Cytosine methylation of mature microRNAs inhibits their functions and is associated with poor prognosis in glioblastoma multiforme

Type Article
Date 2020-02
Language English
Author(s) Cheray Mathilde1, 2, Etcheverry Amandine3, 4, 5, 6, Jacques Camille7, Pacaud Romain1, 2, 8, Bougras-Cartron Gwenola1, 8, Aubry Marc3, 4, 5, Denoual Florent6, Peterlongo Pierre9, Nadaradjane Arulraj2, Briand Josephine2, 8, Akcha Farida10, Heymann Dominique2, 8, Vallette François M.1, 2, 8, Mosser Jean3, 4, 5, 6, Ory Benjamin7, Cartron Pierre-François1, 2, 8, 11
Affiliation(s) 1 : RCINA, INSERM, Université de Nantes, Nantes, France
2 : Faculté de Médecine, Université de Nantes, Nantes, France
3 : CNRS, UMR 6290, Institut de Génétique et Développement de Rennes (IGdR), F-35043, Rennes, France
4 : Université Rennes1, UEB, UMS 3480 Biosit, Faculté de Médecine, F-35043, Rennes, France
5 : Plate-forme Génomique Environnementale et Humaine Biosit, Université Rennes1, F-35043, Rennes, France
6 : CHU Rennes, Service de Génétique Moléculaire et Génomique, F-35033, Rennes, France
7 : INSERM, UMR 1238, équipe labellisée ligue 2012, 1 Rue Gaston Veil, 44035, Nantes, France
8 : LaBCT, Institut de Cancérologie de l’Ouest, Saint Herblain, France
9 : IRISA Inria Rennes Bretagne Atlantique, équipe GenScale, Campus de Beaulieu, 35042, Rennes, France
10 : Ifremer, Laboratoire d’Ecotoxicologie, Rue de l’Ile d’Yeu, BP21105, cedex 03 44311, Nantes, France
11 : Institut de Cancérologie de l’Ouest, CRCINA INSERM U1232, Equipe 9 –Apoptose et Progression tumorale, LaBCT, Boulevard du Pr J Monod, 44805, Saint-Herblain, France
Source Molecular Cancer (1476-4598) (Springer Science and Business Media LLC), 2020-02 , Vol. 19 , N. 1 , P. 36 (16p.)
DOI 10.1186/s12943-020-01155-z
WOS© Times Cited 50
Keyword(s) miRNA, Cytosine-methylation, Epigenetics, DNMT3A, AGO4, Glioblastoma, Epitranscriptomics
Abstract Background Literature 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). Methods RNA 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. Results Our 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. Conclusion Together, our results indicate that the DNMT3A/AGO4-mediated cytosine methylation of miRNA negatively.
Full Text
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Publisher's official version 16 2 MB Open access
Additional file 1. Schematic representation of BS experiments. 1 28 KB Open access
Additional file 2. Supplementary figures and tables. 22 8 MB Open access
Additional file 3. Statistical analysis of BS experiments. 11 KB Open access
Additional file 4. Results of BS experiments. 226 KB Open access
Additional file 5. Sequence of selected miRNA-BS. 177 KB Open access
Additional file 6. Results of selected miRNA-BS. 146 KB Open access
Additional file 7. Results of 5mC-RIP-Array analyses. 66 KB Open access
Additional file 8. Extended experimental procedures. 7 647 KB Open access
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Cheray Mathilde, Etcheverry Amandine, Jacques Camille, Pacaud Romain, Bougras-Cartron Gwenola, Aubry Marc, Denoual Florent, Peterlongo Pierre, Nadaradjane Arulraj, Briand Josephine, Akcha Farida, Heymann Dominique, Vallette François M., Mosser Jean, Ory Benjamin, Cartron Pierre-François (2020). Cytosine methylation of mature microRNAs inhibits their functions and is associated with poor prognosis in glioblastoma multiforme. Molecular Cancer, 19(1), 36 (16p.). Publisher's official version : , Open Access version :