Integrated Enhancer Regulatory Network by Enhancer-Promoter Looping in Gastric Cancer (RNA-seq)

Enhancer as a cis-regulatory element (CRE), plays a critical role in gene regulation at many stages of cell growth. Meanwhile, enhancers in cancer cells also act as a regulator controlling oncogene transcription. In this study, we did a comprehensive analysis of long-range chromatin interactions, histone modifications, chromatin status, and expressions of two gastric cancer (GC) cell lines, compared with a human normal epithelial cell line. We found that GC-specific enhancers marked by histone modifications in gastric cancer can activate a population of genes including some oncogenes by enhancer-promoter (E-P) interactions. In addition, motif analysis of E-P interacted enhancers showed that three GC-specific TF sequences may regulate enhancer-interacting gene activation. We found MYB which is crucial for GC cellular growth and is activated by TCF7 regulated E-P loop. To validate this mechanism, we applied CRISPRi (dCas9-KRAB) induced transcriptional regulatory element inactivation in our target enhancer, the growth assay resulted in significant inhibition of cell growth in two GC cell lines. In conclusion, we found oncogene activation by aberrant loop formation that contributes to tumorigenesis in GC. To investigate the function of MYB and TCF7 in gastric cancer cells, we obtained MYB and TCF7 knockdown SNU719 and YCC10 gastric cancer cell lines by siRNA and CRISPRi We then performed gene expression profiling analysis using data obtained from RNA-seq of before and after MYB or TCF7 knockdown in two cell lines (SNU719_siCtrl, SNU719_siMYB, YCC10_siCtrl, YCC10_siMYB; SNU719_siCtrl, SNU719_siTCF7, YCC10_siCtrl, YCC10_siTCF7; SNU719_CRISPRi_sgCtrl, SNU719_CRISPRi_sgMYB-E3, YCC10_CRISPRi_sgCtrl, YCC10_CRISPRi_sgMYB-E3).