Replication fork integrity and intra-S phase checkpoint suppress gene amplification |
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Authors: | Anna Kondratova Takaaki Watanabe Michael Marotta Matthew Cannon Anca M Segall David Serre Hisashi Tanaka |
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Institution: | 1Department of Molecular Genetics, Cleveland Clinic Lerner Research Institute, Cleveland, OH, USA;2Department of Surgery, Cedars-Sinai Medical Center, West Hollywood, CA, USA;3Genomic Medicine Institute, Cleveland Clinic Lerner Research Institute, Cleveland, OH, USA;4Department of Biology, San Diego State University, San Diego, CA 92182, USA |
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Abstract: | Gene amplification is a phenotype-causing form of chromosome instability and is initiated by DNA double-strand breaks (DSBs). Cells with mutant p53 lose G1/S checkpoint and are permissive to gene amplification. In this study we show that mammalian cells become proficient for spontaneous gene amplification when the function of the DSB repair protein complex MRN (Mre11/Rad50/Nbs1) is impaired. Cells with impaired MRN complex experienced severe replication stress and gained substrates for gene amplification during replication, as evidenced by the increase of replication-associated single-stranded breaks that were converted to DSBs most likely through replication fork reversal. Impaired MRN complex directly compromised ATM/ATR-mediated checkpoints and allowed cells to progress through cell cycle in the presence of DSBs. Such compromised intra-S phase checkpoints promoted gene amplification independently from mutant p53. Finally, cells adapted to endogenous replication stress by globally suppressing genes for DNA replication and cell cycle progression. Our results indicate that the MRN complex suppresses gene amplification by stabilizing replication forks and by securing DNA damage response to replication-associated DSBs. |
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