Fig. 4

STAT3 increases PD-L1 expression through direct binding to the PD-L1 promoter. a The cytoplasm and nuclear translocation of STAT3 analyzed using cell fractionation in H460 cells after niclosamide treatment. b The putative binding mode of niclosamide and STAT3. STAT3 was shown as marine Cartoon and key residues was shown as marine sticks. Niclosamide was shown as light orange sticks. Hydrogen bonds were depicted as yellow dashed lines. c Relative mRNA expression levels of PD-L1 were decreased by niclosamide treatment in tumor cells. d-e Tumor cells expressing shSTAT3 or control were evaluated for STAT3 and PD-L1 expression by qRT-PCR and Western blot. f The − 765 to − 587 nucleotide sequence of the 5′-flanking region of PDL1 is shown. Underlined sequences are putative STAT3 transcription factor binding sites, as predicted by PROMO. g Overview of the four PD-L1 promoter fragments cloned into pGL3-Basic vector. h Luciferase activity measured and normalized according to Renilla luciferase activity in 293 T cells transiently transfected individually with the four promoter fragment constructs and empty luciferase vector pGL3-Basic for 48 h. Results are displayed as means ± S.D. of a representative experiment performed in triplicate. i Analysis of PD-L1 promoter fragment A construct in 293 T cells transiently transfected with STAT3 for 48 h. Relative luciferase activity was determined as described. Results are represented as means ± S.D. of a representative experiment performed in triplicate. * P < 0.05, ** P < 0.01, *** P < 0.001. Error bars represent SD of three independent experiments