Short oligomers rather than rings of human RAD52 promote single-strand annealing

Genome maintenance and stability rely on the repair of DNA double-strand breaks. The break repair can be mediated by the single-strand annealing protein RAD52. RAD52 forms rings that are thought to promote annealing. However, RAD52s annealing activity decreases with increasing concentrations that favor ring formation. Thus, which oligomeric form and how RAD52 anneals DNA strands and detects sequence homology is unclear. We combine mass photometry with biochemical assays to quantify oligomeric states of human RAD52 with and without DNA and put forward an alternative mechanism illustrating the critical role of short oligomers for single-stranded DNA annealing. We found that while truncated RAD52 formed undecameric rings at nanomolar concentrations, full-length RAD52 was mostly monomeric at lower nanomolar, physiological concentrations. At higher concentrations, it formed rings with a variable stoichiometry from heptamers to tridecamers. At low concentrations, with hardly any rings present, RAD52 already promoted single-strand annealing. Rings and short oligomers could bind at least two single DNA strands, but if complementary strands were both bound to rings annealing was inhibited. Our findings suggest that single-strand annealing and homology detection is mediated by short oligomers of RAD52 instead of rings.