecipitation (IP) using either immunoglobulin G (IgG, as control) or EXOSC10-specific antibodies to precipitate endogenous EXOSC10. Co-IP proteins were analyzed by WB using the indicated antibodies. Input, 5% of material used for the IP. See also Figures S6e,g. g, Nuclear protein extracts of MLE-12 cells, that were treated or untreated with RNaseA or DNase1, were analyzed by immunoprecipitation (IP) using either immunoglobulin G (IgG, as control) or EXOSC10-specific antibodies to precipitate endogenous EXOSC10. Co-IP proteins were analyzed by WB using the indicated antibodies. Input, 5% of material used for the IP. See Fig. 6a-b. f, qRT-PCR-based analysis of the indicated ncRNAs after miR-Pd from cells transfected with biotinylated Mirctrl or biotinylated Mirlet7d. Nuclear protein lysate of biotinylated Mirlet7d transfected cells were treated with Proteinase K before miR-Pd as indicated. See Fig. 6a-b. h, ChIP analysis of Mafg- and Chmp2b-promoters using the indicated antibodies in MLE-12 cells transfected with Mirctrl, Mirlet7d or/and C1d-specific shRNA constructs. See Fig. 6c. In a, d-g, representative results from 2 independent experiments (see Supplementary Data Set 3). In all plots, data are shown as means ± s.e.m (n=3 independent experiments); asterisks, P values after one–way ANOVA, ***P˂0.001; **P˂0.01; *P˂0.05; ns, non-significant. The statistical test values of each plot are shown in the Supplementary Data Set 4