Abstract
Cytidine base editors, composed of a cytidine deaminase fused to Cas9 nickase, enable efficient C-to-T conversion in various organisms. However, current base editors suffer from severe trade-off between editing efficiency and precision. Here, based on rationally mutated cytidine deaminase domain, we develop a new base editor, YFE-BE4max, effectively narrow the editing width to as little as approximately three nucleotides while maintaining high efficiency in rabbits. Moreover, YFE-BE4max successfully mediated the Tyr p. Q68Stop and Lmna p. G607G mutation in F0 rabbit with high efficiency and precision, which precisely recapitulates the pathological features of human OCA1 and HGPS, respectively. Collectively, YFE-BE4max system provide promising tools to perform efficient base editing with high precision in rabbits and enhances its capacity to precisely model human diseases.
Highlights
The clustered regularly interspaced short palindromic repeat (CRISPR) system has exhibited powerful genome manipulation capability in various organisms[1,2]
base editor 3 (BE3) can potentially induce unwanted C-to-T substitutions when more than one C is present in the large ~5nucleotide window, which can negatively affect the precision of targeted base editing
Our previous study demonstrated that BE3 system can be used to mimic human pathologies by efficiently mediate C-to-T conversions in rabbits[6]
Summary
The clustered regularly interspaced short palindromic repeat (CRISPR) system has exhibited powerful genome manipulation capability in various organisms[1,2]. BE3 can potentially induce unwanted C-to-T substitutions when more than one C is present in the large ~5nucleotide window, which can negatively affect the precision of targeted base editing. Such system is not ideal for precise disease modeling and gene therapy when accurate single C substitution is required. To overcome this limitation, Liu et al optimized rA1 with mutant deaminase domains (YE base editors), representatively termed YE1 (W90Y + R126E) and YEE (W90Y + R126E + R132E), to effectively narrow the width of the editing window from ~5 nucleotides to as little as 1–2 nucleotides in human cells[4]. YEE-BE3 showed better accuracy than YE1-BE3, but has lowered editing efficiency at target loci[4,5]
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