Abstract
The DNA damage checkpoint is activated in response to DNA double-strand breaks (DSBs). We had previously shown that chromatin assembly mediated by the histone chaperone Asf1 triggers inactivation of the DNA damage checkpoint in yeast after DSB repair, also called checkpoint recovery. Here we show that chromatin assembly factor 1 (CAF-1) also contributes to chromatin reassembly after DSB repair, explaining its role in checkpoint recovery. Towards understanding how chromatin assembly promotes checkpoint recovery, we find persistent presence of the damage sensors Ddc1 and Ddc2 after DSB repair in asf1 mutants. The genes encoding the E3 ubiquitin ligase complex Rtt101Mms1 are epistatic to ASF1 for survival following induction of a DSB, and Rtt101Mms1 are required for checkpoint recovery after DSB repair but not for chromatin assembly. By contrast, the Mms22 substrate adaptor that is degraded by Rtt101Mms1 is required for DSB repair per se. Deletion of MMS22 blocks loading of Rad51 at the DSB, while deletion of ASF1 or RTT101 leads to persistent Rad51 loading. We propose that checkpoint recovery is promoted by Rtt101Mms1-mediated ubiquitylation of Mms22 in order to halt Mms22-dependent loading of Rad51 onto double-stranded DNA after DSB repair, in concert with the chromatin assembly-mediated displacement of Rad51 and checkpoint sensors from the site of repair.
Highlights
DNA double-strand breaks (DSBs) occur often, arising on average ten times per cell per day [1]
We show that chromatin assembly factor 1 (CAF-1) contributes to the assembly of chromatin after DSB repair
We had previously shown that yeast lacking the H3-H4 histone chaperone CAF-1 are sensitive to DNA double-strand damaging agents even though they have no defect in repair of DSBs per se [33]
Summary
DNA double-strand breaks (DSBs) occur often, arising on average ten times per cell per day [1]. While DSBs are common, they are the most deleterious of genotoxic lesions, as they can result in translocations if misrepaired and loss of chromosomal segments if unrepaired. The cell has developed multiple pathways to try to ensure the accurate repair of DSBs and maintain genomic integrity. DSB repair pathways fall into two main classes.
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