Hek293t

The GC cell lines (MGC803, MKN45, HGC27, SGC7901, and AGS), normal gastric cells GES1, and HEK293T were obtained from the China Infrastructure of Cell Line Resources (Beijing, China). The GC cells were cultured in RPMI-1640 medium while the GES1 and HEK293T were cultured in DMEM medium. All the media were complemented with 10% fetal bovine serum (FBS, Hyclone, Logan, UT, USA), 100 units/mL penicillin, and 100 μg/mL streptomycin (Living, Beijing, China). All the cells were maintained in a humidified incubator containing 5% CO₂ at 37 °C. The cobalt chloride (#232696) and glycine (#V900144) were purchased from Sigma-Aldrich (St. Louis, MO, USA). The formic acid was purchased from Thermo Fisher Scientific (#85178, Waltham, MA, USA). The doxycycline was purchased from Topscience (#T1687, Shanghai, China).

Human embryonic kidney HEK293T cells (HEK293T), Madin-Darby canine kidney cells (MDCK), and human lung adenocarcinoma epithelial cells (A549) were obtained from China Infrastructure of Cell Line Resources and maintained in Dulbecco’s modified Eagle’s medium (DMEM) (Invitrogen) supplemented with 10% heat-inactivated fetal bovine serum (FBS) and antibiotics (100 U/ml penicillin G and 100 g/ml streptomycin).
The A/WSN/33 (H1N1) (WSN) strain of IAV were grown in MDCK cells.
Anti-M1 monoclonal antibody and anti-NP polyclonal antibody were gifts from Prof. Wenjun Liu. Anti-Myc (9E10) and FLAG (M2) antibodies were purchased commercially (Sigma). Anti-His (H-3) and anti-β-actin (1–19) antibodies were also purchased commercially (Santa Cruz Biotechnology). Anti-NEDD8 (Y297) was purchased commercially (Abcam).

A549, HEK293T and MDCK cell lines were obtained from the China Infrastructure of Cell Line Resources and cultured in Dulbecco’s modified Eagle’s medium (DMEM, GIBCO, Waltham, MA, USA). All cell cultures were supplemented with 10% fetal bovine serum (FBS, GIBCO) and maintained at 37 °C in a humidified incubator with 5% CO₂. HEK293T-IAV-Luc cells were generated by transfection of plasmid pREP-FluA-Luc (kindly provided by Prof. Andrew Pekosz, Johns Hopkins University, Baltimore, MD, USA) where the firefly reporter gene is right behind the NP promoter of influenza virus and which expresses firefly luciferase upon influenza A virus infection. lncRPS6P3-konckdown A549 cells were generated using shRNA. The influenza A virus strains A/WSN/33 (A/WSN; H1N1), A/Puerto Rico/8/34 (A/PR8; H1N1) and A/California/2009/04 (A/CA04; H1N1) (kindly provided by Prof. Wenjun Liu, Institute of Microbiology, Chinese Academy of Sciences, Beijing, China) were propagated in MDCK cells.