U2OS and HCT116 cells (ATCC, Manassas, VA, USA) were grown in high glucose Dulbecco's modified Eagle's media (DMEM) with pyruvate (Thermo Fisher Scientific, Darmstadt, Germany). RPE-1 hTERT cells (ATCC) were cultured in DMEM:F12 media (Thermo Fisher Scientific). Culture media were supplemented with 10% fetal bovine serum (FBS; Thermo Fisher Scientific) and penicillin/streptomycin (Thermo Fisher Scientific). Cell lines were tested at least twice a year for Mycoplasma contamination using the LookOut Detection Kit (Sigma), and all tests were negative.
Cells were treated with DMSO (0.15%; Carl Roth, Karlsruhe, Germany), Nutlin-3a (10 μM; Sigma Aldrich, Darmstadt, Germany), Actinomycin D (5 nM; Cayman Chemicals, Ann Arbor, MI, USA), or Doxorubicin (1 μM or as indicated; Cayman Chemicals) for 24 h. For knockdown experiments, cells were seeded in six-well plates or 6 cm dishes and reverse transfected with 5 nM Silencer Select siRNAs (Thermo Fisher Scientific) using RNAiMAX (Thermo Fisher Scientific) and Opti-MEM (Thermo Fisher Scientific) following the manufacturer protocol.
Images of cells were taken using an Evos M5000 microscope (Thermo Fisher Scientific) or a ChemiDoc MP documentation system (Bio-Rad, Feldkirchen, Germany).
Cells were treated with DMSO (0.15%; Carl Roth, Karlsruhe, Germany), Nutlin-3a (10 μM; Sigma Aldrich, Darmstadt, Germany), Actinomycin D (5 nM; Cayman Chemicals, Ann Arbor, MI, USA), or Doxorubicin (1 μM or as indicated; Cayman Chemicals) for 24 h. For knockdown experiments, cells were seeded in six-well plates or 6 cm dishes and reverse transfected with 5 nM Silencer Select siRNAs (Thermo Fisher Scientific) using RNAiMAX (Thermo Fisher Scientific) and Opti-MEM (Thermo Fisher Scientific) following the manufacturer protocol.
Images of cells were taken using an Evos M5000 microscope (Thermo Fisher Scientific) or a ChemiDoc MP documentation system (Bio-Rad, Feldkirchen, Germany).