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UV-DDB, consisting of subunits DDB1 and DDB2, recognizes UV-induced photoproducts during global genome nucleotide excision repair (GG-NER). We recently demonstrated a noncanonical role of UV-DDB in stimulating base excision repair (BER) which raised several questions about the timing of UV-DDB arrival at 8-oxoguanine (8-oxoG), and the dependency of UV-DDB on the recruitment of downstream BER and NER proteins. Using two different approaches to introduce 8-oxoG in cells, we show that DDB2 is recruited to 8-oxoG immediately after damage and colocalizes with 8-oxoG glycosylase (OGG1) at sites of repair. 8-oxoG removal and OGG1 recruitment is significantly reduced in the absence of DDB2. NER proteins, XPA and XPC, also accumulate at 8-oxoG. While XPC recruitment is dependent on DDB2, XPA recruitment is DDB2-independent and transcription-coupled. Finally, DDB2 accumulation at 8-oxoG induces local chromatin unfolding. We propose that DDB2-mediated chromatin decompaction facilitates the recruitment of downstream BER proteins to 8-oxoG lesions. Nucleotide excision repair proteins are involved in the repair of UV-induced DNA damage. Here, the authors show that NER proteins, DDB2, XPC, and XPA play a vital role in the 8-oxoguanine repair by coordinating with base excision repair protein OGG1.

DNA damage sensors DDB2 and XPC initiate global genome nucleotide excision repair (NER) to protect DNA from mutagenesis caused by helix-distorting lesions. XPC recognizes helical distortions by binding to unpaired ssDNA opposite DNA lesions. DDB2 binds to UV-induced lesions directly and facilitates efficient recognition by XPC. We show that not only lesion-binding but also timely DDB2 dissociation is required for DNA damage handover to XPC and swift progression of the multistep repair reaction. DNA-binding-induced DDB2 ubiquitylation and ensuing degradation regulate its homeostasis to prevent excessive lesion (re)binding. Additionally, damage handover from DDB2 to XPC coincides with the arrival of the TFIIH complex, which further promotes DDB2 dissociation and formation of a stable XPC-TFIIH damage verification complex. Our results reveal a reciprocal coordination between DNA damage recognition and verification within NER and illustrate that timely repair factor dissociation is vital for correct spatiotemporal control of a multistep repair process. DNA damage sensors DDB2 and XPC are fundamental factors to initiate global genome nucleotide excision repair and protect DNA from mutagenesis. Here the authors reveal that ubiquitin and TFIIH-stimulated DDB2 dissociation promotes DNA damage handover to XPC in nucleotide excision repair.

The integrity of DNA is continuously challenged by metabolism-derived and environmental genotoxic agents that cause a variety of DNA lesions, including base alterations and breaks. DNA damage interferes with vital processes such as transcription and replication, and if not repaired properly, can ultimately lead to premature aging and cancer. Multiple DNA pathways signaling for DNA repair and DNA damage collectively safeguard the integrity of DNA. Chromatin plays a pivotal role in regulating DNA-associated processes, and is itself subject to regulation by the DNA-damage response. Chromatin influences access to DNA, and often serves as a docking or signaling site for repair and signaling proteins. Its structure can be adapted by post-translational histone modifications and nucleosome remodeling, catalyzed by the activity of ATP-dependent chromatin-remodeling complexes. In recent years, accumulating evidence has suggested that ATP-dependent chromatin-remodeling complexes play important, although poorly characterized, roles in facilitating the effectiveness of the DNA-damage response. In this review, we summarize the current knowledge on the involvement of ATP-dependent chromatin remodeling in three major DNA repair pathways: nucleotide excision repair, homologous recombination, and non-homologous end-joining. This shows that a surprisingly large number of different remodeling complexes display pleiotropic functions during different stages of the DNA-damage response. Moreover, several complexes seem to have multiple functions, and are implicated in various mechanistically distinct repair pathways.

