Background: High expression of inhibitor of DNA binding 1 (ID1) correlates with poor prognosis in colorectal cancer (CRC). Aberrant enhancer activation in regulatingID1transcription is limited. Methods: Immunohistochemistry (IHC), quantitative RT-PCR (RT-qPCR) and Western blotting (WB) were used to determine the expression ofID1. CRISPR-Cas9 was used to generateID1or enhancer E1 knockout cell lines. Dual-luciferase reporter assay, chromosome conformation capture assay and ChIP-qPCR were used to determine the active enhancers ofID1. Cell Counting Kit 8, colony-forming, transwell assays and tumorigenicity in nude mice were used to investigate the biological functions ofID1and enhancer E1. Results: Human CRC tissues and cell lines expressed a higher level ofID1than normal controls.ID1promoted CRC cell proliferation and colony formation. Enhancer E1 actively regulatedID1promoter activity. Signal transducer and activator of transcription 3 (STAT3) bound toID1promoter and enhancer E1 to regulate their activity. The inhibitor of STAT3 Stattic attenuatedID1promoter and enhancer E1 activity and the expression ofID1. Enhancer E1 knockout down-regulatedID1expression level and cell proliferation in vitro and in vivo. Conclusions: Enhancer E1 is positively regulated by STAT3 and contributes to the regulation ofID1to promote CRC cell progression and might be a potential target for anti-CRC drug studies.