Effects Of Super-enhancer-driven Epha2 On The Biological Characteristics Of HCT-116 Cells

Super Enhancers(SEs)are highly active and cell type specific DNA sequences in the genome.They determine the fate of cells by regulating the temporal and spatial expression of cell type specific genes.Studies have shown that super enhancers can not only drive the transcriptional expression of non-coding RNAs(nc RNAs),and indirectly regulate biological functions,but also play a key role in the occurrence and development of tumors.In the previous study,a new super enhancer EphA2-SE that is active in a variety of tumors has been identified and authenticated.However,the specific mechanism of its component enhancers in colon cancer is still unknown.Therefore,this project further explores its function in the occurrence and development of colon cancer by establishing a knockout cell line of EphA2-SE component enhancer.First of all,this subject used CRISPR/Cas9 gene editing technology to construct a knockout cell line with EphA2-SE component enhancer deletion in HCT-116 cells.Through testing,it’s found that no off-target effects have occurred.The results of realtime quantitative PCR,Western Blot and immunofluorescence experiments showed that the absence of EphA2-SE and its component enhancers can reduce the expression of the target gene EphA2 in HCT-116 cells.The results of MTT colorimetry,cell plate cloning experiment and Transwell experiment showed that the absence of EphA2-SE and its component enhancers can inhibit the proliferation,migration and invasion of HCT-116 cells.In summary,EphA2-SE and its component enhancers mediate the biological characteristics of colon cancer HCT-116 cells by affecting the expression of the target gene EphA2.It lays a good foundation for further elucidating the functions of super enhancers in tumorigenesis and development.