Abstract
The RSC chromatin remodeling complex has been implicated in contributing to DNA double-strand break (DSB) repair in a number of studies. Both survival and levels of H2A phosphorylation in response to damage are reduced in the absence of RSC. Importantly, there is evidence for two isoforms of this complex, defined by the presence of either Rsc1 or Rsc2. Here, we investigated whether the two isoforms of RSC provide distinct contributions to DNA damage responses. First, we established that the two isoforms of RSC differ in the presence of Rsc1 or Rsc2 but otherwise have the same subunit composition. We found that both rsc1 and rsc2 mutant strains have intact DNA damage-induced checkpoint activity and transcriptional induction. In addition, both strains show reduced non-homologous end joining activity and have a similar spectrum of DSB repair junctions, suggesting perhaps that the two complexes provide the same functions. However, the hypersensitivity of a rsc1 strain cannot be complemented with an extra copy of RSC2, and likewise, the hypersensitivity of the rsc2 strain remains unchanged when an additional copy of RSC1 is present, indicating that the two proteins are unable to functionally compensate for one another in DNA damage responses. Rsc1, but not Rsc2, is required for nucleosome sliding flanking a DNA DSB. Interestingly, while swapping the domains from Rsc1 into the Rsc2 protein does not compromise hypersensitivity to DNA damage suggesting they are functionally interchangeable, the BAH domain from Rsc1 confers upon Rsc2 the ability to remodel chromatin at a DNA break. These data demonstrate that, despite the similarity between Rsc1 and Rsc2, the two different isoforms of RSC provide distinct functions in DNA damage responses, and that at least part of the functional specificity is dictated by the BAH domains.
Highlights
Access to chromatin can be regulated by covalent post-translational modification of histone proteins and by the action of ATP-dependent remodeling complexes
An additional copy of RSC1 in a rsc2 mutant strain does not increase the DNA damage resistance of that strain (Fig. 4A). These plasmids were able to rescue DNA damage hypersensitivity when Rsc1 was expressed in a rsc1 mutant strain and when Rsc2 was expressed in a rsc2 mutant strain (Figure 4B). While both proteins contribute to non-homologous end joining (NHEJ) and have similar phenotypes, these data suggest that Rsc1 and Rsc2 provide distinct functions in mediating survival after DNA damage and cannot entirely compensate for one another
We show that Rsc1 and Rsc2 exist in two separate RSC complexes that appear to be otherwise identical in subunit composition
Summary
Access to chromatin can be regulated by covalent post-translational modification of histone proteins and by the action of ATP-dependent remodeling complexes. ATP-dependent remodelers are large multisubunit complexes that couple ATP hydrolysis to movement of histones or nucleosomes. A number of different remodeling activities can be performed by these complexes, including exchange or incorporation of core histones or histone variants, eviction of histones or nucleosomes and repositioning or sliding of nucleosomes [1]. Chromatin remodeling activities are known to facilitate the repair of DNA double-strand breaks (DSBs). One such activity is the RSC (Remodels the Structure of Chromatin) complex originally identified by Cairns et al [2]. The catalytic activity is provided by Sth, which is encoded by an essential gene, and there are currently 17 known subunits (Sth, Rsc, Rsc, Rsc, Rsc, Rsc, Rsc7/Npl, Rsc, Rsc, Sfh, Arp, Arp, Rsc, Htl, Rtt102, Rsc and Ldb7)
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