293t cell line

Fourth-generation lentivirus packing systems (Lenti-X, Takara-Clontech) were used to generate stable copGFP-expressing and mCherry-expressing cell lines. Briefly, puromycin-resistant – copGFP (pLenti-CMV-GFP-Puro) and blasticidin-resistant mCherry (pLV-Bsd-CMV-mCherry) vectors were obtained and amplified in stable competent EColi (New England Biolabs) by employing standard molecular biology protocol with 100 µg/mL Ampicillin selection, purified for lentivirus packing using commercial plasmid prep kit (Takara-Clontech). For generating lentivirus, Lenti-X reagent was mixed with 6 µg of lentivirus plasmid and transfected in 293T cell line (Takara-Clontech) for packing. Harvested lentivirus was used to transduce target cell lines using polybrene by following established protocol. Hepa1-6, HepG2, and Hep3B cell lines were transduced with copGFP lentivirus, and cell lines were selected with 3 µg/mL puromycin (Sigma-Aldrich) and expanded after confirmation by fluorescence microscope for copGFP expression (Supplementary Figure 4). Hepa1-6 cell line was transduced with mCherry lentivirus, and selected with 10 µg/mL blasticidin (Sigma-Aldrich).

Human prostatic carcinoma cell line PC3 was obtained from the American Type Culture Collection (Manassas, VA, USA). Human hepatocellular carcinoma cell line HuH7 and human uterine cervical carcinoma cell line HeLa were supplied by Cell Resource Center for Biomedical Research, Institute of Development, Aging and Cancer, Tohoku University (Sendai, Japan). The 293T cell line, which is a highly transfectable derivative of human embryonic kidney cell line 293, was obtained from Clontech Laboratories, Inc. (Takara Bio USA, Inc., Mountain View, CA, USA). PC3, HuH7 and HeLa cells were maintained in RPMI‐1640 medium (Nissui Pharmaceutical, Tokyo, Japan) supplemented with 10% fetal bovine serum (Gibco, Thermo Fisher Scientific, Waltham, MA, USA), 2 mm l‐glutamine, 2.4 g·L⁻¹ NaHCO₃, 100 units·mL⁻¹ penicillin, and 100 μg·mL⁻¹ streptomycin. The 293T cells were maintained in Dulbecco's modified Eagle's medium (Gibco), 10% fetal bovine serum, 100 units·mL⁻¹ penicillin, and 100 μg·mL⁻¹ streptomycin. Cells were incubated at 37 °C under a humidified atmosphere of 5% CO₂. Pifithrin‐μ (PFT‐μ), VER‐155008, and JG98 were purchased from Tocris Bioscience (Bio‐Techne Corp., Minneapolis, MN, USA), Cayman Chemical Company (Ann Arbor, MI, USA), and Selleck Chemicals (Houston, TX, USA), respectively. These compounds were dissolved in dimethylsulfoxide at 10–100 mm as a stock solution.