Substrate-Selective Repair and Restart of Replication Forks by DNA Translocases

• Published in: Cell reports (Cambridge) Vol. 3; no. 6; pp. 1958 - 1969
• Main Authors: Bétous, Rémy, Couch, Frank B., Mason, Aaron C., Eichman, Brandt F., Manosas, Maria, Cortez, David
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 27.06.2013; Elsevier
• Subjects: ADN; Animals; Baculoviridae - genetics; Cancer cells; Cèl·lules canceroses; DNA; DNA - biosynthesis; DNA - genetics; DNA - metabolism; DNA Damage; DNA Helicases - genetics; DNA Helicases - metabolism; DNA Repair; DNA Replication; Escherichia coli; Escheríchia coli; HEK293 Cells; Humans; Insecta - cytology; Insecta - virology; Life Sciences; Protein Binding; Proteins; Proteïnes; Reparació de l'ADN; Replication Origin; Templates, Genetic; Replication fork regression; DNA translocase; DNA replication stress
• ISSN: 2211-1247; 2211-1247
• DOI: 10.1016/j.celrep.2013.05.002

Stalled replication forks are sources of genetic instability. Multiple fork-remodeling enzymes are recruited to stalled forks, but how they work to promote fork restart is poorly understood. By combining ensemble biochemical assays and single-molecule studies with magnetic tweezers, we show that SMARCAL1 branch migration and DNA-annealing activities are directed by the single-stranded DNA-binding protein RPA to selectively regress stalled replication forks caused by blockage to the leading-strand polymerase and to restore normal replication forks with a lagging-strand gap. We unveil the molecular mechanisms by which RPA enforces SMARCAL1 substrate preference. E. coli RecG acts similarly to SMARCAL1 in the presence of E. coli SSB, whereas the highly related human protein ZRANB3 has different substrate preferences. Our findings identify the important substrates of SMARCAL1 in fork repair, suggest that RecG and SMARCAL1 are functional orthologs, and provide a comprehensive model of fork repair by these DNA translocases. [Display omitted] •RPA directs SMARCAL1 to regress stalled replication forks with a leading-strand gap•RPA directs SMARCAL1 to restore normal forks with a lagging-strand gap•Single-molecule analyses indicate RPA controls bursts of SMARCAL1 translocation•E. coli RecG, but not ZRANB3, works similarly to SMARCAL1 DNA translocases including SMARCAL1 catalyze remodeling of stalled replication forks to promote repair. Manosas, Cortez, and colleagues demonstrate that the single-stranded binding protein RPA directs SMARCAL1 to both promote fork regression and restoration. RPA dictates substrate preference such that the net effect is reversal of a stalled fork with a leading-strand gap and restoration of a normal fork with a lagging-strand gap. The E. coli RecG fork repair enzyme shares this substrate preference suggesting similarity in function.