Search

Increasing evidence supports the involvement of DNA damage in several neurodegenerative diseases, including amyotrophic lateral sclerosis (ALS). Elevated levels of DNA damage are consistently observed in both sporadic and familial forms of ALS and may also play a role in Western Pacific ALS, which is thought to have an environmental cause. The cause of DNA damage in ALS remains unclear but likely differs between genetic subgroups. Repeat expansion in the C9ORF72 gene is the most common genetic cause of familial ALS and responsible for about 10% of sporadic cases. These genetic mutations are known to cause R-loops, thus increasing genomic instability and DNA damage, and generate dipeptide repeat proteins, which have been shown to lead to DNA damage and impairment of the DNA damage response. Similarly, several genes associated with ALS including TARDBP, FUS, NEK1, SQSTM1 and SETX are known to play a role in DNA repair and the DNA damage response, and thus may contribute to neuronal death via these pathways. Another consistent feature present in both sporadic and familial ALS is the ability of astrocytes to induce motor neuron death, although the factors causing this toxicity remain largely unknown. In this review, we summarise the evidence for DNA damage playing a causative or secondary role in the pathogenesis of ALS as well as discuss the possible mechanisms involved in different genetic subtypes with particular focus on the role of astrocytes initiating or perpetuating DNA damage in neurons.

R-loops are by-products of transcription that must be tightly regulated to maintain genomic stability and gene expression. Here, we describe a mechanism for the regulation of the R-loop-specific helicase, senataxin (SETX), and identify the ubiquitin specific peptidase 11 (USP11) as an R-loop regulator. USP11 de-ubiquitinates SETX and its depletion increases SETX K48-ubiquitination and protein turnover. Loss of USP11 decreases SETX steady-state levels and reduces R-loop dissolution. Ageing of USP11 knockout cells restores SETX levels via compensatory transcriptional downregulation of the E3 ubiquitin ligase, KEAP1. Loss of USP11 reduces SETX enrichment at KEAP1 promoter, leading to R-loop accumulation, enrichment of the endonuclease XPF and formation of double-strand breaks. Overexpression of KEAP1 increases SETX K48-ubiquitination, promotes its degradation and R-loop accumulation. These data define a ubiquitination-dependent mechanism for SETX regulation, which is controlled by the opposing activities of USP11 and KEAP1 with broad applications for cancer and neurological disease., DNA:RNA hybrids (R-loops) are products of transcription that impact genome integrity and gene expression. Here the authors reveal a mechanism for regulating R-loops in a ubiquitination-dependent manner controlled by the activities of USP11 and KEAP1

Tumour hypoxia is associated with poor patient prognosis and therapy resistance. A unique transcriptional response is initiated by hypoxia which includes the rapid activation of numerous transcription factors in a background of reduced global transcription. Here, we show that the biological response to hypoxia includes the accumulation of R-loops and the induction of the RNA/DNA helicase SETX. In the absence of hypoxia-induced SETX, R-loop levels increase, DNA damage accumulates, and DNA replication rates decrease. Therefore, suggesting that, SETX plays a role in protecting cells from DNA damage induced during transcription in hypoxia. Importantly, we propose that the mechanism of SETX induction in hypoxia is reliant on the PERK/ATF4 arm of the unfolded protein response. These data not only highlight the unique cellular response to hypoxia, which includes both a replication stress-dependent DNA damage response and an unfolded protein response but uncover a novel link between these two distinct pathways., Hypoxia induces a change in transcriptional response in mammalian cells. Here the authors reveal a role for the RNA/DNA helicase Senataxin in protecting cells from DNA damage induced during transcription in hypoxia.

