TLR3 served as a downstream gene of BHLHE40 and promoted migration and invasion of PCA cells. (A) The JASPAR database predicts binding sites between BHLHE40 and the promoters of GCNT4 and TLR3; (B) knockdown of BHLHE40 repressed the expression of TLR3 in PANC-1 cells; (C) by using ChIP assay, BHLHE40 was immunoprecipitated from PANC-1 cells, and the binding of BHLHE40 to the promoter of TLR3 was detected using qRT-PCR with the indicated primers; (D) knockdown of TLR3 by using siTLR3 repressed the expression of TLR3 in PANC-1 cells; (E) cell migration was assessed using a wound healing assay upon treatment with siTLR3 (×40); (F) the invasion ability of PANC-1 cells transfected with siTLR3 was evaluated by using transwell assay (×200), in which cells were stained with 0.1% crystal violet; (G) the invasion ability of PANC-1 cells co-transfected with siBHLHE40 and siTLR3 was determined by transwell assay (×200), in which cells were stained with 0.1% crystal violet. All data are presented as the means ± SD of 3 independent experiments. *, P<0.05; **, P<0.01; ***, P<0.001. PCA, pancreatic cancer; TSS, transcription start site; ChIP, chromatin immunoprecipitation; qRT-PCR, quantitative real-time polymerase chain reaction; siRNAs, small interfering RNAs; SD, standard deviation; siTLR3, siRNAs targeting TLR3.