title: RNF4 and PLK1 are required for replication fork collapse in ATR-deficient cells.
abstract: The ATR-CHK1 axis stabilizes stalled replication forks and prevents their collapse into DNA double-strand breaks (DSBs). Here, we show that fork collapse in Atr-deleted cells is mediated through the combined effects the sumo targeted E3-ubiquitin ligase RNF4 and activation of the AURKA-PLK1 pathway. As indicated previously, Atr-deleted cells exhibited a decreased ability to restart DNA replication following fork stalling in comparison with control cells. However, suppression of RNF4, AURKA, or PLK1 returned the reinitiation of replication in Atr-deleted cells to near wild-type levels. In RNF4-depleted cells, this rescue directly correlated with the persistence of sumoylation of chromatin-bound factors. Notably, RNF4 repression substantially suppressed the accumulation of DSBs in ATR-deficient cells, and this decrease in breaks was enhanced by concomitant inhibition of PLK1. DSBs resulting from ATR inhibition were also observed to be dependent on the endonuclease scaffold protein SLX4, suggesting that RNF4 and PLK1 either help activate the SLX4 complex or make DNA replication fork structures accessible for subsequent SLX4-dependent cleavage. Thus, replication fork collapse following ATR inhibition is a multistep process that disrupts replisome function and permits cleavage of the replication fork.
authors.affiliation: Abramson Family Cancer Research Institute, Department of Cancer Biology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania 19104, USA;
authors.forename: Ryan L
authors.initials: RL
authors.lastname: Ragland
authors.forename: Sima
authors.initials: S
authors.lastname: Patel
authors.forename: Rebecca S
authors.initials: RS
authors.lastname: Rivard
authors.forename: Kevin
authors.initials: K
authors.lastname: Smith
authors.forename: Ashley A
authors.initials: AA
authors.lastname: Peters
authors.forename: Anja-Katrin
authors.initials: AK
authors.lastname: Bielinsky
authors.forename: Eric J
authors.initials: EJ
authors.lastname: Brown
mesh_terms: D016475:3T3 Cells; D000818:Animals; D064007:Ataxia Telangiectasia Mutated Proteins / Q000235:genetics; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism*; D002843:Chromatin / Q000378:metabolism; D053903:DNA Breaks, Double-Stranded; D004261:DNA Replication; D051379:Mice; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism*; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000378:metabolism*; D045522:Recombinases / Q000378:metabolism; D058207:Sumoylation; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D044767:Ubiquitin-Protein Ligases; D000097869:Polo-Like Kinase 1
chemical_list: D064007:Ataxia Telangiectasia Mutated Proteins; D018797:Cell Cycle Proteins; D002843:Chromatin; D009687:Nuclear Proteins; D017346:Protein Serine-Threonine Kinases; D011518:Proto-Oncogene Proteins; D045522:Recombinases; D014157:Transcription Factors; D044767:Ubiquitin-Protein Ligases; D000097869:Polo-Like Kinase 1; C486410:Rnf4 protein, mouse; C576897:Atr protein, mouse; C543807:Slx4 protein, mouse
keywords: ATR; PLK1; RNF4; SLX4; SUMO; replication fork collapse