Human ISWI complexes are targeted by SMARCA5 ATPase and SLIDE domains to help resolve lesion-stalled transcription.
Bottom Line: Chromatin compaction of deoxyribonucleic acid (DNA) presents a major challenge to the detection and removal of DNA damage.Using live cell imaging, we identify a novel function for two distinct mammalian ISWI adenosine triphosphate (ATP)-dependent chromatin remodeling complexes in resolving lesion-stalled transcription.Our studies support a model in which SMARCA5 targeting to DNA damage-stalled transcription sites is controlled by an ATP-hydrolysis-dependent scanning and proofreading mechanism, highlighting how SWI2/SNF2 chromatin remodelers identify and bind nucleosomes containing damaged DNA.
Affiliation: Department of Genetics, Medical Genetics Cluster, Cancer Genomics Netherlands, Erasmus MC, Rotterdam, 3015 GE, The Netherlands.Show MeSH
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Mentions: TC-NER factors such as CSB localize to DNA damage induced by a 266-nm UV-C laser, which specifically induces CPD and 6-4PP photolesions (Figure 2A and Supplementary Movie S1) (35,41). Stably expressed GFP-tagged SMARCA5 also rapidly accumulated at local UV-C damage, in both U2OS and MRC5 cells (Figure 2A, Supplementary Figure S2A and Supplementary Movie S2), in a dose-dependent manner (Supplementary Figure S2B). This was confirmed by local UV damage induction using a microporous filter (Supplementary Figure S2C) (43). The association of TC-NER factors with TC-NER complexes depends on stalling of RNApolII complexes and thus on active transcription (4,36). Inhibition of RNApolII activity using α-amanitin (44) indeed decreased the accumulation of both CSB and SMARCA5 at local damage (Figure 2B). SMARCA5 recruitment was also attenuated by the transcription elongation inhibitor DRB (Supplementary Figure S3A, in which both recruitment to the center and periphery of the damaged area are quantified as explained below) (45). These results confirm a function of SMARCA5 in TC-NER and suggest that this protein may localize to UV damage depending on RNApolII stalling.
Affiliation: Department of Genetics, Medical Genetics Cluster, Cancer Genomics Netherlands, Erasmus MC, Rotterdam, 3015 GE, The Netherlands.