RecFOR proteins target RecA protein to a DNA gap with either DNA or RNA at the 5' terminus: implication for repair of stalled replication forks.

The repair of single-stranded gaps in duplex DNA by homologous recombination requires the proteins of the RecF pathway. The assembly of RecA protein onto gapped DNA (gDNA) that is complexed with the single-stranded DNA-binding protein is accelerated by the RecF, RecO, and RecR (RecFOR) proteins. Here, we show the RecFOR proteins specifically target RecA protein to gDNA even in the presence of a thousand-fold excess of single-stranded DNA (ssDNA). The binding constant of RecF protein, in the presence of the RecOR proteins, to the junction of ssDNA and dsDNA within a gap is 1-2 nm, suggesting that a few RecF molecules in the cell are sufficient to recognize gDNA. We also found that the nucleation of a RecA filament on gDNA in the presence of the RecFOR proteins occurs at a faster rate than filament elongation, resulting in a RecA nucleoprotein filament on ssDNA for 1000-2000 nucleotides downstream (5' → 3') of the junction with duplex DNA. Thus, RecA loading by RecFOR is localized to a region close to a junction. RecFOR proteins also recognize RNA at the 5'-end of an RNA-DNA junction within an ssDNA gap, which is compatible with their role in the repair of lagging strand gaps at stalled replication forks.