Effect Of PIWIL1 Gene On Biological Behavior Of Human Colon Cancer Cells

Aims In the Drosophila ovary,PIWI is required in somatic signaling cells,which is specifically expressed in testis and plays important roles in stem cell self-renew,RNA silencing,and translational regulation in diverse organisms.PIWIL1 encodes a member of the PIWI subfamily.Although recent researches have showed that PIWIL1 is specifically expressed in tumors and associated with tumor development and growth,its biological function is not well known.In this study,colon carcinoma cell line SW620 is used as object,the objective is to reveal the biological function of PIWIL1 in the development of tumor at the molecular level.Methods In order to study the differential expression patterns of PIWI gene family,we used Real-time PCR to detect the mRNA level of PIWIL1,PIWIL2,PIWIL3 and PIWIL4 in human colon cancer cell line SW620.RNA interfering technology was applied to silence the expression of PIWIL1 in colon carcinoma cell line SW620 and the effects on cells’ biological function were observed.Then,the plasmid which expresses hairpin siRNA for PIWEL1 gene was constructed and transfected into SW620 cells via lipofectamine 2000.The RNA interference plasmid efficiency which can be detected by Western blot.MTT assay,colony formation assay,monolayer wound assay,aggregation experiment and transwell assay were used to study the effect of PIWL1-shRNA on cell viability,proliferation capacity,adhesion,movement and invasion in SW620 cells.To detect the effect of PIWIL1 silencing on the sensitivity of SW620 to 5-FU,MTT assay was employed to detect the inhibition rate of 5-FU.Results The PIWIL1 mRNA level was higher in SW620 cells compared to PIWIL2,PIWIL3 and PIWIL4.The recombinant vector which expresses hairpin siRNA for PIWIL1 gene was successfully constructed as the results of endonuclease digesting.The Western blot showed that the PIWIL1 shRNA expression vector markedly suppressed PIWIL1 expression in the level of protein.In PIWTL1-depleted SW620 cells,the cell viability and proliferation were inhibited and the invasive ability was also significantly reduced.However,the experiment showed a significant increase of cell aggregation in PIWIL1-shRNA transfected cells compared with un-transfecterd cells or control vector transfected cells.The down-regulation of PIWIL1 increased the inhibition rate of 5-Fu.Conclusion Above the results primarily demonstrated that the recombinant PIWIL1 shRNA expression vector can reduced the level of PIWIL1 expression by RNAi effectively.The down regulation of PIWIL1 can inhibit the growth of colon cancer cells and increase the sensitivity of PIWIL1 to 5-FU.The recombinant vector could increase the capability of cell-cell adhesion and reduce the invasive ability of cell.PIWIL1 may play an important role in the pathogenesis of human colon cancer.