Fig. 7.

β-Catenin is a transcription factor targeting the PD-L1 gene. a, b qRT-PCR and agarose gel electrophoresis showing interaction of the PD-L1 promoter fragment with β-catenin in ISG12a-silenced Huh7 and HGC-27 cells, as determined using the ChIP assay. The immunoprecipitated DNA is presented as the signal relative to the total amount of input chromatin (a). Histone H3 ChIP for RPL30 served as the positive control, and IgG ChIP served as the negative control. The data are presented on the log₂ scale. c Fold enrichment of the interaction fragment of the PD-L1 promoter with β-catenin in Huh7 and HGC-27 cells with ISG12a knockdown, as determined by qRT-PCR using the ChIP assay. d, e Transcriptional activity of the PD-L1 promoter (pGL3-CD274) was modulated by CTNNB1 in HEK293T cells (d) or ISG12a and CTNNB1 in Huh7 (e) cells, as examined using dual-luciferase reporter assays. f Immunoblots for PD-L1 and Flag in Huh7 cells, and the expression level of PD-L1 relative to GAPDH between the two groups was compared by grayscale analysis. Cells were transfected with p3×Flag-CTNNB1 or vector for 48 h. g Immunoblots showing levels of PD-L1 and β-catenin in HGC-27 cells. Cells stably transfected with plasmids for silencing ISG12a were further transfected with plasmids for 48 h to silence CTNNB1. Experiments were independently repeated two (a–e, g) or three (f) times, with similar results. A two-sided Student’s t test (f) or one-way ANOVA with the Tukey–Kramer or Games-Howell post hoc test (a, c–e) was performed to analyze the results, and data are presented as means ± SD of three biological replicates. *p < 0.05, **p < 0.01, and ***p < 0.001