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Methods Mol Biol. 2012;815:91-102. doi: 10.1007/978-1-61779-424-7_8.

Detection of RNA editing events in human cells using high-throughput sequencing.

Author information

1
Laboratory of Molecular Immunology, National Heart, Lung and Blood Institute, National Institutes of Health, Bethesda, MD 20892, USA. chepelevi@mail.nih.gov

Abstract

RNA editing can lead to amino acid substitutions in protein sequences, alternative pre-mRNA splicing, and changes in gene expression levels. The exact in vivo modes of interaction of the RNA editing enzymes with their targets are not well understood. Alterations in RNA editing have been linked to various human disorders and the improved understanding of the editing mechanism and specificity can explain the phenotypes that result from misregulation of RNA editing. Unbiased high-throughput methods of detection of RNA editing events genome-wide in human cells are necessary for the task of deciphering the RNA editing regulatory code. With the rapidly falling cost of genome resequencing, the future method of choice for the detection of RNA editing events will be whole-genome gDNA and cDNA sequencing. We describe a detailed procedure for the computational identification of RNA editing targets using the data from the deep sequencing of DNA and RNA from the peripheral blood mononuclear cells of a human individual with severe hemophilia A who is resistant to HIV infection. Interestingly, we find that mRNAs of the cyclin-dependent kinase CDK13 and the DNA repair enzyme NEIL1 undergo extensive A → I RNA editing that leads to amino acid substitutions in protein sequences.

PMID:
22130986
PMCID:
PMC4184133
DOI:
10.1007/978-1-61779-424-7_8
[Indexed for MEDLINE]
Free PMC Article
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