Effect Of C-Myc On The Progression Of Oral Squamous Cell Carcinoma Through Lnc-CCDC117-1

Objective To illuminate the targeted regulatory relationship between oncogenic transcription factor c-Myc and long non-coding RNA(lnc RNAs)lnc-CCDC117-1,and investigate the effect of lnc-CCDC117-1 knockdown、overexpression on the development of oral squamous cell carcinoma.Methods Lnc RNAs regulated by c-Myc were preliminarily screened by gene chip technology to further verify the expression difference of lnc RNAs in oral squamous cell HN6.The regulatory effect of c-Myc on lnc-CCDC117-1 was preliminatively evaluated.Lentiviral vectors carrying flag-c-Myc,plko.1-shc-Myc and their controls were transfected into oral squamous cell HN6,SCC9,CAL27 respectively.The expression of lnc-CCDC117-1 was detected by quantitative real-time PCR(QRT-PCR).The promoter sequence of the 2000 bp region upstream of lnc-CCDC117-1 was queried by UCSC database,after predicting the possible c-Myc binding sites in this region with JASPAR database,a dual luciferase reporter system was established to verify the prediction results.In addition,the intracellular localization of lnc-CCDC117-1 was detected by fluorescence in situ hybridization.Based on this,overexpression vector and knockdown vector carrying lnc-CCDC117-1 was constructed separately and transfected into HN6,SCC9,CAL27 cells,the transfection efficiency was evaluated by using QRT-PCR.Then the cell proliferation of oral squamous cell carcinoma cells was detected by CCK-8 and colony formation assays.Results Lnc RNAs positively regulated by c-Myc were initially screened by gene chip technology.After overexpression or c-Myc knockdown in oral squamous cell HN6,these lnc RNAs were verified to be consistent with the results of the chip,and the expression difference of lnc-CCDC117-1 was found to be the most significant.Real-time fluorescence quantitative PCR test showed that c-Myc had a positive regulatory effect on lnc-CCDC117-1.When c-Myc was overexpressed,the expression of lnc-CCDC117-1could be significantly up-regulated.lnc-CCDC117-1 expression was significantly downregulated by c-Myc knockdown.Dual luciferase reporter genes showed that c-Myc could target the regulation of lnc-CCDC117-1,and c-Myc was involved in regulating and enhancing the transcriptional activity of lnc-CCDC117-1.Lnc-CCDC117-1 mainly existed in the nucleus of the cell.QRT-PCR results showed that the expression of lncCCDC117-1 and c-Myc was significantly decreased by shlnc-CCDC117-1.The expression of lnc-CCDC117-1 was significantly up-regulated by +lnc-CCDC117-1.The results of growth curve assay,CCK-8 assay,cell scratch assay and cloning formation showed that overexpression of lnc-CCDC117-1 significantly promoted the proliferation and migration of oral squamous cell carcinoma,while knockdown of lnc-CCDC117-1could inhibit the growth and proliferation of oral squamous cell carcinoma.Conclusion lnc-CCDC117-1 may be positively regulated by c-Myc.Overexpression of lnc-CCDC117-1 in HN6,SCC9 and CAL27 cells would promote cells proliferation,and otherwise will inhibit cells growth.