Phosphorylated CtIP Functions as a Co-factor of the MRE11-RAD50-NBS1 Endonuclease in DNA End Resection

• Published in: Molecular cell 2016-12, Vol.64 (5), p.940-950
• Main Authors: Anand, Roopesh, Ranjha, Lepakshi, Cannavo, Elda, Cejka, Petr
• Format: Article
• Language: English
• Subjects: Acid Anhydride Hydrolases; Carrier Proteins; Cell Cycle Proteins - genetics; Cell Cycle Proteins - metabolism; DNA Breaks, Double-Stranded; DNA end resection; DNA End-Joining Repair - genetics; DNA Helicases - genetics; DNA Helicases - metabolism; DNA Repair Enzymes; DNA-Binding Proteins; double-strand DNA break; Endodeoxyribonucleases; helicase; homologous recombination; Homologous Recombination - genetics; Humans; MRE11 Homologue Protein; Multiprotein Complexes - genetics; Nuclear Proteins; nuclease; Phosphorylation; Recombinant Proteins - genetics; Recombinant Proteins - metabolism
• ISSN: 1097-2765; 1097-4164
• DOI: 10.1016/j.molcel.2016.10.017

To repair a DNA double-strand break (DSB) by homologous recombination (HR), the 5′-terminated strand of the DSB must be resected. The human MRE11-RAD50-NBS1 (MRN) and CtIP proteins were implicated in the initiation of DNA end resection, but the underlying mechanism remained undefined. Here, we show that CtIP is a co-factor of the MRE11 endonuclease activity within the MRN complex. This function is absolutely dependent on CtIP phosphorylation that includes the key cyclin-dependent kinase target motif at Thr-847. Unlike in yeast, where the Xrs2/NBS1 subunit is dispensable in vitro, NBS1 is absolutely required in the human system. The MRE11 endonuclease in conjunction with RAD50, NBS1, and phosphorylated CtIP preferentially cleaves 5′-terminated DNA strands near DSBs. Our results define the initial step of HR that is particularly relevant for the processing of DSBs bearing protein blocks. [Display omitted] •Phosphorylated CtIP promotes the endonuclease of MRE11-RAD50-NBS1•The cleavage is dependent on the nuclease of MRE11 and the ATPase of RAD50•NBS1 and CtIP have structural roles to promote DNA cleavage by MRE11-RAD50•The endonuclease preferentially cleaves 5′ DNA strands near protein adducts Anand et al. demonstrate that phosphorylated CtIP stimulates the MRE11 endonuclease within the MRE11-RAD50-NBS1 complex. This activity initiates DNA end resection of broken DNA. This reconstitutes the first steps in DNA double-strand break repair by homologous recombination.