Journal Article

ATR and ATM differently regulate WRN to prevent DSBs at stalled replication forks and promote replication fork recovery

Franchitto, Annapaola, Pirzio, Livia Maria, Ammazzalorso, Francesca, Pichierri, Pietro, Bignami, Margherita

2010

Overview

Accurate response to replication arrest is crucial to preserve genome stability and requires both the ATR and ATM functions. The Werner syndrome protein (WRN) is implicated in the recovery of stalled replication forks, and although an ATR/ATM‐dependent phosphorylation of WRN was observed after replication arrest, the function of such modifications during the response to perturbed replication is not yet appreciated. Here, we report that WRN is directly phosphorylated by ATR at multiple C‐terminal S/TQ residues. Suppression of ATR‐mediated phosphorylation of WRN prevents proper accumulation of WRN in nuclear foci, co‐localisation with RPA and causes breakage of stalled forks. On the other hand, inhibition of ATM kinase activity or expression of an ATM‐unphosphorylable WRN allele leads to retention of WRN in nuclear foci and impaired recruitment of RAD51 recombinase resulting in reduced viability after fork collapse. Altogether, our findings indicate that ATR and ATM promote recovery from perturbed replication by differently regulating WRN at defined moments of the response to replication fork arrest. By dissecting the so far loosely defined roles of the Werner syndrome helicase at stalled replication forks, this study shows that two highly related kinases can differentially regulate specific WRN phosphorylation sites and distinct WRN functions at various stages after replication arrest.

• Publisher: John Wiley & Sons, Ltd,Nature Publishing Group UK,Springer Nature B.V,Nature Publishing Group
• Subject: Ataxia Telangiectasia Mutated Proteins / Blotting, Western / Cell Cycle / Cell Cycle Proteins - antagonists & inhibitors / Cell Cycle Proteins - genetics / Cell Cycle Proteins - metabolism / Cell division / Cell Nucleus - metabolism / DNA Breaks, Double-Stranded / DNA Repair / DNA Replication / DNA-Binding Proteins - genetics / DNA-Binding Proteins - metabolism / EMBO13 / Exodeoxyribonucleases - genetics / Exodeoxyribonucleases - metabolism / Flow Cytometry / Fluorescent Antibody Technique / Gene expression / genome instability / Genomics / Humans / Immunoprecipitation / Molecular biology / Phosphorylation / Protein Serine-Threonine Kinases - antagonists & inhibitors / Protein Serine-Threonine Kinases - genetics / Protein Serine-Threonine Kinases - metabolism / Proteins / Rad51 Recombinase - antagonists & inhibitors / Rad51 Recombinase - genetics / Rad51 Recombinase - metabolism / RecQ helicases / RecQ Helicases - genetics / RecQ Helicases - metabolism / replication fork arrest / Retention / Reverse Transcriptase Polymerase Chain Reaction / RNA, Messenger - genetics / RNA, Small Interfering - pharmacology / Tumor Suppressor Proteins - genetics / Tumor Suppressor Proteins - metabolism / Werner syndrome / Werner Syndrome Helicase