The biological principles and advanced applications of DSB repair in CRISPR-mediated yeast genome editing

Abstract

To improve the performance of yeast cell factories for industrial production, extensive CRISPR-mediated genome editing systems have been applied by artificially creating double-strand breaks (DSBs) to introduce mutations with the assistance of intracellular DSB repair. Diverse strategies of DSB repair are required to meet various demands, including precise editing or random editing with customized gRNAs or a gRNA library. Although most yeasts remodeling techniques have shown rewarding performance in laboratory verification, industrial yeast strain manipulation relies only on very limited strategies. Here, we comprehensively reviewed the molecular mechanisms underlying recent industrial applications to provide new insights into DSB cleavage and repair pathways in both Saccharomyces cerevisiae and other unconventional yeast species. The discussion of DSB repair covers the most frequently used homologous recombination (HR) and nonhomologous end joining (NHEJ) strategies to the less well-studied illegitimate recombination (IR) pathways, such as single-strand annealing (SSA) and microhomology-mediated end joining (MMEJ). Various CRISPR-based genome editing tools and corresponding gene editing efficiencies are described. Finally, we summarize recently developed CRISPR-based strategies that use optimized DSB repair for genome-scale editing, providing a direction for further development of yeast genome editing.