Autophagy induction enhances homologous recombination-associated CRISPR–Cas9 gene editing

Abstract CRISPR (clustered regularly interspaced short palindromic repeats)–Cas9 (CRISPR-associated protein 9)-based gene editing via homologous recombination (HR) enables precise gene correction and insertion. However, its low efficiency poses a challenge due to the predominance of nonhomologous end-joining during DNA repair processes. Although numerous efforts have been made to boost HR efficiency, there remains a critical need to devise a novel method that can be universally applied across cell types and in vivo animals, which could ultimately facilitate therapeutic treatments. This study demonstrated that autophagy induction using different protocols, including nutrient deprivation or chemical treatment, significantly improved HR-associated gene editing at diverse genomic loci in mammalian cells. Notably, interacting cofactor proteins that bind to Cas9 under the autophagic condition have been identified, and autophagy induction could also enhance in vivo HR-associated gene editing in mice. These findings pave the way for effective gene correction or insertion for in vivo therapeutic treatments.