Repair of gaps opposite lesions by homologous recombination in mammalian cells.
Damages in the DNA template inhibit the progression of replication, which may cause single-stranded gaps. Such situations can be tolerated by translesion DNA synthesis (TLS), or by homology-dependent repair (HDR), which is based on transfer or copying of the missing information from the replicated sister chromatid. Whereas it is well established that TLS plays an important role in DNA damage tolerance in mammalian cells, it is unknown whether HDR operates in this process. Using a newly developed plasmid-based assay that distinguishes between the three mechanisms of DNA damage tolerance, we found that mammalian cells can efficiently utilize HDR to repair DNA gaps opposite an abasic site or benzo[a]pyrene adduct. The majority of these events occurred by a physical strand transfer (homologous recombination repair; HRR), rather than a template switch mechanism. Furthermore, cells deficient in either the human RAD51 recombination protein or NBS1, but not Rad18, exhibited decreased gap repair through HDR, indicating a role for these proteins in DNA damage tolerance. To our knowledge, this is the first direct evidence of gap-lesion repair via HDR in mammalian cells, providing further molecular insight into the potential activity of HDR in overcoming replication obstacles and maintaining genome stability.
Adar
Sheera
S
Department of Biological Chemistry, Weizmann Institute of Science, Rehovot 76100, Israel.
Izhar
Lior
L
Hendel
Ayal
A
Geacintov
Nicholas
N
Livneh
Zvi
Z
D000818:Animals; D018797:Cell Cycle Proteins / Q000502:physiology; D045744:Cell Line, Tumor; D002478:Cells, Cultured; D004247:DNA / Q000378:metabolism; D004249:DNA Damage; D004260:DNA Repair; D004268:DNA-Binding Proteins / Q000502:physiology; D006801:Humans; D051379:Mice; D009687:Nuclear Proteins / Q000502:physiology; D051135:Rad51 Recombinase / Q000502:physiology; D011995:Recombination, Genetic
D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; C112156:NBN protein, human; D009687:Nuclear Proteins; C485953:Rad18 protein, mouse; D004247:DNA; C495153:RAD51 protein, human; D051135:Rad51 Recombinase