Abstract：The regulatory network constituted by transcription factors, enhancers, and microRNAs (miRNAs) is critical to the transcriptional regulation and progression of cancer. TEA domain transcription factor 1 (TEAD1) is a member of the TEA/ATTS domain family. However, it is unclear whether TEAD1, as a cancer-related transcription factor, is involved in the enhancer-miRNA network and the occurrence and develop-ment of hepatocellular carcinoma. Herein, 14 286 active enhancers that transcribe stable and unstable enhan-cer RNAs (eRNAs) were firstly identified by integrating CAGE-seq and GRO-seq data of HepG2 cell line, and confirmed to have previously reported histone modification characteristics (high H3K27ac signal and H3K4me1/H3K4me3 signal ratio). Subsequently, 2 550 TEAD1-binding enhancers (EnhTEAD1) were identified by in-tegrating 35 883 TEAD1-DNA binding sites obtained from ChIP-seq data. Compared with no TEAD1-binding enhancers (EnhnoTEAD1), the expression of active enhancer markers (H3K27ac, H3K4me1 and H3K4me3) and eRNAs on EnhTEAD1 was significantly increased. Furthermore, TEAD1 may act synergistically with four transcription factors (GATA4, HNF4A, YY1, and CTCF) to promote chromatin accessibility and spatial cyclization mediated by the EnhTEAD1 region. To study the regulatory network between EnhTEAD1 and miRNAs, small RNA sequencing was performed in HepG2 cells after TEAD1 was interfered using small in-terfering RNAs (siRNAs). A total of 68 differentially expressed miRNAs (EnhTEAD1-miRNAs) regulated by EnhTEAD1 were obtained by RNA-seq and Hi-C, and were found to be significantly involved in a variety of cancer-related biological processes and pathways. In summary, the study elucidates the regulatory mecha-nism of EnhTEAD1-miRNA network in hepatocellular carcinoma, and also provides a new potential target for treatment of hepatocellular carcinoma.
