Journal
NATURE METHODS
Volume 12, Issue 9, Pages 866-+Publisher
NATURE PUBLISHING GROUP
DOI: 10.1038/NMETH.3490
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Funding
- Sidaction
- French Agence Nationale de Recherches sur le SIDA et les Hepatites Virales (ANRS)
- ANRS
- Initiative d'excellence (IDEX), Par-dela les frontieres, l'Universite de Strasbourg
- US National Institutes of Health (NIH)
- Australian National Health and Medical Research Council (NHMRC)
- Australian Research Council (ARC)
- German Ministry for Education Science (BMBF) [031A307]
- Einstein Center for Mathematics Berlin (ECMath) [CH5]
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RNA regulates many biological processes; however, identifying functional RNA sequences and structures is complex and time-consuming. We introduce a method, mutational interference mapping experiment (MIME), to identify, at single-nucleotide resolution, the primary sequence and secondary structures of an RNA molecule that are crucial for its function. MIME is based on random mutagenesis of the RNA target followed by functional selection and next-generation sequencing. Our analytical approach allows the recovery of quantitative binding parameters and permits the identification of base-pairing partners directly from the sequencing data. We used this method to map the binding site of the human immunodeficiency virus-1 (HIHIV-1) Pr55(Gag) protein on the viral genomic RNA in vitro, and showed that, by analyzing permitted base-pairing patterns, we could model RNA structure motifs that are crucial for protein binding.
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