Abstract
Genome stability involves accurate replication and DNA repair. Broken replication forks, such as those encountering a nick, lead to double strand breaks (DSBs), which are preferentially repaired by sister-chromatid recombination (SCR). To decipher the role of chromatin in eukaryotic DSB repair, here we analyze a collection of yeast chromatin-modifying mutants using a previously developed system for the molecular analysis of repair of replication-born DSBs by SCR based on a mini-HO site. We confirm the candidates through FLP-based systems based on a mutated version of the FLP flipase that causes nicks on either the leading or lagging DNA strands. We demonstrate that Rpd3L and Hda1 histone deacetylase (HDAC) complexes contribute to the repair of replication-born DSBs by facilitating cohesin loading, with no effect on other types of homology-dependent repair, thus preventing genome instability. We conclude that histone deacetylation favors general sister chromatid cohesion as a necessary step in SCR.
Highlights
Genome stability involves accurate replication and DNA repair
We previously developed a yeast TINV-HO plasmid system based on a 24-bp mini-HO site (HOr) to induce single-strand DNA (ssDNA) breaks that resulted in double strand breaks (DSBs) after replication and permitted the molecular analysis of their repair by SCR6,10
The TINV-HO system is based on two leu[2] inverted repeats, one of which contains the HOr site, which is inefficiently targeted by the endonuclease HO leading mainly to DNA nicks that are converted into DSBs by the replication fork (Fig. 1a)[6,10]
Summary
Genome stability involves accurate replication and DNA repair. Broken replication forks, such as those encountering a nick, lead to double strand breaks (DSBs), which are preferentially repaired by sister-chromatid recombination (SCR). We demonstrate that Rpd3L and Hda[1] histone deacetylase (HDAC) complexes contribute to the repair of replication-born DSBs by facilitating cohesin loading, with no effect on other types of homology-dependent repair, preventing genome instability. The DNA damage response (DDR) promotes the establishment of such damage-induced cohesion at the break site as well as at other genomic regions independently of replication[15,16,17]. Damage-induced cohesion promotes SCR and is favored by certain factors such as the Smc5/6 complex[18,19], the constitutive methylation of H3K79 (ref. 20), and by chromatin remodeling by the RSC complex[21]
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