Fig. 4: USP22 specifically interacts with lipid metabolism key transcription factor of PPARγ.

a Tandem affinity purification–mass spectrometry detection of USP22-interacting proteins (obtained from S-beads pulldown) after HEK293T cells were transfected with SFB-USP22 for 24 h. b Cell lysates of MHCC-97H cells were immunoprecipitated with IgG or USP22 antibodies, and immunoblot assays were performed using USP22, PPARγ, PPARα, PPARδ and SREBF1 antibodies. c Cell lysates of MHCC-97L, HUH7, HepG2, SNU-449, and Bel-7402 cells were immunoprecipitated with IgG or USP22 antibodies, and immunoblot assays were performed using USP22 or PPARγ antibodies. d GST pulldown assay with purified His-USP22 and GST-PPARγ. PD Pulldown. e HEK293T cells were transfected for 24 h with plasmids encoding either Flag-PPARγ or Myc-USP22 alone or in combination. Cell lysates were immunoprecipitated with Flag and Myc antibodies, and immunoblotting was performed using Myc or Flag antibodies. f Triple immunoflorescence (IF) staining for USP22 (red), PPARγ (green), and nuclei (DAPI, blue) was performed in MHCC-97L and MHCC-97H cells. Scale bars, 10 μm. g Plasmids containing FL (full length), AF-1, DBD, Hinge-LBD domain of PPARγ were co-expressed with SFB-USP22 in 293 T cells. Lysates were immunoprecipitated with S-beads. h GSEA of signaling pathway with USP22-correlated genes based on TCGA LIHC database. All experiments were performed independently at least three times.