Abstract
SummaryAccurate DNA replication is essential to preserve genomic integrity and prevent chromosomal instability-associated diseases including cancer. Key to this process is the cells’ ability to stabilize and restart stalled replication forks. Here, we show that the EXD2 nuclease is essential to this process. EXD2 recruitment to stressed forks suppresses their degradation by restraining excessive fork regression. Accordingly, EXD2 deficiency leads to fork collapse, hypersensitivity to replication inhibitors, and genomic instability. Impeding fork regression by inactivation of SMARCAL1 or removal of RECQ1’s inhibition in EXD2−/− cells restores efficient fork restart and genome stability. Moreover, purified EXD2 efficiently processes substrates mimicking regressed forks. Thus, this work identifies a mechanism underpinned by EXD2’s nuclease activity, by which cells balance fork regression with fork restoration to maintain genome stability. Interestingly, from a clinical perspective, we discover that EXD2’s depletion is synthetic lethal with mutations in BRCA1/2, implying a non-redundant role in replication fork protection.
Highlights
Faithful duplication of the genome during cell division ensures accurate transmission of genetic information to daughter cells
EXD2 Is Recruited to Replication Forks following Replication Stress Recently, we have employed isolation of proteins on nascent DNA coupled with mass spectrometry to identify factors recruited to stalled replication forks (Higgs et al, 2015)
To test if EXD2 associates with replication forks, we performed an isolation of proteins on nascent DNA (iPOND) analysis coupled with a thymidine-chase
Summary
Faithful duplication of the genome during cell division ensures accurate transmission of genetic information to daughter cells. Replication fork stability is constantly challenged by damage to the DNA template or progression through chromosomal regions that are inherently difficult to replicate (Bhowmick and Hickson, 2017; Gaillard et al, 2015; Kolinjivadi et al, 2017a) These blockades can collapse replication forks, contributing to tumor progression by driving chromosomal instability (Burrell et al, 2013; Hills and Diffley, 2014; Jeggo et al, 2016; Kass et al, 2016; Zeman and Cimprich, 2014). The importance of these responses is highlighted by several cancer-predisposing human diseases caused by mutations in various proteins contributing to replication fork stability (e.g., Seckel, Bloom, Werner, or Fanconi anemia syndromes)
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