Werner syndrome protein participates in a complex with RAD51, RAD54, RAD54B and ATR in response to ICL-induced replication arrest.
Werner syndrome (WS) is a rare genetic disorder characterized by genomic instability caused by defects in the WRN gene encoding a member of the human RecQ helicase family. RecQ helicases are involved in several DNA metabolic pathways including homologous recombination (HR) processes during repair of stalled replication forks. Following introduction of interstrand DNA crosslinks (ICL), WRN relocated from nucleoli to arrested replication forks in the nucleoplasm where it interacted with the HR protein RAD52. In this study, we use fluorescence resonance energy transfer (FRET) and immune-precipitation experiments to demonstrate that WRN participates in a multiprotein complex including RAD51, RAD54, RAD54B and ATR in cells where replication has been arrested by ICL. We verify the WRN-RAD51 and WRN-RAD54B direct interaction in vitro. Our data support a role for WRN also in the recombination step of ICL repair.
Otterlei
Marit
M
Laboratory of Molecular Gerontology, National Institute on Aging, NIH, 5600 Nathan Shock Dr., Baltimore, MD 21224, USA. marit.otterlei@ntnu.no
Bruheim
Per
P
Ahn
Byungchan
B
Bussen
Wendy
W
Karmakar
Parimal
P
Baynton
Kathy
K
Bohr
Vilhelm A
VA
D064007:Ataxia Telangiectasia Mutated Proteins; D015153:Blotting, Western; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism*; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism; D002470:Cell Survival / Q000187:drug effects; D003432:Cross-Linking Reagents / Q000737:chemistry / Q000494:pharmacology*; D004265:DNA Helicases / Q000235:genetics / Q000378:metabolism*; D004261:DNA Replication / Q000187:drug effects; D004268:DNA-Binding Proteins; D004797:Enzyme-Linked Immunosorbent Assay; D005090:Exodeoxyribonucleases; D031541:Fluorescence Resonance Energy Transfer; D006367:HeLa Cells; D006801:Humans; D047468:Immunoprecipitation; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D018613:Microscopy, Confocal; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism*; D021381:Protein Transport / Q000187:drug effects; D051135:Rad51 Recombinase / Q000235:genetics / Q000378:metabolism*; D053484:RecQ Helicases / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D000071657:Werner Syndrome Helicase
D018797:Cell Cycle Proteins; D003432:Cross-Linking Reagents; D004268:DNA-Binding Proteins; D008164:Luminescent Proteins; D009687:Nuclear Proteins; C491221:RAD54B protein, human; D011993:Recombinant Fusion Proteins; C576597:ATR protein, human; D064007:Ataxia Telangiectasia Mutated Proteins; D017346:Protein Serine-Threonine Kinases; D051135:Rad51 Recombinase; D005090:Exodeoxyribonucleases; D004265:DNA Helicases; C103532:RAD54L protein, human; D053484:RecQ Helicases; C098785:WRN protein, human; D000071657:Werner Syndrome Helicase