STAT3-Mediated Promoter-Enhancer Interaction Up-Regulates Inhibitor of DNA Binding 1 (ID1) to Promote Colon Cancer Progression

Abstract

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