PrimPol primase mediates replication traverse of DNA interstrand crosslinks

Abstract

Phone +34 91 732 8000 ext 3490 González-Acosta et al., 2020 2 ABSTRACT Interstrand crosslinks (ICLs) are DNA lesions frequently induced by chemotherapy that interfere with essential processes such as replication and transcription. ICL repair may be initiated by the convergence of two replication forks at the crosslink, which results in a termination-like DNA structure recognized and processed by the Fanconi Anemia (FA)pathway. An alternative possibility to generate a suitable substrate for ICL repair involves "ICL traverse", a DNA damage tolerance mechanism in which a single fork arriving at the ICL can skip the lesion and restart DNA synthesis from a downstream point. This reaction requires FANCM translocase, the BLM/TOP3A/RMI1-2 (BTR) complex and other factors.Here we report that PrimPol, the second primase-polymerase identified in mammalian cells after Pola/Primase, interacts with BTR and participates in the ICL traverse reaction.A functional complementation assay reveals that the primase activity of PrimPol is required, confirming the need for re-priming events during ICL traverse. Genetic ablation of PRIMPOL strongly impaired this tolerance mechanism, making cells more dependent on fork convergence to initiate ICL repair. PRIMPOL KO cells and mice display hypersensitivity to ICL-inducing drugs, opening the possibility of targeting PrimPol activity to enhance the efficacy of chemotherapy based on DNA crosslinking agents.

Mechanisms for Maintaining Eukaryotic Replisome Progression in the Presence of DNA Damage

“…In particular, these studies have revealed that DDT mechanisms function not only to relieve polymerase stalling, but also to maintain CMG translocation in the presence of obstacles previously thought to pose a complete block to template unwinding. Here, the combined action of transient opening of the MCM ring, unwinding of the parental duplex by accessory helicases, and recoupling of the nascent leading strand by classic DDT mechanisms has expanded the range of obstacles considered able to be tolerated by the replisome ( Sparks et al, 2019 ; Wasserman et al, 2019 ; González-Acosta et al, 2020 ; Sato et al, 2020 , preprint). Meanwhile, the chronology of pathway choice for recoupling leading strand replication is beginning to be deciphered.…”

“…This suggests that PrimPol may function to rescue uncoupled forks when Pol δ alone or TLS cannot readily bypass a leading-strand obstacle ( Figure 5A ). In support of this, PrimPol is required for the tolerance of impediments not efficiently bypassed by TLS, including chain-terminating nucleosides ( Kobayashi et al, 2016 ), DNA secondary structures ( Schiavone et al, 2016 ), R-loops ( Šviković et al, 2019 ), cisplatin-induced adducts and hydroxyurea ( Quinet et al, 2020 ), bulky DNA adducts that induce recombination ( Piberger et al, 2020 ), and interstrand crosslinks (ICLs) ( González-Acosta et al, 2020 , Preprint). Indeed, although Rev1 is involved in bypass of G4-quadruplexes (G4s)—DNA secondary structures formed by Hoogsteen base-pairing between guanines—containing long loops ( Schiavone et al, 2014 ), PrimPol is required for tolerance of G4s that are more thermodynamically stable ( Schiavone et al, 2016 ).…”

“…BLM might therefore play an analogous role to RTEL1 during DPC bypass by unwinding the parental strands downstream of the ICL ( Ling et al, 2016 ). BTR also interacts with both RPA ( Wu et al, 2018 ) and PrimPol ( González-Acosta et al, 2020 , preprint). Recent work demonstrated that PrimPol participates in ICL bypass, likely by reinitiating leading-strand synthesis to restore rapid fork rates ( González-Acosta et al, 2020 , preprint).…”

“…BTR also interacts with both RPA ( Wu et al, 2018 ) and PrimPol ( González-Acosta et al, 2020 , preprint). Recent work demonstrated that PrimPol participates in ICL bypass, likely by reinitiating leading-strand synthesis to restore rapid fork rates ( González-Acosta et al, 2020 , preprint). Traversal would leave the X-shaped ICL structure behind the re-established replication fork to be repaired by the Fanconi anaemia pathway in a postreplicative manner ( Lopez-Martinez et al, 2016 ).…”

Identification and Validation of a DNA Damage Repair-Related Signature for Diffuse Large B-Cell Lymphoma

“…PRIMPOL is thought to contribute to cellular tolerance to DNA damage by facilitating damage bypass [67]. Further studies have shown that loss of PRIMPOL leads to increased cellular sensitivity to DNA cross-linking agents such as 17 BioMed Research International mitomycin C, suggesting that repriming of PRIMPOL may represent a novel target for improving the sensitivity of cancer cells to DNA-damaging chemotherapies [68,69]. Rad23 homolog B plays a critical role in nucleotide excision repair (NER) in humans [70].…”