Promoter Hypomethylation Is Responsible For Upregulated Expression Of MiR-21-5p In Clear Cell Renal Cell Carcinoma

Objective:renal clear cell carcinoma(ccRCC)has seriously threatened human health.miR-21 is an oncogenic miRNA which is upregulated in many solid tumors.However,the effect of miR-21 hypomethylation on miR-21-5p expression in cancers and the mechanism of miR-21-5p DNA demethylation remains unclear.Methods:The expression level of miR-21-5p and methylation of CpG locus in promoter region were evaluated by bioinformatics analysis using The Cancer Genome Atlas(TCGA),dbDEMC and starBase.The correlation between expression of miR-21-5p and prognosis of patients was performed by the OncoLnc.The expression and methylation level of miR-21-5p were analyzed by in microRNA quantitative PCR and DNA methylation pyrophosphate sequencing in ccRCC specimens.The expression of miR21-5p was induced upon treatment with 5-Aza-dC in cell.qRT-PCR was used to evaluate the expression of miR-21-5p in ACHN transfected with siTET3 and siTDG.JASPAR predicts the possible binding of transcription factors at CpG locus.Results:We found that the mRNA expression of miR-21-5p in ccRCC both TCGA and clinical samples was consistently revealed to be significantly higher compared with that in normal tissues.Survival analysis from the OncoLnc revealed that the high expression of miR-21-5p was associated with a short overall survival.After the cell were treated with 5-Aza-dC,the expression of miR-21-5p was activated.In addition,the overexpression of miR-21-5p is caused by hypomethylation of two key CpG locus(cg04276626 and cg02515217)in the promoter region.qRT-PCR showed that both TET3 and TDG were highly expressed in renal cancer cell lines,while the expression of miR-21-5p was down-regulated when the expression of TET3 and or TDG was down-regulated in ACHN.Moreover,we found that the cg02515217 CpG loci in the promoter region of miR-21-5p may be the binding region of transcription factors CEBPB,MEIS3 and TEAD4.Conclusions:Promoter hypomethylation was responsible for upregulated expression of miR-21-5p in tumors,especially in ccRCC miR-21-5p could be used as potential biomarker for diagnosis and prognosis.