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Open AccessArticle

Allele-Specific Knockout by CRISPR/Cas to Treat Autosomal Dominant Retinitis Pigmentosa Caused by the G56R Mutation in NR2E3

1
INM, University of Montpellier, Inserm, 34091 Montpellier, France
2
National Reference Centre for Inherited Sensory Diseases, University of Montpellier, CHU, 34295 Montpellier, France
*
Author to whom correspondence should be addressed.
Academic Editor: Jan Wijnholds
Int. J. Mol. Sci. 2021, 22(5), 2607; https://doi.org/10.3390/ijms22052607
Received: 3 February 2021 / Revised: 1 March 2021 / Accepted: 3 March 2021 / Published: 5 March 2021
Retinitis pigmentosa (RP) is an inherited retinal dystrophy that causes progressive vision loss. The G56R mutation in NR2E3 is the second most common mutation causing autosomal dominant (ad) RP, a transcription factor that is essential for photoreceptor development and maintenance. The G56R variant is exclusively responsible for all cases of NR2E3-associated adRP. Currently, there is no treatment for NR2E3-related or, other, adRP, but genome editing holds promise. A pertinent approach would be to specifically knockout the dominant mutant allele, so that the wild type allele can perform unhindered. In this study, we developed a CRISPR/Cas strategy to specifically knockout the mutant G56R allele of NR2E3 and performed a proof-of-concept study in induced pluripotent stem cells (iPSCs) of an adRP patient. We demonstrate allele-specific knockout of the mutant G56R allele in the absence of off-target events. Furthermore, we validated this knockout strategy in an exogenous overexpression system. Accordingly, the mutant G56R-CRISPR protein was truncated and mis-localized to the cytosol in contrast to the (peri)nuclear localizations of wild type or G56R NR2E3 proteins. Finally, we show, for the first time, that G56R iPSCs, as well as G56R-CRISPR iPSCs, can differentiate into NR2E3-expressing retinal organoids. Overall, we demonstrate that G56R allele-specific knockout by CRISPR/Cas could be a clinically relevant approach to treat NR2E3-associated adRP. View Full-Text
Keywords: inherited retinal dystrophies; autosomal dominant retinitis pigmentosa; rod-cone dystrophy; NR2E3; CRISPR/Cas; photoreceptor; allele-specific; knockout; iPSC; retinal organoids inherited retinal dystrophies; autosomal dominant retinitis pigmentosa; rod-cone dystrophy; NR2E3; CRISPR/Cas; photoreceptor; allele-specific; knockout; iPSC; retinal organoids
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MDPI and ACS Style

Diakatou, M.; Dubois, G.; Erkilic, N.; Sanjurjo-Soriano, C.; Meunier, I.; Kalatzis, V. Allele-Specific Knockout by CRISPR/Cas to Treat Autosomal Dominant Retinitis Pigmentosa Caused by the G56R Mutation in NR2E3. Int. J. Mol. Sci. 2021, 22, 2607. https://doi.org/10.3390/ijms22052607

AMA Style

Diakatou M, Dubois G, Erkilic N, Sanjurjo-Soriano C, Meunier I, Kalatzis V. Allele-Specific Knockout by CRISPR/Cas to Treat Autosomal Dominant Retinitis Pigmentosa Caused by the G56R Mutation in NR2E3. International Journal of Molecular Sciences. 2021; 22(5):2607. https://doi.org/10.3390/ijms22052607

Chicago/Turabian Style

Diakatou, Michalitsa; Dubois, Gregor; Erkilic, Nejla; Sanjurjo-Soriano, Carla; Meunier, Isabelle; Kalatzis, Vasiliki. 2021. "Allele-Specific Knockout by CRISPR/Cas to Treat Autosomal Dominant Retinitis Pigmentosa Caused by the G56R Mutation in NR2E3" Int. J. Mol. Sci. 22, no. 5: 2607. https://doi.org/10.3390/ijms22052607

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