Saw1 localizes to repair sites but is not required for recruitment of Rad10 to repair intermediates bearing short non-homologous 3′ flaps during single-strand annealing in S. cerevisiae
| Autor: | Eric P. Kelson, Paula L. Fischhaber, Aaron D. Miller, Melina Mardirosian, Claire Phan, Linette Nalbandyan |
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| Rok vydání: | 2015 |
| Předmět: |
0301 basic medicine
medicine.medical_specialty Saccharomyces cerevisiae Proteins DNA Repair DNA repair Clinical Biochemistry Saccharomyces cerevisiae DNA-binding protein Article 03 medical and health sciences chemistry.chemical_compound medicine Homologous chromosome DNA Breaks Double-Stranded Molecular Biology Nuclease biology Single-Strand Specific DNA and RNA Endonucleases fungi Cell Biology General Medicine Cell cycle biology.organism_classification eye diseases Double Strand Break Repair Surgery Cell biology DNA-Binding Proteins 030104 developmental biology chemistry biology.protein DNA |
| Zdroj: | Molecular and Cellular Biochemistry. 412:131-139 |
| ISSN: | 1573-4919 0300-8177 |
| DOI: | 10.1007/s11010-015-2616-7 |
| Popis: | SAW1 is required for efficient removal by the Rad1-Rad10 nuclease of 3′ non-homologous DNA ends (flaps) formed as intermediates during two modes of double-strand break repair in S. cerevisiae, single-strand annealing (SSA) and synthesis-dependent strand annealing (SDSA). Saw1 was shown in vitro to bind flaps with high affinity, but displayed diminished affinity when flaps were short (< 30 deoxynucleotides [nt]), consistent with it not being required for short flap cleavage. Accordingly, this study, using in vivo fluorescence microscopy showed that SAW1 was not required for recruitment of Rad10-YFP to DNA double-strand breaks (DSBs) during their repair by SSA when the flaps were ~10 nt. In contrast, recruitment of Rad10-YFP to DSBs when flaps were ~500 nt did require SAW1 in G1 phase of cell cycle. Interestingly, we observed a substantial increase in colocalization of Saw1-CFP and Rad10-YFP at DSBs when short flaps were formed during repair, especially in G1, indicating significant recruitment of Saw1 despite there being no requirement for Saw1 to recruit Rad10. Saw1-CFP was seldom observed at DSBs without Rad10-YFP. Together these results support a model in which Saw1, and Rad1-Rad10 are recruited as a complex to short and long flaps in all phases of cell cycle, but that Saw1 is only required for recruitment of Rad1-Rad10 to DSBs when long flaps are formed in G1. |
| Databáze: | OpenAIRE |
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