We used human hepatoma cell lines established from hepatocellular carcinoma (Huh7) and a highly differentiated immortalized human hepatocyte (OUMS-29) [44 (link)]. Huh7 cells were obtained from RIKEN Bioresource center (Tsukuba, Japan). Cells were cultured in Dulbecco’s modified Eagle’s medium (DMEM) supplemented with 10% fetal bovine serum and antibiotics. Cells were maintained in a 37 °C incubator with 5% CO2.
The following materials were used: ALLN and epoxomicin as PIs (Calbiochem, La Jolla, CA, USA); copper sulfate (NACALAI TESQUE, Inc., Kyoto, Japan); PBA (Sigma-Aldrich, St. Louis, MO, USA); tunicamycin (Sigma-Aldrich); and zinc acetate (Wako Pure Chemical Industries, Ltd., Osaka, Japan).
Antibodies to the following antigens were used: LC3 (Medical and Biological Laboratories); Beclin 1 (Novus Biologicals, Littleton, CO, USA); Atg7, p-eIF2α (Cell Signaling Technology, Danvers, MA, USA); ubiquitin, XBP1, p62 (Santa Cruz Biotechnology, CA, USA); K8, actin (Sigma-Aldrich); Rubicon (Abcam, Cambridge, MA, USA); and ubiquitin (Dako, Glostrup, Denmark). Lamp1 antibody was a kind gift from J.T. August (Johns Hopkin University, Baltimore, MD, USA).
The following materials were used: ALLN and epoxomicin as PIs (Calbiochem, La Jolla, CA, USA); copper sulfate (NACALAI TESQUE, Inc., Kyoto, Japan); PBA (Sigma-Aldrich, St. Louis, MO, USA); tunicamycin (Sigma-Aldrich); and zinc acetate (Wako Pure Chemical Industries, Ltd., Osaka, Japan).
Antibodies to the following antigens were used: LC3 (Medical and Biological Laboratories); Beclin 1 (Novus Biologicals, Littleton, CO, USA); Atg7, p-eIF2α (Cell Signaling Technology, Danvers, MA, USA); ubiquitin, XBP1, p62 (Santa Cruz Biotechnology, CA, USA); K8, actin (Sigma-Aldrich); Rubicon (Abcam, Cambridge, MA, USA); and ubiquitin (Dako, Glostrup, Denmark). Lamp1 antibody was a kind gift from J.T. August (Johns Hopkin University, Baltimore, MD, USA).