Rescue of stalled replication forks by RecG: Simultaneous translocation onthe leading and lagging strand templates supports an active DNA unwinding model of fork reversal and Holliday junction formation

Modification of damaged replication forks is emerging as a crucial factor f or efficient chromosomal duplication and the avoidance of genetic instabili ty. The RecG helicase of Escherichia coil, which is involved in recombinati on and DNA repair, has been postulated to act on stalled replication forks to promote replication restart via the formation of a four-stranded (Hollid ay) junction. Here we show that RecC can actively unwind the leading and ta gging strand arms of model replication fork structures in vitro. Unwinding is achieved in each case by simultaneous interaction with and translocation along both the leading and lagging strand templates at a fork. Disruption of either of these interactions dramatically inhibits unwinding of the oppo sing duplex arm. Thus, RecG translocates simultaneously along two DNA stran ds, one with 5 ' -3 ' and the otherwith 3 ' -5 ' polarity. The unwinding of both nascent strands at a damaged fork, and their subsequent annealing to form a Holliday junction, may explain the ability of RecC to promote replic ation restart. Moreover, the preferential binding of partial forks lacking a leading strand suggests that RecC may have the ability to target stalled replication intermediates in vivo in which lagging strand synthesis has con tinued beyond the leading strand.