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© Biologie structurale et chimie
Structure du domaine en doigt de zinc de la protéine NEMO, déterminée par Résonance magnétique nucléaire (RMN). Cette protéine jouant un rôle dans des maladies (cancer, inflammation), les connaissances acquises sur sa structure offrent de précieuses informations sur sa fonction.
Publication : Nature Communications

CtIP fusion to Cas9 enhances transgene integration by homology-dependent repair.

Scientific Fields
Diseases
Organisms
Applications
Technique

Published in Nature Communications - 19 Mar 2018

Charpentier M, Khedher AHY, Menoret S, Brion A, Lamribet K, Dardillac E, Boix C, Perrouault L, Tesson L, Geny S, De Cian A, Itier JM, Anegon I, Lopez B, Giovannangeli C and Concordet JP

Link to Pubmed [PMID] – 29556040

Nat Commun. 2018 Mar 19;9(1):1133

In genome editing with CRISPR–Cas9, transgene integration often remains challenging. Here, we present an approach for increasing the efficiency of transgene integration by homology- dependent repair (HDR). CtIP, a key protein in early steps of homologous recombination, is fused to Cas9 and stimulates transgene integration by HDR at the human AAVS1 safe harbor locus. A minimal N-terminal fragment of CtIP, designated HE for HDR enhancer, is sufficient to stimulate HDR and this depends on CDK phosphorylation sites and the multimerization domain essential for CtIP activity in homologous recombination. HDR stimulation by Cas9–HE, however, depends on the guide RNA used, a limitation that may be overcome by testing multiple guides to the locus of interest. The Cas9–HE fusion is simple to use and allows obtaining twofold or more efficient transgene integration than that with Cas9 in several experimental systems, including human cell lines, iPS cells, and rat zygotes.