The accumulation of MMS-induced single strand breaks in G1 phase is recombinogenic in DNA polymerase beta defective mammalian cells.
DNA polymerase (Pol) beta null mouse embryonic fibroblasts provide a useful cell system to investigate the effects of alterations in base excision repair (BER) on genome stability. These cells are characterized by hypersensitivity to the cytotoxic effects of methyl methanesulfonate (MMS) and by decreased repair of the MMS-induced DNA single strand breaks (SSB). Here, we show that, in the absence of Pol beta, SSB accumulate in G1 phase cells, accompanied by the formation of proliferating cell nuclear antigen foci in the nuclei. When replicating Pol beta null cells are treated with MMS, a rapid phosphorylation of histone H2AX is detected in the nuclei of S phase cells, indicating that double strand breaks (DSB) are formed in response to unrepaired SSB. This is followed by relocalization within the nuclei of Rad51 protein, which is essential for homologous recombination (HR). These findings are compatible with a model where, in mammalian cells, unrepaired SSB produced during BER are substrates for the HR pathway via DSB formation. This is an example of a coordinated effort of two different repair pathways, BER and HR, to protect mammalian cells from alkylation-induced cytotoxicity.
Pascucci
Barbara
B
Istituto di Cristallografia, CNR, Sezione di Roma PO Box 10, 00016 Monterotondo Stazione, Roma, Italy.
Russo
Maria Teresa
MT
Crescenzi
Marco
M
Bignami
Margherita
M
Dogliotti
Eugenia
E
D000818:Animals; D002843:Chromatin / Q000378:metabolism; D004249:DNA Damage; D019951:DNA Polymerase beta / Q000235:genetics*; D004260:DNA Repair; D004268:DNA-Binding Proteins / Q000032:analysis; D016193:G1 Phase / Q000187:drug effects / Q000235:genetics*; D017353:Gene Deletion; D042822:Genomic Instability; D006657:Histones / Q000032:analysis; D008741:Methyl Methanesulfonate / Q000633:toxicity; D051379:Mice; D009153:Mutagens / Q000633:toxicity; D018809:Proliferating Cell Nuclear Antigen / Q000378:metabolism; D051135:Rad51 Recombinase; D011995:Recombination, Genetic
D002843:Chromatin; D004268:DNA-Binding Proteins; D006657:Histones; D009153:Mutagens; D018809:Proliferating Cell Nuclear Antigen; C475961:gamma-H2AX protein, mouse; D008741:Methyl Methanesulfonate; D051135:Rad51 Recombinase; C495154:Rad51 protein, mouse; D019951:DNA Polymerase beta