Journal
NUCLEIC ACIDS RESEARCH
Volume 50, Issue 19, Pages 10857-10868Publisher
OXFORD UNIV PRESS
DOI: 10.1093/nar/gkac897
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Funding
- ProQR Therapeutics
- Rett Syndrome Research Trust
- United States Department of Agriculture National Institute of Food and Agriculture Hatch Grant [CA-D-MCB-2629-H]
- National Institutes of Health [F31CA265135-01]
- National Institute of General Medical Sciences from the National Institutes of Health [P30 GM124165]
- NIH-ORIP HEI grant [S10OD021527]
- DOE Office of Science [DE-AC02-06CH11357]
- Rett Syndrome Research Trust funding
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ADARs, which correct disease-causing mutations at the RNA level, can be targeted to sites in the transcriptome using complementary guide strands. However, ADARs have bias against editing adenosines with a guanosine 5' nearest neighbor, limiting its effectiveness. This study reveals that certain nucleosides and specific duplex structures can enhance ADAR editing efficiency at 5'-GA sites, providing insights for the design of therapeutic RNA editing strategies.
ADARs (adenosine deaminases acting on RNA) can be directed to sites in the transcriptome by complementary guide strands allowing for the correction of disease-causing mutations at the RNA level. However, ADARs show bias against editing adenosines with a guanosine 5 ' nearest neighbor (5 '-GA sites), limiting the scope of this approach. Earlier studies suggested this effect arises from a clash in the RNA minor groove involving the 2-amino group of the guanosine adjacent to an editing site. Here we show that nucleosides capable of pairing with guanosine in a syn conformation enhance editing for 5 '-GA sites. We describe the crystal structure of a fragment of human ADAR2 bound to RNA bearing a G:G pair adjacent to an editing site. The two guanosines form a G(syn):G(anti) pair solving the steric problem by flipping the 2-amino group of the guanosine adjacent to the editing site into the major groove. Also, duplexes with 2 '-deoxyadenosine and 3-deaza-2 '-deoxyadenosine displayed increased editing efficiency, suggesting the formation of a G(syn):AH(anti)(+) pair. This was supported by X-ray crystallography of an ADAR complex with RNA bearing a G:3-deaza dA pair. This study shows how non-Watson-Crick pairing in duplex RNA can facilitate ADAR editing enabling the design of next generation guide strands for therapeutic RNA editing.
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