Fig. 3

SEC63 increases the stability of ACLY. A Whole cell lysates extracted from the Huh7 or PLC/PRF/5 cells were subjected to western blot using anti-SEC63 or anti-ACLY antibodies. B-C Huh7 cells were transfected as indicated. Then, western blot was performed. D Total RNA extracted from the SEC63-depleted cells or control cells were subjected to RT-qPCR using the ACLY primers. E The ctrl cells or SEC63-depleted cells were treated with CHX (50 μg/mL) as indicated time points. Western blot was performed for evaluating ACLY and SEC63 level. ACLY expression level was summarized from three independent experiments (lower panel). F Huh7 cells were transfected as indicated and treated with CHX (50 μg/mL) as indicated time points. Western blot was performed for evaluating ACLY and SEC63 level. ACLY expression level was summarized (right panel). G The ctrl cells or SEC63-depleted cells were treated with MG132 (10 μM) for 4 h. Western blot was performed as indicated. H The ctrl cells or SEC63-depleted cells were treated as indicated. The ubiquitination level of ACLY was analyzed by immunoprecipitation. I Huh7 cells were treated with MG132 (10 μM) for 4 h. The ubiquitination level of ACLY was analyzed by immunoprecipitation. J After MG132 treatment, the ubiquitination level of ACLY was analyzed by immunoprecipitation in Huh7 cells. K Huh7 cells were transfected with indicated plasmids and cell lysates were subjected to Co-IP using anti-ACLY antibody. L Ctrl cells or SEC63-depleted cells were treated with the indicated drugs. The cell lysates were subjected to Co-IP using anti-ACLY antibody