The Effect Of M~6A Methyilransferase METTL3 On The Biological Characteristics Of HCT-116 Cells

N6-methyl adenosine(m~6A)is the methylation of adenosine at the N6 site,which is the most common internal modification of eukaryotic m RNA.As a core component of m~6A methylation,METTL3 mediates the "Writers" process of m~6A methylation,which has been proved to play a carcinogenesis or cancer suppression role in a variety of cancers.METTL3 is a double-edged sword for cancer.As one of the top ten malignant tumors,colon cancer seriously threatens human health.Although current methods of surgery,chemotherapy and molecular targeted therapy have improved patient outcomes,there are still more than 1000000 new cases and approximately 600000 deaths worldwide each year.In order to study the effect of METTL3 on the biological characteristics of human colon cancer HCT-116,the METTL3 knockdown vector was constructed by using the CRISPR/Cas9 gene editing technology,transfected with colon cancer HCT-116 cells,the target clone was screened by limiting dilution,and established by Western Blot.METTL3 knocks down the cell line.In addition,by constructing a lentiviral vector overexpressing the METTL3 gene,the PLVSIN-METTL3,PLVSIN-EV vectors and helper plasmids were co-transfected into HEK-293 T cells to package the lentivirus.Colon cancer HCT-116 cell line was infected with the packaged virus,and METTL3 overexpression cell line was established by Western Blot screening.Using real-time quantitative PCR to detect the gene expression levels of the four cell lines WT-HCT-116 cell line,KD-METTL3 cell line,PLV-METTL3 cell line and PLV-EV cell line shows that the PLV-METTL3 cell line is significantly higher than Gene expression level of PLV-EV cell line.In order to detect the effect of METTL3 on the behavior of HCT-116 cells,the MTT cell proliferation experiment was used to find that for the WT-HCT-116 cell line and KD-METTL3 cell line,the value-added ability of the KD-METTL3 cell line was reduced;while for PLV-METTL3.For the cell line and the PLV-EV cell line,the PLV-METTL3 cell line has a high proliferative ability.In general,overexpression of METTL3 promotes the proliferation of colon cancer HCT-116 cells.In order to further study the effect of METTL3 on the cloning ability of cells,a cell clone formation experiment was used,which showed that after knocking down METTL3,it would inhibit the clonal formation of cells,that is,after downregulation of METTL3,it would inhibit the cloning ability of HCT-116 cells,namely HCT-116 cells Survivability is reduced.In summary,the down-regulation of METTL3 expression inhibits the proliferation and colony-forming ability of colon cancer HCT-116 cells,while up-regulation of METTL3 will promote the proliferation of colon cancer HCT-116 cells,thus preliminarily inferring METTL3 in human colon cancer HCT-116 plays the role of oncogene.