Fig. 6

The risk allele of rs560789 reduced activity of epithelial-specific enhancer and prevented the binding of transcription factors related to epithelial development. a UCSC genome browser tracks of genomic fragments containing rs560789 that inserted into GFP reporter constructs and luciferase-based reporter vector. b Sanger-sequencing results of the products cloned into the GFP reporter plasmid showing the mutation of non-risk allele (A) to risk allele (G) in rs560789 (located at chr11:117228892). c, d Lateral views of wild-type zebrafish embryos at 11 hpf injected at the 1-cell stage with GFP reporter constructs inserted with enhancer containing non-risk allele (c) or risk allele (d). e 100% stacked column showing the relative percentage of the number of zebrafish embryos that expressed strong, weak, or non-fluorescent after GFP reporter injection. 156 and 120 zebrafish embryos were injected with plasmids containing fragments with non-risk allele and risk allele of rs560789, respectively. f Dual-luciferase assay for non-risk and risk allele of rs560789 in HIOEC and HEPM cells. ctrl, group without transfection; pGL3, group transfected with pGL3-promoter vector; ns, non-significance; ****P < 0.000 1. g Venn diagram showing 50 transcription factor motifs lost and six transcription factor motifs gained identified by Jaspar2022 after rs560789 mutated to risk allele. h qPCR of SOX2 ChIP at rs560789 (SOX2-target) or in VIM locus (SOX2-Offtarget, a region without SOX2 motif) using HIOEC cells. i Dual-luciferase assay for co-transfection of non-risk and risk alleles of rs560789 with pcDNA3.1(+) vector or SOX2 overexpression plasmid (pcDNA3.1(+)-Sox2) in HIOEC. j Examples of transcription factor that were lost or gained with rs560789 mutation of risk allele. k Dot plot showed the Gene Ontology (GO) enrichment assay for the 50 lost transcription factors. –log(P value) and false discovery rate (FDR) (enrichment value) for each term were generated by Metascape. l The new WashU epigenome browser tracks showing assay for ATAC-seq, H3K27ac ChIP-seq, and DLO Hi-C results of HIOEC at chr11:108 Mb–122 Mb. ATAC-seq and H3K27ac ChIP-seq results showed two replicates. Regions marked with light purple show examples of the promoters interacting with rs560789. Red arcs showed the interactions