Edem1

Total proteins were extracted from liver tissues and cells using radio-immunoprecipitation buffer (RIPA). Equal amounts of denatured proteins were separated on 10% SDS-PAGE and analyzed by immunoblotting with primary antibodies against GCLC (1:1000; Proteintech, 12601-1-AP), α-SMA (1:1000; Proteintech, 14395-1-AP), COL1 (1:1000; Abcam, ab6308), glucose-regulated protein 78 (GRP78; 1:1000; Proteintech, 11587-1-AP), CCAAT/enhancer-binding protein homologous protein (CHOP; 1:1000; Proteintech, 15204-1-AP), phosphorylated inositol-requiring enzyme 1 (p-IRE1; 1:1000; Abcam, ab48187), endoplasmic reticulum degradation enhancer, mannosidase alpha-like 1 (EDEM1; 1:500; Proteintech, 26226-1-AP); protein kinase inhibitor (p58^IPK; 1:1000; Abcam, ab70840); nuclear factor kappa B (NF-κB; 1:1000; Proteintech, 14220-1-AP), inhibitor of KB kinase b (IKKB; 1:1000; Proteintech, 15649-1-AP), tumor necrosis factor α (TNFα; 1:1000; Abcam, ab6671), transforming growth factor 1 (TGFβ1; 1:1000; Proteintech, 21898-1-AP), tissue inhibitor of the metalloproteinase (TIMP1; 1:1000; Proteintech, 16644-1-AP), Matrix metalloproteinase 2 (MMP2; 1:1000; Proteintech, 10373-2-AP), and β-actin (1:10000; Proteintech, 14395-1-AP). After the incubation with HRP-conjugated secondary antibodies, the signals were visualized by the ECL system (Thermo Fisher Scientific).

Rabbit polyclonal antibodies were used to detect ERdj3, EDEM1, and ERp57 (Proteintech, Chicago, IL); others were to detect AAT (DAKO, Carpentaria, CA), GAPDH (Santa Cruz, Dallas, TX), Lamp1, P62, LC3B (all from Cell Signaling, Danvers, MA), and calnexin and calreticulin (both from Stressgen biotechnologies, San Diego, CA). Bafilomycin A1, MG132, Brefeldin A, and mouse monoclonal anti‐β‐actin were purchased from Sigma (St. Louis, MO), and 2C1 against human AAT polymers was purchased from Hycult biotech (Netherlands). Mouse monoclonal antibodies against BiP and AAT were purchased from BD bioscience (San Jose, CA) and R&D systems (Minneapolis, MN), respectively. Alexa Fluor 488 goat anti‐mouse IgG and Alexa Fluor 594 goat anti‐rabbit IgG were purchased from Invitrogen. Primers, Dynabeads protein A and G, Superscript VILO cDNA synthesis kit, and ERdj3 siRNA were purchased from Life Technologies, and HP DNA transfection reagent and TaqMan Universal PCR Master Mix were purchased from Roche Applied Science. Disuccinimidyl suberate (DSS) cross‐linker was purchased from Thermo Fisher scientific (Waltham, MA).