title: DNA double-strand break repair pathway choice is directed by distinct MRE11 nuclease activities.
abstract: MRE11 within the MRE11-RAD50-NBS1 (MRN) complex acts in DNA double-strand break repair (DSBR), detection, and signaling; yet, how its endo- and exonuclease activities regulate DSBR by nonhomologous end-joining (NHEJ) versus homologous recombination (HR) remains enigmatic. Here, we employed structure-based design with a focused chemical library to discover specific MRE11 endo- or exonuclease inhibitors. With these inhibitors, we examined repair pathway choice at DSBs generated in G2 following radiation exposure. While nuclease inhibition impairs radiation-induced replication protein A (RPA) chromatin binding, suggesting diminished resection, the inhibitors surprisingly direct different repair outcomes. Endonuclease inhibition promotes NHEJ in lieu of HR, while exonuclease inhibition confers a repair defect. Collectively, the results describe nuclease-specific MRE11 inhibitors, define distinct nuclease roles in DSB repair, and support a mechanism whereby MRE11 endonuclease initiates resection, thereby licensing HR followed by MRE11 exonuclease and EXO1/BLM bidirectional resection toward and away from the DNA end, which commits to HR.
authors: Genome Damage and Stability Centre, University of Sussex, Brighton BN1 9RQ, UK; Advanced Scientific Research Leaders Development Unit, Gunma University, Maebashi, Gunma 371-8511, Japan.
authors: Atsushi
authors: A
authors: Shibata
authors: Skaggs Institute of Chemical Biology, The Scripps Research Institute, La Jolla, CA 92037, USA; The Life Sciences Division, Lawrence Berkeley National Laboratory, Berkeley, CA 94720, USA.
authors: Davide
authors: D
authors: Moiani
authors: Skaggs Institute of Chemical Biology, The Scripps Research Institute, La Jolla, CA 92037, USA; The Life Sciences Division, Lawrence Berkeley National Laboratory, Berkeley, CA 94720, USA.
authors: Andrew S
authors: AS
authors: Arvai
authors: Skaggs Institute of Chemical Biology, The Scripps Research Institute, La Jolla, CA 92037, USA; The Life Sciences Division, Lawrence Berkeley National Laboratory, Berkeley, CA 94720, USA; The School of Biotechnology, Amrita University, Kollam, Kerala 690525, India.
authors: Jefferson
authors: J
authors: Perry
authors: Departments of Radiation Oncology and Medical Biophysics, University of Toronto, ON M5G 2M9, Canada.
authors: Shane M
authors: SM
authors: Harding
authors: Genome Stability Laboratory, Laval University Cancer Research Center, Hôtel-Dieu de Québec, 9 McMahon, QC G1R 2J6, Canada.
authors: Marie-Michelle
authors: MM
authors: Genois
authors: Genome Stability Laboratory, Laval University Cancer Research Center, Hôtel-Dieu de Québec, 9 McMahon, QC G1R 2J6, Canada.
authors: Ranjan
authors: R
authors: Maity
authors: Department of Radiation Oncology, Department of Genetics, Erasmus University Medical Center, P.O. Box 2040, Rotterdam 3000 CA, the Netherlands.
authors: Sari
authors: S
authors: van Rossum-Fikkert
authors: Department of Radiation Oncology, Department of Genetics, Erasmus University Medical Center, P.O. Box 2040, Rotterdam 3000 CA, the Netherlands.
authors: Aryandi
authors: A
authors: Kertokalio
authors: Dipartimento Farmaco Chimico Tecnologico, Università degli Studi di Siena, Via A. Moro, 53100 Siena, Italy.
authors: Filippo
authors: F
authors: Romoli
authors: Genome Damage and Stability Centre, University of Sussex, Brighton BN1 9RQ, UK.
authors: Amani
authors: A
authors: Ismail
authors: Dipartimento Farmaco Chimico Tecnologico, Università degli Studi di Siena, Via A. Moro, 53100 Siena, Italy.
authors: Ermal
authors: E
authors: Ismalaj
authors: Dipartimento Farmaco Chimico Tecnologico, Università degli Studi di Siena, Via A. Moro, 53100 Siena, Italy.
authors: Elena
authors: E
authors: Petricci
authors: Genome Damage and Stability Centre, University of Sussex, Brighton BN1 9RQ, UK.
authors: Matthew J
authors: MJ
authors: Neale
authors: Departments of Radiation Oncology and Medical Biophysics, University of Toronto, ON M5G 2M9, Canada.
authors: Robert G
authors: RG
authors: Bristow
authors: Genome Stability Laboratory, Laval University Cancer Research Center, Hôtel-Dieu de Québec, 9 McMahon, QC G1R 2J6, Canada.
authors: Jean-Yves
authors: JY
authors: Masson
authors: Department of Radiation Oncology, Department of Genetics, Erasmus University Medical Center, P.O. Box 2040, Rotterdam 3000 CA, the Netherlands.
authors: Claire
authors: C
authors: Wyman
authors: Genome Damage and Stability Centre, University of Sussex, Brighton BN1 9RQ, UK. Electronic address: p.a.jeggo@sussex.ac.uk.
authors: Penny A
authors: PA
authors: Jeggo
authors: Skaggs Institute of Chemical Biology, The Scripps Research Institute, La Jolla, CA 92037, USA; The Life Sciences Division, Lawrence Berkeley National Laboratory, Berkeley, CA 94720, USA. Electronic address: jatainer@lbl.gov.
authors: John A
authors: JA
authors: Tainer
mesh_terms: D002460:Cell Line; D002843:Chromatin / Q000235:genetics / Q000378:metabolism; D053903:DNA Breaks, Double-Stranded; D059766:DNA End-Joining Repair; D045643:DNA Repair Enzymes / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000737:chemistry*; D005090:Exodeoxyribonucleases / Q000235:genetics / Q000378:metabolism; D016195:G2 Phase; D005720:Gamma Rays / Q000009:adverse effects; D006801:Humans; D000076228:MRE11 Homologue Protein; D059767:Recombinational DNA Repair; D051716:Replication Protein A / Q000235:genetics / Q000378:metabolism
chemical_list: D002843:Chromatin; D004268:DNA-Binding Proteins; D004791:Enzyme Inhibitors; C121766:MRE11 protein, human; D051716:Replication Protein A; C114331:EXO1 protein, human; D005090:Exodeoxyribonucleases; D000076228:MRE11 Homologue Protein; D045643:DNA Repair Enzymes