The MSH2 and MSH6 luciferase reporter and NRIP1 expression vectors were previously described [26 (link),27 (link),28 (link),29 (link)]. HCT116 cells were transfected in 96-well plates (2.5 × 104 cells per well) 24 h prior to DNA transfection with Jet-PEI (275 ng of total DNA). Increasing doses of pEF-c-myc-RIP140 or pEF-c-myc-RIPMSI were cotransfected with the pGL3-MSH2-Luc or the Sp1 mutant pGL3-MSH2m1-Luc (kind gifts of E. Huang [28 (link)]). Similar experiments were performed with the pGL3-MSH6-Luc reporter vector and a Sp1 mutant pGL3-MSH6M1-2/7-Luc (kind gifts of R.D. Kolodner [29 (link)]). The pRL-CMV-renilla plasmid (Promega, Charbonnières-les-Bains, France) was used to normalize transfection efficiency. Firefly luciferase values were measured and normalized by the Renilla luciferase activity. Values were expressed as the mean ratio of luciferase activities.
ChIP assays for proteins at the MSH2 and MSH6 promoters were performed in HT29 cells using the CHIP-IT kit (Active Motif, Carlsbad, CA, USA). Sonicated chromatin was immunoprecipitated with antibodies against IgG (sc-3739, Santa Cruz Biotechnology, Inc, Heidelberg, Germany), H3pan (CC16310135, Diagenode, Liège, Belgium) and NRIP1 (ab42126, Abcam, Paris, France). Immunoprecipitated DNA was amplified by qPCR using the primers listed inSupplementary Table S1 .
ChIP assays for proteins at the MSH2 and MSH6 promoters were performed in HT29 cells using the CHIP-IT kit (Active Motif, Carlsbad, CA, USA). Sonicated chromatin was immunoprecipitated with antibodies against IgG (sc-3739, Santa Cruz Biotechnology, Inc, Heidelberg, Germany), H3pan (CC16310135, Diagenode, Liège, Belgium) and NRIP1 (ab42126, Abcam, Paris, France). Immunoprecipitated DNA was amplified by qPCR using the primers listed in
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