Systematic Functional Interrogation of Genes in GWAS Loci Identified ATF1 as a Key Driver in Colorectal Cancer Modulated by a Promoter-Enhancer Interaction

Genome-wide association studies (GWASs) have identified approximately 100 colorectal cancer (CRC) risk loci. However, the causal genes in these loci have not been systematically interrogated. We conducted a high-throughput RNA-interference functional screen to identify the genes essential for proliferation in the CRC risk loci of Asian populations. We found that ATF1 , located in the 12q13.12 region, functions as an oncogene that facilitates cell proliferation; ATF1 has the most significant effect of the identified genes and promotes CRC xenograft growth by affecting cell apoptosis. Next, by integrating a fine-mapping analysis, a two-stage affected-control study consisting of 6,213 affected individuals and 10,388 controls, and multipronged experiments, we elucidated that two risk variants, dbSNP: rs61926301 and dbSNP: rs7959129, that located in the ATF1 promoter and first intron, respectively, facilitate a promoter-enhancer interaction, mediated by the synergy of SP1 and GATA3, to upregulate ATF1 expression, thus synergistically predisposing to CRC risk (OR = 1.77, 95% CI = 1.42–2.21, p = 3.16 × 10 −7 ; P multiplicative-interaction = 1.20 × 10 −22 ; P additive-interaction = 6.50 × 10 −3 ). Finally, we performed RNA-seq and ChIP-seq assays in CRC cells treated with ATF1 overexpression in order to dissect the target programs of ATF1 . Results showed that ATF1 activates a subset of genes, including BRAF , NRAS , MYC , BIRC2 , DAAM1 , MAML2 , STAT1 , ID1 , and NKD2 , related to apoptosis, Wnt, TGF-β, and MAPK pathways, and these effects could cooperatively increase the risk of CRC. These findings reveal the clinical potential of ATF1 in CRC development and illuminate a promoter-enhancer interaction module between the ATF1 regulatory elements dbSNP: rs61926301 and dbSNP: rs7959129, and they bring us closer to understanding the molecular drivers of cancer.