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. 2015 Jun:30:38-45.
doi: 10.1016/j.dnarep.2015.03.006. Epub 2015 Mar 25.

SWI/SNF recruitment to a DNA double-strand break by the NuA4 and Gcn5 histone acetyltransferases

Gwendolyn Bennett  1 , Craig L Peterson  2
Affiliations

SWI/SNF recruitment to a DNA double-strand break by the NuA4 and Gcn5 histone acetyltransferases

Gwendolyn Bennett et al. DNA Repair (Amst). 2015 Jun.

Abstract

The DNA damage response to double-strand breaks (DSBs) is critical for cellular viability. Recent work has shown that a host of chromatin regulators are recruited to a DSB, and that they are important for the DNA damage response. However, the functional relationships between different chromatin regulators at DSBs remain unclear. Here we describe a conserved functional interaction among the chromatin remodeling enzyme, SWI/SNF, the NuA4 and Gcn5 histone acetyltransferases, and phosphorylation of histone H2A.X (γH2AX). Specifically, we find that the NuA4 and Gcn5 enzymes are both required for the robust recruitment of SWI/SNF to a DSB, which in turn promotes the phosphorylation of H2A.X.

Keywords: Chromatin; Gcn5; Histone acetyltransferase; Homologous recombination; NuA4; SWI/SNF.

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Figures

Figure 1
Figure 1. Gcn5 promotes H2A.X phosphorylation and the recruitment of multiple chromatin regulators to a DSB
(a) Schematic of the donorless yeast strain harboring a galactose inducible HO endonuclease, which cuts at the MAT locus of chromosome III. The approximate location of regions amplified for ChIP analyses to the right (“+”) of the break are shown in red and labeled according to their distance from the DSB in kilobases (kb). (b, c) Wild-type (wt), and gcn5Δ strains were arrested in G2/M using nocodazole, and analyzed by ChIP for recruitment of the indicated chromatin modifying enzyme subunits to the DSB region at the specified time points after DSB induction. (d) Input DNA at the break site relative to an unbroken locus (ACT1) and normalized to pre-induction levels. (e) As in (b), γH2AX levels determined by ChIP at the indicated time points after break induction.
Figure 2
Figure 2. NuA4 promotes SWI/SNF recruitment to a DSB and H2A.X phosphorylation
(a) Schematic representing the yeast culture growth and treatments. (b) Western blot of samples from ESA1-AID containing yeast strains treated with either ethanol or 500μm NAA. untr. = untreated samples. (c,d) The ESA1-AID yeast strain treated with ethanol or NAA and analyzed by ChIP for the indicated chromatin modifying enzyme subunits after DSB induction. (e) γH2AX levels determined by ChIP at the indicated time points after break induction. (f) Input DNA at the break site relative to an unbroken locus (ACT1) and normalized to pre-induction levels.
Figure 3
Figure 3. NuA4 acetylation of chromatin specifically promotes SWI/SNF recruitment to a DSB
(a,b) ChIP analyses of the indicated chromatin modifying enzyme subunits after an induced break in isogenic wt and yng2Δ yeast strains. (c) γH2AX levels determined by ChIP at the indicated time points after break induction. (d) Input DNA in at the break site relative to an unbroken locus (ACT1) and normalized to pre-induction levels.
Figure 4
Figure 4. SWI/SNF’s bromodomain is key for DSB recruitment and promotes H2A.X phosphorylation
(a) Isogenic wt and swi2Δbr yeast strains were synchronized in G2/M and analyzed by ChIP for the indicated chromatin modifying enzyme subunits at (a) the GAL1/10 promoter region, and (b,c) the double-strand break site. (d) γH2AX levels determined by ChIP at the indicated time points after break induction. (e) Input DNA at the break site relative to an unbroken locus (ACT1) and normalized to pre-induction levels.
Figure 5
Figure 5. A model for the interaction of NuA4 acetylation, SWI/SNF DSB recruitment, and γH2AX during DSB damage response
See text for details
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References

    1. Papamichos-Chronakis M, Peterson CL. Chromatin and the genome integrity network. Nat. Rev. Genet. 2013;14:62–75. - PMC - PubMed
    1. Peterson CL, Almouzni G. Nucleosome dynamics as modular systems that integrate DNA damage and repair. Cold Spring Harb. Perspect. Biol. 2013;5 - PMC - PubMed
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