Distinct roles for RSC and SWI/SNF chromatin remodelers in genomic excision repair

Abstract

Nucleosomes are a significant barrier to the repair of UV damage because they impede damage recognition by nucleotide excision repair (NER). The RSC and SWI/SNF chromatin remodelers function in cells to promote DNA access by moving or evicting nucleosomes, and both have been linked to NER in yeast. Here, we report genome-wide repair maps of UV-induced cyclobutane pyrimidine dimers (CPDs) in yeast cells lacking RSC or SWI/SNF activity. Our data indicate that SWI/SNF is not generally required for NER but instead promotes repair of CPD lesions at specific yeast genes. In contrast, mutation or depletion of RSC subunits causes a general defect in NER across the yeast genome. Our data indicate that RSC is required for repair not only in nucleosomal DNA but also in neighboring linker DNA and nucleosome-free regions (NFRs). Although depletion of the RSC catalytic subunit also affects base excision repair (BER) of N-methylpurine (NMP) lesions, RSC activity is less important for BER in linker DNA and NFRs. Furthermore, our data indicate that RSC plays a direct role in transcription-coupled NER (TC-NER) of transcribed DNA. These findings help to define the specific genomic and chromatin contexts in which each chromatin remodeler functions in DNA repair, and indicate that RSC plays a unique function in facilitating repair by both NER subpathways.

Set2 histone methyltransferase regulates transcription coupled-nucleotide excision repair in yeast

“…In WT cells, there were fewer unrepaired CPDs along the TS, beginning immediately after the TSS, while more unrepaired CPDs were present along the NTS, particularly at nucleosomes ( Fig 3A ). Slower repair of CPD lesions in nucleosomal DNA relative to adjacent linker DNA ( Fig 3A ) is consistent with our previous findings [ 26 , 39 , 41 ]. Comparing the WT and set2 Δ mutant data revealed more unrepaired CPDs in the set2 Δ mutant throughout the TS ( Fig 3B ), consistent with the model that Set2 promotes efficient TC-NER.…”

“…To measure the impact of Set2 on TC-NER, we analyzed the log ratio of unrepaired CPDs on the TS relative to the NTS across ~5000 yeast genes. This method quantifies the magnitude of the repair asymmetry between the TS and NTS of yeast genes due to TC-NER, and therefore can be used to characterize changes in TC-NER efficiency, even if GG-NER is also altered in the mutant cells [ 41 ]. In WT cells, the log 2 TS/NTS ratio was much less than zero in the transcribed DNA (i.e., region between TSS and TES; see Fig 2C ), due to faster repair of the TS by TC-NER.…”

“…Bin analysis for CPD repair along the transcribed strand (TS) and non-transcribed strand (NTS) of ~5000 yeast genes was performed as previously described [ 39 ], using transcription start site (TSS) and polyadenylation site (PAS, also referred to as transcription termination site (TTS) or transcription end site (TES)) coordinates from [ 39 ]. A similar gene bin analysis was used to display the fraction of CPDs remaining for yeast genes using the Java Treeview program [ 75 , 76 ], as previously described [ 41 ]. Genes were sorted by transcription rate [ 42 ] or by whether the gene is associated with a polyA SRAT.…”

“…Genes were sorted by transcription rate [ 42 ] or by whether the gene is associated with a polyA SRAT. Transcriptional asymmetry analysis calculated the log 2 ratio of the fraction of unrepaired CPDs in the TS relative to the NTS for each bin, as previously described [ 41 ]. Single nucleotide resolution repair analysis adjacent to the TSS was performed for the ~5200 yeast genes with published TSS coordinates [ 37 ], as previously described [ 39 , 54 ].…”

Nucleotide excision repair leaves a mark on chromatin: DNA damage detection in nucleosomes

“…Loss of SWI/SNF subunits was also found to confer UV hypersensitivity in yeast and C. elegans , suggestive of functional evolutionary conservation [ 95 , 96 ]. Although the reported interaction of two subunits with Rad4 in yeast may point to a more direct role in GG-NER in this species, mapping of genome-wide repair in yeast lacking SWI/SNF subunits shows that this complex is only required for GG-NER in a small subset of genes [ 97 ]. Instead, the related RSC ATP-dependent remodeling complex was found to promote GG-NER in both nucleosomal and non-nucleosomal DNA throughout the yeast genome.…”