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Ultrastructural localization of 5-methylcytosine on DNA and RNA

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Abstract

DNA methylation is the major epigenetic modification and it is involved in the negative regulation of gene expression. Its alteration can lead to neoplastic transformation. Several biomolecular approaches are nowadays used to study this modification on DNA, but also on RNA molecules, which are known to play a role in different biological processes. RNA methylation is one of the most common RNA modifications and 5-methylcytosine presence has recently been suggested in mRNA. However, an analysis of nucleic acid methylation at electron microscope is still lacking. Therefore, we visualized DNA methylation status and RNA methylation sites in the interphase nucleus of HeLa cells and rat hepatocytes by ultrastructural immunocytochemistry and cytochemical staining. This approach represents an efficient alternative to study nucleic acid methylation. In particular, this ultrastructural method makes the visualization of this epigenetic modification on a single RNA molecule possible, thus overcoming the technical limitations for a (pre-)mRNA methylation analysis.

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Acknowledgements

The authors are grateful for the technical assistance of Ms. Francine Flach and Ms. Paola Veneroni and for the critical review of the manuscript of Prof. Carlo Pellicciari and Prof. Antonella Forlino.

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Authors and Affiliations

  1. Laboratory of Cell Biology and Neurobiology, Department of Biology and Biotechnology, University of Pavia, Via Ferrata 9, 27100, Pavia, Italy

    Irene Masiello & Marco Biggiogera

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  1. Irene Masiello
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  2. Marco Biggiogera
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Correspondence to Marco Biggiogera.

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Funding

This work was supported by Fondi di Ateneo per la Ricerca (F.A.R. 2013–2014) from the University of Pavia.

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The experiments described in this manuscript comply with the current Italian laws.

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The authors declare that no competing interests exist.

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Masiello, I., Biggiogera, M. Ultrastructural localization of 5-methylcytosine on DNA and RNA. Cell. Mol. Life Sci. 74, 3057–3064 (2017). https://doi.org/10.1007/s00018-017-2521-1

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