Abstract
BackgroundLiterature reports that mature microRNA (miRNA) can be methylated at adenosine, guanosine and cytosine. However, the molecular mechanisms involved in cytosine methylation of miRNAs have not yet been fully elucidated. Here we investigated the biological role and underlying mechanism of cytosine methylation in miRNAs in glioblastoma multiforme (GBM).MethodsRNA immunoprecipitation with the anti-5methylcytosine (5mC) antibody followed by Array, ELISA, dot blot, incorporation of a radio-labelled methyl group in miRNA, and miRNA bisulfite sequencing were perfomred to detect the cytosine methylation in mature miRNA. Cross-Linking immunoprecipiation qPCR, transfection with methylation/unmethylated mimic miRNA, luciferase promoter reporter plasmid, Biotin-tagged 3’UTR/mRNA or miRNA experiments and in vivo assays were used to investigate the role of methylated miRNAs. Finally, the prognostic value of methylated miRNAs was analyzed in a cohorte of GBM pateints.ResultsOur study reveals that a significant fraction of miRNAs contains 5mC. Cellular experiments show that DNMT3A/AGO4 methylated miRNAs at cytosine residues inhibit the formation of miRNA/mRNA duplex and leading to the loss of their repressive function towards gene expression. In vivo experiments show that cytosine-methylation of miRNA abolishes the tumor suppressor function of miRNA-181a-5p miRNA for example. Our study also reveals that cytosine-methylation of miRNA-181a-5p results is associated a poor prognosis in GBM patients.ConclusionTogether, our results indicate that the DNMT3A/AGO4-mediated cytosine methylation of miRNA negatively.Graphical abstract
Highlights
Literature reports that mature microRNA can be methylated at adenosine, guanosine and cytosine
We extended our study to a cellular system in which siRNA-DNA methyltransferase 3A (DNMT3A), siRNA-AGO4 and αAGO41–164 were used to decrease the integrity of the DNMT3A/AGO4 complex
Cytosine-methylation of miRNA-181a-5p abolishes the formation of the miRNA-181a-5p-3’untranslated region (UTR)/BIM duplex We studied the formation of miRNA-messenger RNAs (mRNAs) duplex by performing biotin-tagged miRNA experiments [22, 23]
Summary
Literature reports that mature microRNA (miRNA) can be methylated at adenosine, guanosine and cytosine. MiRNAs target the RNA interference effector complex (RISC) of specific messenger RNAs (mRNAs) through partial base paring of sequences found predominantly in the 3’ untranslated region (UTR) of the gene. This reaction in turn increases the degradation of the mRNA and/or decreases its translation [3]. Several studies have shown that epigenetic changes in the promoter or coding region of miRNAs regulate their expression and the whole gene expression profile [6, 7] In addition to this regulation, miRNAs can be regulation via mechanims of base or phosphate modifications. The undoubtable importance of this first report concerning the cytosine methylation of miRNAs, many scientific questions concerning cytosine methylation remaind unanswered: how cytosine methylation of miRNAs impacts their functionality? What is the molecular player of miRNAs cytosine methylation?
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
