Standard methods of Western blotting and immunoprecipitation were used. Briefly, cells were lysed using RIPA buffer (Beyotime, Shanghai, China) and the supernatant was resolved by SDS-PAGE after centrifugation and transferred to PVDF membranes for Western blotting. Then, the membranes incubated with ECL luminescence reagent (Meilun Biotechnology Co., Ltd, Dalian, China) and exposed using Tanon 5200 Chemiluminescence Imaging System (Tanon Technology Co., Ltd, Shanghai, China).
The glutathione S-transferase (GST) pull-down assays and immunoprecipitation experiments were performed as described before [21 (link)]. The supernatant was incubated FLAG-tagged beads (A2220; Sigma-Aldrich, St. Louis, MO, USA) or HA-tagged beads (11815016001; Roche, Mannheim, Germany) or Glutathione Beads (SA008005; Smart-Lifesciences Biotechnology Co., Ltd, Changzhou, China) for 4 h. The precipitates were then washed five times and analyzed by Western blotting.
The glutathione S-transferase (GST) pull-down assays and immunoprecipitation experiments were performed as described before [21 (link)]. The supernatant was incubated FLAG-tagged beads (A2220; Sigma-Aldrich, St. Louis, MO, USA) or HA-tagged beads (11815016001; Roche, Mannheim, Germany) or Glutathione Beads (SA008005; Smart-Lifesciences Biotechnology Co., Ltd, Changzhou, China) for 4 h. The precipitates were then washed five times and analyzed by Western blotting.
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