The effectiveness of many DNA-damaging chemotherapeutic drugs depends on their ability to form monoadducts, intrastrand crosslinks and/or interstrand crosslinks (ICLs) that interfere with transcription and replication. The ERCC1–XPF endonuclease plays a critical role in removal of these lesions by incising DNA either as part of nucleotide excision repair (NER) or interstrand crosslink repair (ICLR). Engagement of ERCC1–XPF in NER is well characterized and is facilitated by binding to the XPA protein. However, ERCC1–XPF recruitment to ICLs is less well understood. Moreover, specific mutations in XPF have been found to disrupt its function in ICLR but not in NER, but whether this involves differences in lesion targeting is unknown. Here, we imaged GFP-tagged ERCC1, XPF and ICLR-defective XPF mutants to investigate how in human cells ERCC1–XPF is localized to different types of psoralen-induced DNA lesions, repaired by either NER or ICLR. Our results confirm its dependence on XPA in NER and furthermore show that its engagement in ICLR is dependent on FANCD2. Interestingly, we find that two ICLR-defective XPF mutants (R689S and S786F) are less well recruited to ICLs. These studies highlight the differential mechanisms that regulate ERCC1–XPF activity in DNA repair.

DNA–protein crosslinks (DPCs) arise from enzymatic intermediates, metabolism or chemicals like chemotherapeutics. DPCs are highly cytotoxic as they impede DNA-based processes such as replication, which is counteracted through proteolysis-mediated DPC removal by spartan (SPRTN) or the proteasome. However, whether DPCs affect transcription and how transcription-blocking DPCs are repaired remains largely unknown. Here we show that DPCs severely impede RNA polymerase II-mediated transcription and are preferentially repaired in active genes by transcription-coupled DPC (TC-DPC) repair. TC-DPC repair is initiated by recruiting the transcription-coupled nucleotide excision repair (TC-NER) factors CSB and CSA to DPC-stalled RNA polymerase II. CSA and CSB are indispensable for TC-DPC repair; however, the downstream TC-NER factors UVSSA and XPA are not, a result indicative of a non-canonical TC-NER mechanism. TC-DPC repair functions independently of SPRTN but is mediated by the ubiquitin ligase CRL4 CSA and the proteasome. Thus, DPCs in genes are preferentially repaired in a transcription-coupled manner to facilitate unperturbed transcription. Three studies identify a transcription-coupled DNA–protein crosslink repair pathway that depends on the Cockayne syndrome proteins and the proteasome.

XPC recognizes UV-induced DNA lesions and initiates their removal by nucleotide excision repair (NER). Damage recognition in NER is tightly controlled by ubiquitin and SUMO modifications. Recent studies have shown that the SUMO-targeted ubiquitin ligase RNF111 promotes K63-linked ubiquitylation of SUMOylated XPC after DNA damage. However, the exact regulatory function of these modifications in vivo remains elusive. Here we show that RNF111 is required for efficient repair of ultraviolet-induced DNA lesions. RNF111-mediated ubiquitylation promotes the release of XPC from damaged DNA after NER initiation, and is needed for stable incorporation of the NER endonucleases XPG and ERCC1/XPF. Our data suggest that RNF111, together with the CRL4 DDB2 ubiquitin ligase complex, is responsible for sequential XPC ubiquitylation, which regulates the recruitment and release of XPC and is crucial for efficient progression of the NER reaction, thereby providing an extra layer of quality control of NER. The SUMO-targeted ubiquitin ligase RNF111 promotes K63-linked ubiquitylation of SUMOylated XPC after DNA damage. Here the authors show that RNF111 is responsible for sequential XPC ubiquitylation, and RNF111-mediated ubiquitylation promotes the release of XPC from damaged DNA after NER initiation.

Congenital nucleotide excision repair (NER) deficiency gives rise to several cancer-prone and/or progeroid disorders. It is not understood how defects in the same DNA repair pathway cause different disease features and severity. Here, we show that the absence of functional ERCC1-XPF or XPG endonucleases leads to stable and prolonged binding of the transcription/DNA repair factor TFIIH to DNA damage, which correlates with disease severity and induces senescence features in human cells. In vivo, in C. elegans , this prolonged TFIIH binding to non-excised DNA damage causes developmental arrest and neuronal dysfunction, in a manner dependent on transcription-coupled NER. NER factors XPA and TTDA both promote stable TFIIH DNA binding and their depletion therefore suppresses these severe phenotypical consequences. These results identify stalled NER intermediates as pathogenic to cell functionality and organismal development, which can in part explain why mutations in XPF or XPG cause different disease features than mutations in XPA or TTDA. Hereditary nucleotide excision repair deficiencies cause different cancerous and progeroid disorders of which the exact etiology is not understood. This study finds that prolonged binding of DNA repair factor TFIIH to DNA damage contributes to a more severe phenotype caused by DNA repair deficiency.

