Fig. 3

CircNF1 manipulates the JAK–STAT3 pathway to regulate ESCC malignant progression and PD-L1 transcription. A Volcano plot showing differentially expressed genes in TE1 cells transfected with si-circNF1 and si-NC. B, C GO analysis B and KEGG pathway analysis (C) of circNF1-regulated genes in RNA-seq dataset. D qRT-PCR validating the expression of five candidate genes involved in the JAK–STAT3 pathway in circNF1 knockdown cells. E Western blotting assay detecting the expression of STAT3 and p-STAT3 in circNF1 knockdown or overexpressing cells, with GAPDH as normalized control. F Western blotting assay detecting PD-L1 expression in circNF1 knockdown or overexpressing cells, with GAPDH as normalized control. G ChIP assays analyzing whether p-STAT3 is bound to the PD-L1 promoter region in KYSE150 and TE1 cells. H, I EdU (H) and Transwell (I) assays evaluating the proliferation and migration ability of ESCC cells treated with DMSO or Stattic in control and circNF1-overexpressed ESCC cells. Scale bar: 100 μm. The data are presented as mean ± SD. *P < 0.05, **P < 0.01, ***P < 0.001