Restoration of Replication Fork Stability in BRCA1- and BRCA2-Deficient Cells by Inactivation of SNF2-Family Fork Remodelers

Abstract

SUMMARY To ensure the completion of DNA replication and maintenance of genome integrity, DNA repair factors protect stalled replication forks upon replication stress. Previous studies have identified a critical role for the tumor suppressors BRCA1 and BRCA2 in preventing the degradation of nascent DNA by the MRE11 nuclease after replication stress. Here we show that depletion of SMARCAL1, a SNF2-family DNA translocase that remodels stalled forks, restores replication fork stability and reduces the formation of replication stress-induced DNA breaks and chromosomal aberrations in BRCA1/2-deficient cells. In addition to SMARCAL1, other SNF2-family fork remodelers, including ZRANB3 and HLTF, cause nascent DNA degradation and genomic instability in BRCA1/2-deficient cells upon replication stress. Our observations indicate that nascent DNA degradation in BRCA1/2-deficient cells occurs as a consequence of MRE11-dependent nucleolytic processing of reversed forks generated by fork remodelers. These studies provide mechanistic insights into the processes that cause genome instability in BRCA1/2-deficient cells.

Replication Fork Reversal: Players and Guardians

“…In particular, Kolinjivadi et al (2017) propose that SMARCAL1 remodels forks with persistent ssDNA gaps at the fork junction into reversed fork structures and Taglialatela et al (2017) suggest that the RPA-binding activity of SMARCAL1 is required for its replication function, in agreement with previous biochemical studies (Betous et al, 2013). At the same time, Vujanovic et al (2017) show that ZRANB3 interacts with polyubiquitinated PCNA to promote fork remodeling (Figure 1A).…”

“…Indeed, SMARCAL1, ZRANB3, and HLTF recognize different types of fork structures in vitro , suggesting that cells might use different factors depending on the particular type of replication intermediate (Betous et al, 2013; Hishiki et al, 2015; Kile et al, 2015). Along the same line, the studies of Kolinjivadi et al (2017), Taglialatela et al (2017), and Vujanovic et al (2017) clearly show that SMARCAL1 or ZRANB3 depletion does not fully abrogate reversed fork formation, supporting the idea that fork reversal is not mediated by a single fork remodeler and that different structures might arise even when using the same type of replication challenge. Moreover, other DNA translocases, including RAD54 (Bugreev et al, 2011) and FANCM (Gari et al, 2008), can mediate fork reversal in vitro , suggesting that additional factors may contribute to this process.…”

“…Interestingly, Kolinjivadi et al (2017), Tagliatela et al (2017), and Vujanovic et al (2017) show that fork reversal is not impaired under conditions that lead to formation of unstable RAD51 nucleofilaments and/or inefficient loading of RAD51 on ssDNA (Figure 1C), such as in BRCA1/2-mutant cells or in cells expressing a dominant RAD51 mutant allele, RAD51-T131P, which was recently reported to destabilize RAD51 nucleofilaments (Wang et al, 2015). The molecular steps that allow replication forks to reverse under conditions of inefficient RAD51 loading remain to be defined.…”

“…However, the molecular determinants required to protect the integrity of regressed arms until forks are restarted are unknown. Here, we review recent articles that provide a fresh perspective on these important issues by defining a key function of two translocases of the SWI/SNF protein family, i.e., ZRANB3 and SMARCAL1, in reversed fork formation (Kolinjivadi et al, 2017; Taglialatela et al, 2017; Vujanovic et al, 2017) and a key function of the breast cancer susceptibility proteins BRCA1 and BRCA2 in reversed fork protection (Kolinjivadi et al, 2017; Lemacon et al, 2017; Mijic et al, 2017; Taglialatela et al, 2017). …”

“…Using an elegant combination of single-molecule DNA fiber and electron microscopy approaches, Kolinjivadi et al (2017), Taglialatela et al (2017) and Vujanovic et al (2017) demonstrate that the translocase activities of SMARCAL1 and ZRANB3 are required for replication fork reversal. In particular, Kolinjivadi et al (2017) propose that SMARCAL1 remodels forks with persistent ssDNA gaps at the fork junction into reversed fork structures and Taglialatela et al (2017) suggest that the RPA-binding activity of SMARCAL1 is required for its replication function, in agreement with previous biochemical studies (Betous et al, 2013).…”