Mutations in SWI/SNF genes are amongst the most common across all human cancers, but efficient therapeutic approaches that exploit vulnerabilities caused by SWI/SNF mutations are currently lacking. Here, we show that the SWI/SNF ATPases BRM/SMARCA2 and BRG1/SMARCA4 promote the expression of p62/GTF2H1 , a core subunit of the transcription factor IIH (TFIIH) complex. Inactivation of either ATPase subunit downregulates GTF2H1 and therefore compromises TFIIH stability and function in transcription and nucleotide excision repair (NER). We also demonstrate that cells with permanent BRM or BRG1 depletion have the ability to restore GTF2H1 expression. As a consequence, the sensitivity of SWI/SNF-deficient cells to DNA damage induced by UV irradiation and cisplatin treatment depends on GTF2H1 levels. Together, our results expose GTF2H1 as a potential novel predictive marker of platinum drug sensitivity in SWI/SNF-deficient cancer cells. SWI/SNF genes are commonly found to be mutated in different cancers. Here the authors report that the remodelers BRM and BRG1 are necessary for efficient nucleotide excision repair by promoting the expression of TFIIH subunit GTF2H1.

Transcription-blocking DNA lesions are specifically targeted by transcription-coupled nucleotide excision repair (TC-NER), which removes a broad spectrum of DNA lesions to preserve transcriptional output and thereby cellular homeostasis to counteract aging. TC-NER is initiated by the stalling of RNA polymerase II at DNA lesions, which triggers the assembly of the TC-NER-specific proteins CSA, CSB and UVSSA. CSA, a WD40-repeat containing protein, is the substrate receptor subunit of a cullin-RING ubiquitin ligase complex composed of DDB1, CUL4A/B and RBX1 (CRL4 CSA ). Although ubiquitination of several TC-NER proteins by CRL4 CSA has been reported, it is still unknown how this complex is regulated. To unravel the dynamic molecular interactions and the regulation of this complex, we apply a single-step protein-complex isolation coupled to mass spectrometry analysis and identified DDA1 as a CSA interacting protein. Cryo-EM analysis shows that DDA1 is an integral component of the CRL4 CSA complex. Functional analysis reveals that DDA1 coordinates ubiquitination dynamics during TC-NER and is required for efficient turnover and progression of this process. Transcription-Coupled Nucleotide Excision Repair (TC-NER) removes transcription-blocking DNA lesions. This study reveals that DDA1 is a crucial player in TC-NER, protecting transcription programs against genotoxic insults.

Cells employ global genome nucleotide excision repair (GGR) to eliminate a broad spectrum of DNA lesions, including those induced by UV light. The lesion-recognition factor XPC initiates repair of helix-destabilizing DNA lesions, but binds poorly to lesions such as CPDs that do not destabilize DNA. How difficult-to-repair lesions are detected in chromatin is unknown. Here, we identify the poly-(ADP-ribose) polymerases PARP1 and PARP2 as constitutive interactors of XPC. Their interaction results in the XPC-stimulated synthesis of poly-(ADP-ribose) (PAR) by PARP1 at UV lesions, which in turn enables the recruitment and activation of the PAR-regulated chromatin remodeler ALC1. PARP2, on the other hand, modulates the retention of ALC1 at DNA damage sites. Notably, ALC1 mediates chromatin expansion at UV-induced DNA lesions, leading to the timely clearing of CPD lesions. Thus, we reveal how chromatin containing difficult-to-repair DNA lesions is primed for repair, providing insight into mechanisms of chromatin plasticity during GGR. Cells employ global genome nucleotide excision repair to repair a broad spectrum of genomic DNA lesions. Here, the authors reveal how chromatin is primed for repair, providing insight into mechanisms of chromatin plasticity during DNA repair.