MicroRNA-346 Promotes The Growth Of Human Cervical Cancer Cells By Targeting HTERT

[Objective]MicroRNAs(miRNAs)are a class of small non-coding RNAs that include 18-26 nucleotides,which post-transcriptionally regulate gene expression.MiRNAs are involved in many physiological and pathological progress?Recent evidence indicates that many miRNAs function as oncogenes or tumor suppressors and play an important role in cancer initiation and progression by regulating their target genes.Using real time PCR method,we found that microRNA-346(miR-346)were significantly up-regulated in human cervical cancer tissue samples.And then,we focused on the effects of miR-346 on the phenotypes of cervical cancer cells as well as the identification of their direct target genes,in order to illuminate the molecular mechanisms of miR-346 in the initiation and progression of cervical cancer.[Methods]We detected the differencial expression of miR-346 in human cervical cancer tissue and adjacent normal tissues by Real-time PCR assay.miR-346 were overexpressed in cervical cancer tissue,and the changes of cell phenotypes were detected both with MTT assay and colony formation assay in HeLa cells and C33A cells.Subsequently,we combined bioinformatics analysis and identified the candidate target genes for miR-346.Then fluorescent reporter assay was performed to confirm the reliability of the direct target genes.Furthermore,the mRNA levels and protein levels of target genes in miR-346 overexpressed cervical cancer cells or tissues were detected with Real-time PCR and Western blot,in order to confirm the regulative role of miR-346 on hTERT expression.Finally,changes of cell phenotypes with MTT assay and colony formation assay by miR-346 was rescued by overexpression of hTERT.[Results]Real-time PCR results showed that the expression of miR-346 in human cervical cancer tissues were higher than those in adjacent normal tissues.We report that after overexpression of miR-346,the proliferation activity and the colony formation activity of human cervical cancer cells were both enhanced,meanwhile,blocking of it cause the opposite effects.Subsequently,we identified oncogene hTERT as congenerous candidate target genes for miR-346.The hTERT mRNA 3'untranslated region(3'UTR)contains the potential binding site of miR-346.The fluorescent reporter assay also confirmed that miR-346 can directly bind to the specific site of hTERT mRNA 3'UTR and positively regulate the gene expression.When miR-346 function was overexpressed in cervical cancer cell HeLa cells,mRNA level and protein level of hTERT were both increased,and mRNA level of hTERT in human cervical cancer tissues were higher than those in adjacent normal tissues.We also discoved that when the target gene was knocked down,the proliferation activity and the colony formation activity of HeLa cells were inhibited apparently.And with the overexpression of hTERT without its UTR,the changed phonetype of HeLa cells was restored.[Conclusions]Our results show that in cervical cancer cells,miR-346 functions as tumor oncogenes and promote cell proliferation.The high-expression of miR-346 in cervical cancer cells lead to an abnormally high expression level of oncogene hTERT and result in the activation of cell proliferation.At the same time,our conclusions take a chanllege to the tranditional regulation function of miRNA,it might be more complicated than we learned.General speaking,the elucidation of the mechanisms of miR-346 in cervical cancer helps us to further understand the mechanism of cervical cancer initiation and progression,and pave a way for clinical diagnosis and therapy of cervical cancer.