Wright et al. Supplemental Figures
Figure S1. Alternative Synthesis-Dependent Strand Annealing model.Synthesis-Dependent Strand Annealing (SDSA) proceeds as depicted in Figure 1 with the difference being that the second end is engaged not by strand annealing but through a second DNA strand invasion step. This model does not require a protein that anneals RPA-coated ssDNA and can rationalize efficient SDSA-mediated HR without RAD52, the only protein presently known to catalyze the annealing of RPA-coated ssDNA. Such a mechanism may require an asymmetry in DSB resection (left end: more resection, right end: less resection), which could be mediated through the elaborate DSB resection control in vertebrates. The second invasion intermediate is further processed to a non-crossover outcome.
Figure S2. Second DSB end invasion in Synthesis-Dependent Strand Annealing and double Holliday junction maturation. Both ends of the broken DNA molecule can form extended D-loops at different times and be disrupted to still achieve repair by Synthesis-Dependent Strand Annealing (SDSA). However, simultaneous invasion of both ends in the donor will risk double Holliday junction (dHJ) formation and consequent crossovers. If both ends initiate DNA synthesis and extend the hDNA regions, the structure may become a dHJ as shown. Disruption activities keep this likelihood low, and SDSA may still occur if the D-loops are disrupted any time before ligation of the nicked junctions.
Figure S3. Crossover formation. The black arrowheads indicate cleavage sites of endonuclease or resolvases on joint molecules. A, Depending on whether the second Holliday junction cleavage occurs in the same plane or not, the double Holliday junction (dHJ) intermediate is resolved into a crossover or non-crossover product. B, Model for structure-selective endonucleases processing of a D-loop into a crossover product without a dHJ intermediate. Cleavage on the donor strand leads to a nicked D-loop. Extension of the nicked D-loop, second end capture, and second end DNA synthesis generate a half-junction as shown. A second cleavage site of the half-junction results in a fully reciprocal crossover.
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