Fig 1: The USP22–PPARγ/ACC/ACLY axis contributed to HCC prognosis.a Representative IHC staining of USP22, PPARγ, ACC, and ACLY in HCC TMAs (HLivH180Su11, contains prognosis information, Shanghai Outdo Biotech Company). Scale bars, 50 μm. b Correlation analysis between USP22 and PPARγ, ACC, ACLY protein expression based on H-Score in HCC TMAs (HLivH180Su11). R represents the Pearson correlation coefficient. c Kaplan–Meier curves of the survival analysis of USP22-positive, PPARγ-positive, USP22& PPARγ copositive, and USP22&ACC&ACLY copositive patients based on HCC TMAs prognosis data (HLivH180Su11). d Kaplan–Meier curves of the survival analysis of USP22-positive, PPARG-positive, USP22&PPARG copositive, and USP22&ACACA&ACLY copositive patients based on the prognosis data of TCGA LIHC database. e Diagram of the proposed mechanism. Source data are provided in the Source Data file.
Fig 2: USP22 increases ACC and ACLY expression by stabilizing PPARγ.a Western blot analysis of USP22 and PPARγ in cytoplasmic and nucleus fractions of MHCC-97H-shUSP22-1/2 cells and MHCC-97L-USP22 cells. LaminB1 and GAPDH were used as nucleus and cytoplasmic markers, respectively. b DNA binding activity of PPARγ in MHCC-97H-shUSP22-1/2 cells and MHCC-97L-USP22 cells. The analysis was performed by PPAR gamma Transcription Factor Assay Kit (ab133101, Abcam, USA). The data shown represent the means (±SD) of biological replicates. One-way ANOVA test. The experiments were repeated five time (n = 5). c Illustration of PPRE site in ACLY promoter and the predicted PPRE site in ACACA promoter. The PPRE motif from ACACA promoter was predicted by web site: https://epd.epfl.ch/index.php. d Chromatin immunoprecipitation (ChIP) analysis of PPARγ binding to the ACLY and ACACA promoters in MHCC-97H-shUSP22-1/2 cells. qPCR was performed with primers specific to the PPARγ-binding motifs. Data were normalized to the input. The data shown represent the means (±SD) of biological triplicates (n = 3). One-way ANOVA test. e, f qRT-PCR (e) and western blot analysis (f) of ACC and ACLY expression in MHCC-97H cells transduced with USP22 shRNA or in combination with PPARγ, and in MHCC-97L cells transduced with USP22 or in combination with PPARγ shRNA. The data shown represent the means (±SD) of biological triplicates (n = 3). One-way ANOVA test. g Representative IHC staining of USP22, PPARγ, ACC, and ACLY in HCC tissue microarrays (LV1021, no prognosis information, Shanxi ChaoYing Biotechnology Company). Scale bars, 50 μm. h Correlation analysis between USP22 and PPARγ, ACC, ACLY protein expression based on H-Score in HCC tissue microarrays (LV1021). R represents Pearson correlation coefficient. i The tissue microarray (LV1021) was stained with HE stain, and the steatosis was interpreted by the pathologist. Two-sided Chi-square test was performed to analyze the correlation between USP22, PPARγ, ACC, ACLY protein expression and steatosis. Source data are provided in the Source Data file.
Supplier Page from Abcam for PPAR gamma Transcription Factor Assay Kit