Dnmt1

Short interfering RNA (siRNA) pre-designed for HDAC1 (Qiagen), RGS10 and DNMT1 (Santa Cruz) were used to knock down expression of HDAC1, RGS10 or DNMT1. Scrambled All Star Control siRNA (Qiagen) was used as a control. A2780-AD cells were transfected with 10 nM of HDAC1 or DNMT1 specific siRNA or All Star scrambled control siRNA using HiPerfect transfection reagent (Qiagen) according to the manufacturer's instruction. Following indicated incubation time, cells were harvested and analyzed in western blot, RNA expression, or chromatin immunoprecipitation experiments.

siCtrl (#1): AllStars Neg. Control siRNA, Cat. No./ID: 1027281 (QIAGEN), siCtrl (#2): CGUACGCGGAAUACUUCGA (Eurofins), siPP2A-A: UUUUCCACUAGCUUCUUC A (Eurofins), siHRAS: GAACCCUCCUGAUGAGAGU (Eurofins), siKRAS: AGAGUGCCUUGACGAUACA (Eurofins), siNRAS: GAAAUACGCCAGUACCGAA, siPME (#1): GGAAGUGAGUCUAUAAGCA, siPME-1 (#2): UCAUAGAGGAAGAAG AAG A, siSET (#1): UGCAGACACUUGUGGAUGG (Eurofins), and siSET (#2): AAUGCA GUGCCUCUUCAUC (Eurofins). All siRNAs (CHD3, DNMT1, DOT1L, KDM1A, MLLT3, RNF168, and SMARCA4) for the cell viability assay were ordered from QIAGEN. AllStars Hs Cell Death siRNA, Cat. No./ID: 1027299 (QIAGEN), was used as a positive control (siCtrl +).
DNMT1 inhibitors (decitabine; AZA), BET inhibitors (iBET151, JQ1, mivebresib), HDAC inhibitors (panobinostat; TSA), KDM1A inhibitors (SP2509), and okadaic acid were used and purchased from SelleckChem. PP2A-reactivating compound DBK1154 was a kind gift of Dr. Michael Ohlmeyer (Atux Iskay; LCC).