Fig. 8: Pol θ-dependent repair is initiated at interrupted MH in vivo.

Examples of proposed mismatched MH used to generate POLQ-dependent deletions, derived from analysis of DSB repair at four CRISPR-Cas9 targets in a sequence derived from the human HBB gene¹². Target sites, deletion size and microhomologies derived from ref. ¹² are summarized in Table 1 for these examples and the full POLQ-dependent dataset. At the top of each panel, the local sequence environment is shown for the two 3ʹ single-stranded DNA ends to be joined. Matched bases in the MH are colored blue, and mismatched bases are colored red. Flanking unpaired bases are removed by nuclease action¹³. Extension by Pol θ is indicated by bases colored green. The internally mismatched bases will be resolved in the cell by mismatch repair or DNA replication, yielding the two outcomes shown at the bottom of each panel. a In target site 3, the same internally mismatched MH, with 3 paired bases, can give rise to two outcomes. These were previously scored as two different 10 bp deletions with MH lengths of 1 or 2. b In target site 2, the same internally mismatched MH, with five paired bases, can give rise to two outcomes. These were previously scored as two different 13 bp deletions with MH lengths of 1 and 4. c In target site 1, the same internally mismatched MH, with two paired bases, can give rise to two outcomes. These were previously scored as two different 4 bp deletions with MH lengths of 0 and 2. d In target site 1, the same internally mismatched MH, with nine paired bases, can give rise to two outcomes. These were previously scored as a POLQ-dependent 21 bp deletion with MH length of 4, and a POLQ-independent 21 bp deletion with MH length of 5.