MRE11-RAD50-NBS1 is a critical regulator of FANCD2 stability and function during DNA double-strand break repair.
Monoubiquitination of the Fanconi anaemia protein FANCD2 is a key event leading to repair of interstrand cross-links. It was reported earlier that FANCD2 co-localizes with NBS1. However, the functional connection between FANCD2 and MRE11 is poorly understood. In this study, we show that inhibition of MRE11, NBS1 or RAD50 leads to a destabilization of FANCD2. FANCD2 accumulated from mid-S to G2 phase within sites containing single-stranded DNA (ssDNA) intermediates, or at sites of DNA damage, such as those created by restriction endonucleases and laser irradiation. Purified FANCD2, a ring-like particle by electron microscopy, preferentially bound ssDNA over various DNA substrates. Inhibition of MRE11 nuclease activity by Mirin decreased the number of FANCD2 foci formed in vivo. We propose that FANCD2 binds to ssDNA arising from MRE11-processed DNA double-strand breaks. Our data establish MRN as a crucial regulator of FANCD2 stability and function in the DNA damage response.
Roques
Céline
C
Genome Stability Laboratory, Laval University Cancer Research Center, Hôtel-Dieu de Québec, Québec, Canada.
Coulombe
Yan
Y
Delannoy
Mathieu
M
Vignard
Julien
J
Grossi
Simona
S
Brodeur
Isabelle
I
Rodrigue
Amélie
A
Gautier
Jean
J
Stasiak
Alicja Z
AZ
Stasiak
Andrzej
A
Constantinou
Angelos
A
Masson
Jean-Yves
JY
D017766:Acid Anhydride Hydrolases; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism*; D053903:DNA Breaks, Double-Stranded; D004260:DNA Repair; D045643:DNA Repair Enzymes / Q000235:genetics / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D015536:Down-Regulation; D052236:Fanconi Anemia Complementation Group D2 Protein / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D000076228:MRE11 Homologue Protein; D008854:Microscopy, Electron; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D055550:Protein Stability; D034741:RNA, Small Interfering / Q000235:genetics
D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; C499905:FANCD2 protein, human; D052236:Fanconi Anemia Complementation Group D2 Protein; C121766:MRE11 protein, human; C112156:NBN protein, human; D009687:Nuclear Proteins; D034741:RNA, Small Interfering; D000076228:MRE11 Homologue Protein; D017766:Acid Anhydride Hydrolases; C112158:RAD50 protein, human; D045643:DNA Repair Enzymes