The Role Of LncRNA RET In Cancer Cell Damage Response Induced By Ionizing Radiation

Ionizing radiation is a risk factor of human health and life.Ionizing radiation causes a series of radiation toxicological effects such as cell growth changes,cell cycles disturbances,apoptosis or death,and DNA damage.Long non-coding RNA?LncRNA?refers to a class of non-coding RNAs that lack the ability to encode proteins and are greater than 200 nts in length.Initially,LncRNA was originally thought to be transcriptional noise,without any function.With the deepening of LncRNA research,it has been found that LncRNA participates in important processes in the life process.Furthermore,in recent studies,LncRNAs are also involved in cellular ionizing radiation injury responses.LOC90024 is a radiation-induced Lnc RNA that has been screened by our research lab,located on chromosome 4q24.We named it Lnc RET?Radiation-Elevated Transcript?.To date,Lnc RET's function has not been reported.Objective:To investigate the effect of LncRET on the biological function and its role in the radiation sensitivity of tumor cells.Methods:Real-time fluorescence quantitative PCR was used to detection whether Lnc RET was induced by ionizing radiation.Using Lnc RET-specific siRNA or lentivirus interference technology constructed He La and A549 cell lines.Real-time fluorescence quantitative PCR,MTT,flow cytometry,Western Blot,immunofluorescence techniques,wound healing assay,Transwell assays and cloning assays were used to test the effects of Knockout LncRET on tumor cell growth,cell cycle,apoptosis,migration and invasion,and cell radiation sensitivity,and furthermore to explored its possible molecular mechanisms.Results:The experimental results showed that after exposure to ⁶⁰Co?-rays in Hela and A549 cells,the LncRET content gradually increased with the irradiation time and dose.MTT assay and plate clone formation experiment showed that compared to NC group,the cell viability of knockdown LncRET has a significant reduce in He La and A549 cells;TDR double blocking method to detected the cell cycle,the results showed that the cell cycle arrest in the G2/M phase;Anncxin?-FITC/PI double marking flow cytometry assay showed that compared to NC group,the apoptosis of knockdown Lnc RET was significantly raised in He La and A549 cells;Western Blot results showed that compared to NC group,knockdown LncRET the expression of Bax was significantly reduce and Bcl-2 expression was increased in HeLa and A549 cells;Wound healing assay and Transwell assay showed that compared to NC group,the cell migration ability and the cell number went through membrane of knockdown LncRET has a significant reduce;Colony formation assay showed that compared to NC group,knockdown LncRET increased cell radiation sensitivity;Flow cytometry,?-H2AX immunoblot analysis,and epithelial mesenchymal transition marker assays showed knockdown LncRET can increase the radiation-induced apoptosis,reduce radiation-induced DNA double-strand break repair and reversal of radiation induced EMT;Western blot results showed that the expression of target protein PTEN increased after LncRET was silent;MTT assay,flow cytometry,and colony formation experiments show that under radiation stress,cell growth,apoptosis,and radiation sensitivity after knockout of PTEN are reversed compared to LncRET silencing,suggesting that LncRET influenced cell process is was regulated by PTEN.Conclusion:1.Down-regulation of Lnc RET expression in tumor can inhibit cells growth,arrest cell cycle,promote cells apoptosis and reduce migration and invasion ability of cells.2.Down-regulation of Lnc RET expression inhibits the growth of tumor cells under radiation pressure and increases the tumor cells radiosensitivity.3.LncRET Regulates Radiation Sensitivity of Tumor Cells by Regulating PTEN.