Fig. 1.

Conditions that influence indel outcomes from DSB EJ: structures of the DNA ends, processing and synapsis of the ends, and ligation. Shown are DSB ends induced by therapeutic clastogens that are often not directly ligatable before end processing, as well DSBs induced by site-specific nucleases (e.g., Streptococcus pyogenes Cas9) that can be readily ligated, but are nonetheless prone to end processing. Shown is a model that end processing to yield blunt ends subsequently requires the C-NHEJ complex (i.e., LIG4 and associated co-factors) for repair. In contrast, end resection that generates 3’ ssDNA with flanking extensive microhomology forms the substrate for Polθ-mediated end synapsis and fill-in synthesis, thereby generating DNA nicks that could be repaired by any of the three ligases (LIG1, LIG3, and LIG4). Shown in the box is the definition of No Indel EJ, which can be detected in genetic assays by examining repair of distal ends from two tandem Cas9 blunt-ended DSBs.