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The 9-1-1 checkpoint clamp coordinates resection at DNA double strand breaks.

Ngo GH, Lydall D - Nucleic Acids Res. (2015)

Bottom Line: However, 9-1-1 also stimulates resection by Exo1- and Dna2-Sgs1-dependent nuclease/helicase activities, and this can be observed in the absence of Rad9(53BP1).Our experiments illustrate the central role of the 9-1-1 checkpoint sliding clamp in the DNA damage response network that coordinates the response to broken DNA ends.Our results have implications in all eukaryotic cells.

View Article: PubMed Central - PubMed

Affiliation: Institute for Cell and Molecular Biosciences (ICaMB), Medical School, Newcastle University, Newcastle upon Tyne, NE2 4HH, UK.

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The 9-1-1 complex stimulates both Exo1- and Dna2-Sgs1-dependent resection in rad9Δ cells. (A-C) Analysis of 3′ ssDNA accumulation in rad9Δ background strains at the indicated loci. The data plotted and the P-values are as described in Figure 1. (D) The 9-1-1 complex stimulates both Exo1 and Dna2-Sgs1 in rad9Δ cells.
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Figure 5: The 9-1-1 complex stimulates both Exo1- and Dna2-Sgs1-dependent resection in rad9Δ cells. (A-C) Analysis of 3′ ssDNA accumulation in rad9Δ background strains at the indicated loci. The data plotted and the P-values are as described in Figure 1. (D) The 9-1-1 complex stimulates both Exo1 and Dna2-Sgs1 in rad9Δ cells.

Mentions: The 9-1-1 complex promotes resection at uncapped telomeres in vivo, where Rad953BP1 binding is low (Figure 3B), and in vitro (in the absence of Rad953BP1) by stimulating both Dna2-Sgs1- and Exo1-dependent resection (24). To test whether 9-1-1 stimulates Dna2-Sgs1- and Exo1-dependent resection at DSBs in vivo, we deleted MEC3 in rad9Δ exo1Δ, rad9Δ sgs1Δ, rad9Δ dna2Δ or rad9Δ sgs1Δ exo1Δ genetic backgrounds and measured ssDNA accumulation (Figure 5A–C, Supplementary Figure S4). In agreement with the results in Figure 4A, mec3Δ decreased resection in all rad9Δ strains following the induction of DSBs (Figure 5A–C, compare rad9Δ and rad9Δ mec3Δ at SNT1). Consistent with the results in RAD9+ strains, deletion of EXO1 or SGS1/DNA2 reduced resection in rad9Δ strains and deletion of both SGS1 and EXO1 completed eliminated resection (Figure 5A–C, Supplementary Figure S4). We conclude that all resection in rad9Δ strains is dependent on Dna2-Sgs1 or Exo1. This suggests that other nucleases, such as MRX-Sae2, do not become hyperactive in the absence of Rad953BP1.


The 9-1-1 checkpoint clamp coordinates resection at DNA double strand breaks.

Ngo GH, Lydall D - Nucleic Acids Res. (2015)

The 9-1-1 complex stimulates both Exo1- and Dna2-Sgs1-dependent resection in rad9Δ cells. (A-C) Analysis of 3′ ssDNA accumulation in rad9Δ background strains at the indicated loci. The data plotted and the P-values are as described in Figure 1. (D) The 9-1-1 complex stimulates both Exo1 and Dna2-Sgs1 in rad9Δ cells.
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Related In: Results  -  Collection

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Figure 5: The 9-1-1 complex stimulates both Exo1- and Dna2-Sgs1-dependent resection in rad9Δ cells. (A-C) Analysis of 3′ ssDNA accumulation in rad9Δ background strains at the indicated loci. The data plotted and the P-values are as described in Figure 1. (D) The 9-1-1 complex stimulates both Exo1 and Dna2-Sgs1 in rad9Δ cells.
Mentions: The 9-1-1 complex promotes resection at uncapped telomeres in vivo, where Rad953BP1 binding is low (Figure 3B), and in vitro (in the absence of Rad953BP1) by stimulating both Dna2-Sgs1- and Exo1-dependent resection (24). To test whether 9-1-1 stimulates Dna2-Sgs1- and Exo1-dependent resection at DSBs in vivo, we deleted MEC3 in rad9Δ exo1Δ, rad9Δ sgs1Δ, rad9Δ dna2Δ or rad9Δ sgs1Δ exo1Δ genetic backgrounds and measured ssDNA accumulation (Figure 5A–C, Supplementary Figure S4). In agreement with the results in Figure 4A, mec3Δ decreased resection in all rad9Δ strains following the induction of DSBs (Figure 5A–C, compare rad9Δ and rad9Δ mec3Δ at SNT1). Consistent with the results in RAD9+ strains, deletion of EXO1 or SGS1/DNA2 reduced resection in rad9Δ strains and deletion of both SGS1 and EXO1 completed eliminated resection (Figure 5A–C, Supplementary Figure S4). We conclude that all resection in rad9Δ strains is dependent on Dna2-Sgs1 or Exo1. This suggests that other nucleases, such as MRX-Sae2, do not become hyperactive in the absence of Rad953BP1.

Bottom Line: However, 9-1-1 also stimulates resection by Exo1- and Dna2-Sgs1-dependent nuclease/helicase activities, and this can be observed in the absence of Rad9(53BP1).Our experiments illustrate the central role of the 9-1-1 checkpoint sliding clamp in the DNA damage response network that coordinates the response to broken DNA ends.Our results have implications in all eukaryotic cells.

View Article: PubMed Central - PubMed

Affiliation: Institute for Cell and Molecular Biosciences (ICaMB), Medical School, Newcastle University, Newcastle upon Tyne, NE2 4HH, UK.

Show MeSH
Related in: MedlinePlus