Sae2 promotes DNA damage resistance by removing the Mre11–Rad50–Xrs2 complex from DNA and attenuating Rad53 signaling

• Published in: Proceedings of the National Academy of Sciences - PNAS Vol. 112; no. 15; pp. E1880 - E1887
• Main Authors: Chen, Huan, Donnianni, Roberto A., Handa, Naofumi, Deng, Sarah K., Oh, Julyun, Timashev, Leonid A., Kowalczykowski, Stephen C., Symington, Lorraine S.
• Format: Journal Article
• Language: English
• Published: United States National Academy of Sciences 14-04-2015; National Acad Sciences
• Series: PNAS Plus
• Subjects: Biological Sciences; Biosensors; Cell Cycle - genetics; Cell Cycle Proteins - genetics; Cell Cycle Proteins - metabolism; Cells; Checkpoint Kinase 2 - genetics; Checkpoint Kinase 2 - metabolism; DNA Breaks, Double-Stranded; DNA Damage; DNA Repair; DNA, Fungal - genetics; DNA, Fungal - metabolism; DNA-Binding Proteins - genetics; DNA-Binding Proteins - metabolism; Endodeoxyribonucleases - genetics; Endodeoxyribonucleases - metabolism; Endonucleases - genetics; Endonucleases - metabolism; Enzymes; Exodeoxyribonucleases - genetics; Exodeoxyribonucleases - metabolism; Kinases; Microscopy, Fluorescence; Multiprotein Complexes - genetics; Multiprotein Complexes - metabolism; Mutation; PNAS Plus; Protein Binding; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins - genetics; Saccharomyces cerevisiae Proteins - metabolism; Signal Transduction - genetics; DNA damage checkpoint; Mre11; Sae2; DNA repair
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.1503331112

Significance Chromosomal double-strand breaks (DSBs) are cytotoxic forms of DNA damage that must be accurately repaired to maintain genome integrity. The conserved Mre11–Rad50–Xrs2/NBS1 nuclease/ATPase complex plays an important role in repair by functioning as a damage sensor and by regulation of DNA end processing to ensure repair by the most appropriate mechanism. Yeast Sae2 is known to function with Mre11 to process DNA ends, but its precise role is poorly understood. Here we show that it is the failure to remove Mre11 from DNA ends, leading to persistent DNA damage signaling and cell cycle arrest, that causes sensitivity of Sae2-deficient cells to DNA damaging agents.
The Mre11–Rad50–Xrs2/NBS1 (MRX/N) nuclease/ATPase complex plays structural and catalytic roles in the repair of DNA double-strand breaks (DSBs) and is the DNA damage sensor for Tel1/ATM kinase activation. Saccharomyces cerevisiae Sae2 can function with MRX to initiate 5′-3′ end resection and also plays an important role in attenuation of DNA damage signaling. Here we describe a class of mre11 alleles that suppresses the DNA damage sensitivity of sae2 Δ cells by accelerating turnover of Mre11 at DNA ends, shutting off the DNA damage checkpoint and allowing cell cycle progression. The mre11 alleles do not suppress the end resection or hairpin-opening defects of the sae2 Δ mutant, indicating that these functions of Sae2 are not responsible for DNA damage resistance. The purified M ᴾ¹¹⁰ᴸRX complex shows reduced binding to single- and double-stranded DNA in vitro relative to wild-type MRX, consistent with the increased turnover of Mre11 from damaged sites in vivo. Furthermore, overproduction of Mre11 causes DNA damage sensitivity only in the absence of Sae2. Together, these data suggest that it is the failure to remove Mre11 from DNA ends and attenuate Rad53 kinase signaling that causes hypersensitivity of sae2 Δ cells to clastogens.