Analysis of strand transfer and template switching mechanisms of DNA gap repair by homologous recombination in Escherichia coli: predominance of strand transfer.
Daughter strand gaps formed upon interruption of replication at DNA lesions in Escherichia coli can be repaired by either translesion DNA synthesis or homologous recombination (HR) repair. Using a plasmid-based assay system that enables discrimination between strand transfer and template switching (information copying) modes of HR gap repair, we found that approximately 80% of strand gaps were repaired by physical strand transfer from the donor, whereas approximately 20% appear to be repaired by template switching. HR gap repair operated on both small and bulky lesions and largely depended on RecA and RecF but not on the RecBCD nuclease. In addition, we found that HR was mildly reduced in cells lacking the RuvABC and RecG proteins involved in resolution of Holliday junctions. These results, obtained for the first time under conditions that detect the two HR gap repair mechanisms, provide in vivo high-resolution molecular evidence for the predominance of the strand transfer mechanism in HR gap repair. A small but significant portion of HR gap repair appears to occur via a template switching mechanism.
Izhar
Lior
L
Department of Biological Chemistry, Faculty of Biochemistry, Weizmann Institute of Science, Rehovot 76100, Israel.
Goldsmith
Moshe
M
Dahan
Ronny
R
Geacintov
Nicholas
N
Lloyd
Robert G
RG
Livneh
Zvi
Z
D001483:Base Sequence; D001564:Benzo(a)pyrene / Q000378:metabolism; D018736:DNA Adducts / Q000378:metabolism; D004260:DNA Repair; D045566:DNA, Cruciform / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics*; D029968:Escherichia coli Proteins / Q000378:metabolism; D043211:Exodeoxyribonuclease V / Q000378:metabolism; D006147:Guanine / Q000378:metabolism; D008954:Models, Biological; D009154:Mutation / Q000235:genetics; D010957:Plasmids; D011938:Rec A Recombinases / Q000378:metabolism; D011995:Recombination, Genetic / Q000235:genetics*; D013698:Templates, Genetic
D018736:DNA Adducts; D045566:DNA, Cruciform; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; C489340:recF protein, E coli; D001564:Benzo(a)pyrene; D006147:Guanine; D011938:Rec A Recombinases; D043211:Exodeoxyribonuclease V; C468322:exodeoxyribonuclease V, E coli