Cockayne syndrome protein B (CSB) is a member of the SNF2/SWI2 ATPase family and is essential for transcription-coupled nucleotide excision DNA repair (TC-NER). CSB also plays critical roles in transcription regulation. CSB can hydrolyze ATP in a DNA-dependent manner, alter protein-DNA contacts and anneal DNA strands. How the different biochemical activities of CSB are utilized in these cellular processes have only begun to become clear in recent years. Mutations in the gene encoding CSB account for majority of the Cockayne syndrome cases, which result in extreme sun sensitivity, premature aging features and/or abnormalities in neurology and development. Here, we summarize and integrate recent biochemical, structural, single-molecule and somatic cell genetic studies that have advanced our understanding of CSB. First, we review studies on the mechanisms that regulate the different biochemical activities of CSB. Next, we summarize how CSB is targeted to regulate transcription under different growth conditions. We then discuss recent advances in our understanding of how CSB regulates transcription mechanistically. Lastly, we summarize the various roles that CSB plays in the different steps of TC-NER, integrating the results of different studies and proposing a model as to how CSB facilitates TC-NER.

Efficient repair of oxidized DNA is critical for genome-integrity maintenance. Cockayne syndrome protein B (CSB) is an ATP-dependent chromatin remodeler that collaborates with Poly(ADP-ribose) polymerase I (PARP1) in the repair of oxidative DNA lesions. How these proteins integrate during DNA repair remains largely unknown. Here, using chromatin co-fractionation studies, we demonstrate that PARP1 and PARP2 promote recruitment of CSB to oxidatively-damaged DNA. CSB, in turn, contributes to the recruitment of XRCC1, and histone PARylation factor 1 (HPF1), and promotes histone PARylation. Using alkaline comet assays to monitor DNA repair, we found that CSB regulates single-strand break repair (SSBR) mediated by PARP1 and PARP2. Strikingly, CSB’s function in SSBR is largely bypassed when transcription is inhibited, suggesting CSB-mediated SSBR occurs primarily at actively transcribed DNA regions. While PARP1 repairs SSBs at sites regardless of the transcription status, we found that PARP2 predominantly functions in actively transcribed DNA regions. Therefore, our study raises the hypothesis that SSBR is executed by different mechanisms based on the transcription status.

Faithful propagation of transcription programs through cell division underlies cell-identity maintenance. Transcriptional regulators selectively bound on mitotic chromatin are emerging critical elements for mitotic transcriptional memory; however, mechanisms governing their site-selective binding remain elusive. By studying how protein-protein interactions impact mitotic chromatin binding of RBPJ, the major downstream effector of the Notch signaling pathway, we found that histone modifying enzymes HDAC1 and KDM5A play critical, regulatory roles in this process. We found that HDAC1 knockdown or inactivation leads to increased RBPJ occupancy on mitotic chromatin in a site-specific manner, with a concomitant increase of KDM5A occupancy at these sites. Strikingly, the presence of KDM5A is essential for increased RBPJ occupancy. Our results uncover a regulatory mechanism in which HDAC1 negatively regulates RBPJ binding on mitotic chromatin in a KDM5A-dependent manner. We propose that relative chromatin affinity of a minimal regulatory complex, reflecting a specific transcription program, renders selective RBPJ binding on mitotic chromatin.

Cockayne syndrome protein B (CSB) is an ATP-dependent chromatin remodeler that relieves oxidative stress by regulating DNA repair and transcription. CSB is proposed to participate in base-excision repair (BER), the primary pathway for repairing oxidative DNA damage, but exactly how CSB participates in this process is unknown. It is also unclear whether CSB contributes to other repair pathways during oxidative stress. Here, using a patient-derived CS1AN-sv cell line, we examined how CSB is targeted to chromatin in response to menadione-induced oxidative stress, both globally and locus-specifically. We found that menadione-induced, global CSB–chromatin association does not require CSB’s ATPase activity and is, therefore, mechanistically distinct from UV-induced CSB–chromatin association. Importantly, poly(ADP-ribose) polymerase 1 (PARP1) enhanced the kinetics of global menadione-induced CSB–chromatin association. We found that the major BER enzymes, 8-oxoguanine DNA glycosylase (OGG1) and apurinic/apyrimidinic endodeoxyribonuclease 1 (APE1), do not influence this association. Additionally, the level of γ-H2A histone family member X (γ-H2AX), a marker for dsDNA breaks, was not increased in menadione-treated cells. Therefore, our results support a model whereby PARP1 localizes to ssDNA breaks and recruits CSB to participate in DNA repair. Furthermore, this global CSB–chromatin association occurred independently of RNA polymerase II–mediated transcription elongation. However, unlike global CSB–chromatin association, both PARP1 knockdown and inhibition of transcription elongation interfered with menadione-induced CSB recruitment to specific genomic regions. This observation supports the hypothesis that CSB is also targeted to specific genomic loci to participate in transcriptional regulation in response to oxidative stress.

Oxidative stress contributes to numerous diseases, including cancer. CSB is an ATP-dependent chromatin remodeler critical for oxidative stress relief. PARP1 is the major sensor for DNA breaks and fundamental for efficient single-strand break repair. DNA breaks activate PARP1, leading to the synthesis of poly(ADP-ribose) (PAR) on itself and neighboring proteins, which is crucial for the recruitment of DNA repair machinery. CSB and PARP1 interact; however, how CSB mechanistically participates in oxidative DNA damage repair mediated by PARP1 remains unclear. Using chromatin immunoprecipitation followed by quantitative PCR, we found that CSB and PARP1 facilitate each other’s chromatin association during the onset of oxidative stress, and that CSB facilitates PARP1 removal when the level of chromatin-bound CSB increases. Furthermore, by monitoring chromatin PAR levels using Western blot analysis, we found that CSB sustains the DNA damage signal initiated by PARP1, and may prevent PARP1 overactivation by facilitating DNA repair. By assaying cell viability in response to oxidative stress, we further demonstrate that PARP1 regulation by CSB is a major CSB function in oxidatively-stressed cells. Together, our study uncovers a dynamic interplay between CSB and PARP1 that is critical for oxidative stress relief.

Efficient DNA lesion repair is crucial for cell survival, especially within actively transcribed DNA regions that contain essential genetic information. Additionally, DNA breaks in regions of active transcription are prone to generating insertions and deletions, which are hallmark features of cancer genomes. Cockayne syndrome protein B (CSB) is the sole ATP-dependent chromatin remodeler that is essential for coupling DNA repair pathways with transcription, leading to more efficient DNA repair in regions of active transcription. CSB is best known for its essential function in transcription-coupled nucleotide excision repair (TC-NER), a process that rapidly removes helix-distorting DNA lesions that stall RNA polymerase II, such as those created by chemotherapeutic platinum compounds and UV irradiation. In addition to NER, CSB has also been reported to couple homologous recombination to transcription. Most recently, CSB has also been shown to couple single-strand DNA break repair to transcription. In this review, we will discuss the overlapping and distinct mechanisms by which CSB couples these different DNA repair pathways to transcription. We will also discuss how these CSB functions may account for Cockayne syndrome and the emerging roles of CSB as an innovative target for cancer therapy.