Fig. 4: Transcription-coupled repair of DPCs by CSA and CSB.
From: Transcription-coupled DNA–protein crosslink repair by CSB and CRL4CSA-mediated degradation
a, DPC-seq reads from MRC-5 WT, CSA and CSB KO cells in the CRIM1 gene directly (0 h) and 4 h after a 30 min pulse of 1 mM FA as described in Fig. 2b. b, Violin plots of per cent of DPC repair in expressed bins (>3 TPM) in MRC-5 WT, CSA and CSB KO cells as described in Fig. 2c. Unpaired two-tailed t-test. c, Relative colony survival of MRC-5 WT and indicated TC-NER KO cells treated with the indicated doses of FA (1 h pulse), UV, 5-Aza-dC (continuous) or cisplatin (1 day pulse). Graphs represent the mean ± s.e.m. from n = 7 (FA) or n = 3 (UV, 5-Aza-dC and cisplatin) independent experiments. Graphs were normalized to the untreated colony number, which was set at 100%. Unpaired two-tailed t-test with values from 2.5 mM FA, 6 J m–2 UV, 400 nM 5-Aza-dC and 3 μM cisplatin. d, Relative colony survival assay of HCT116 WT, CSB mutants and TC-NER KO cell lines treated with 1 h of FA pulse (left) or UV (right). The endogenous KI of CSB(K538R) disrupts ATPase activity, whereas the CSB-GG mutant (L1427G and L1428G) has a deficient UBD. Graphs were normalized to the untreated colony number, which was set at 100%. Graphs represent the mean ± s.e.m. n = 3 independent experiments. Unpaired two-tailed t-test using values from 1 mM FA and 3 J m–2 UV. e, Top: scheme of procedure. Bottom: FA (left) and ultraviolet-B irradiation (right) survival assays of WT, csa-1, csb-1, uvs-1 and xpa-1 mutant C. elegans. Shown is the mean ± s.e.m. of three independent experiments. Unpaired two-tailed t-test using values from 12 mM FA and 80 J m–2 UV. f, Top: scheme of procedure. Bottom: developmental growth of WT, csa-1, csb-1, uvs-1 and xpa-1 mutant C. elegans after control RNA interference (RNAi) or depletion of alh-1 or adh-5. For each condition, the developmental stage was counted as ‘adult, L4’, ‘L3, L2, L1’ or ‘egg’. Shown is the average ± s.e.m. of three independent experiments. Unpaired two-tailed t-test using values from ‘adult, L4’. Source numerical data are available in the source data. Schemes in e and f were created using BioRender (https://biorender.com).