Eukaryotic topoisomerase 1 (TOP1) regulates DNA topology to ensure efficient DNA replication and transcription. TOP1 is also a major driver of endogenous genome instability, particularly when its catalytic intermediate—a covalent TOP1-DNA adduct known as a TOP1 cleavage complex (TOP1cc)—is stabilised. TOP1ccs are highly cytotoxic and a failure to resolve them underlies the pathology of neurological disorders but is also exploited in cancer therapy where TOP1ccs are the target of widely used frontline anti-cancer drugs. A critical enzyme for TOP1cc resolution is the tyrosyl-DNA phosphodiesterase (TDP1), which hydrolyses the bond that links a tyrosine in the active site of TOP1 to a 3’ phosphate group on a single-stranded (ss)DNA break. However, TDP1 can only process small peptide fragments from ssDNA ends, raising the question of how the ~90 kDa TOP1 protein is processed upstream of TDP1. Here we find that TEX264 fulfils this role by forming a complex with the p97 ATPase and the SPRTN metalloprotease. We show that TEX264 recognises both unmodified and SUMO1-modifed TOP1 and initiates TOP1cc repair by recruiting p97 and SPRTN. TEX264 localises to the nuclear periphery, associates with DNA replication forks, and counteracts TOP1ccs during DNA replication. Altogether, our study elucidates the existence of a specialised repair complex required for upstream proteolysis of TOP1ccs and their subsequent resolution., Eukaryotic topoisomerase 1 (TOP1) regulates DNA topology to ensure efficient DNA replication and transcription. Here, the authors reveal insights into the molecular resolution of topoisomerase 1-DNA adducts by TEX264, p97 and SPRTN.

Salmonella Typhi activates the host DNA damage response through the typhoid toxin, facilitating typhoid symptoms and chronic infections. Here we reveal a non-canonical DNA damage response, which we call RING (response induced by a genotoxin), characterized by accumulation of phosphorylated histone H2AX (γH2AX) at the nuclear periphery. RING is the result of persistent DNA damage mediated by toxin nuclease activity and is characterized by hyperphosphorylation of RPA, a sensor of single-stranded DNA (ssDNA) and DNA replication stress. The toxin overloads the RPA pathway with ssDNA substrate, causing RPA exhaustion and senescence. Senescence is also induced by canonical γΗ2ΑΧ foci revealing distinct mechanisms. Senescence is transmitted to non-intoxicated bystander cells by an unidentified senescence-associated secreted factor that enhances Salmonella infections. Thus, our work uncovers a mechanism by which genotoxic Salmonella exhausts the RPA response by inducing ssDNA formation, driving host cell senescence and facilitating infection., Salmonella Typhi produces the typhoid toxin, which damages host cell DNA. Here, Ibler et al. show that the toxin induces a non-canonical DNA damage response mediated by the RPA pathway that drives host cell senescence and facilitates infection.

The mammalian genome is constantly challenged by exogenous and endogenous threats. Although much is known about the mechanisms that maintain DNA and RNA integrity, we know surprisingly little about the mechanisms that underpin the pathology and tissue specificity of many disorders caused by defective responses to DNA or RNA damage. Of the different types of endogenous damage, protein-linked DNA breaks (PDBs) are emerging as an important player in cancer development and therapy. PDBs can arise during the abortive activity of DNA topoisomerases, a class of enzymes that modulate DNA topology during several chromosomal transactions, such as gene transcription and DNA replication, recombination and repair. In this Review, we discuss the mechanisms underpinning topoisomerase-induced PDB formation and repair with a focus on their role during gene transcription and the development of tissue-specific cancers.

The topoisomerase I (Top 1) poison irinotecan is an important component of the modern treatment of colorectal cancer. By stabilising Top 1-DNA complexes, irinotecan generates Top 1-linked DNA single-strand breaks that can evolve into double-strand breaks and ultimately cause cell death. However, cancer cells may overcome cell killing by releasing the stalled topoisomerase from DNA termini, thereby reducing the efficacy of Top 1 poisons in clinics. Thus, understanding the DNA repair mechanisms involved in the repair of Top 1-mediated DNA damage provides a useful tool to identify potential biomarkers that predict response to this class of chemotherapy. Furthermore, targeting these pathways could enhance the therapeutic benefits of Top 1 poisons. In this review, we describe the cellular mechanisms and consequences of targeting Top 1 activity in cells. We summarise preclinical data and discuss the potential clinical utility of small-molecule inhibitors of the key proteins.

Ataxia oculomotor apraxia-1 (AOA1) is a neurological disorder caused by mutations in the gene (APTX) encoding aprataxin1, 2. Aprataxin is a member of the histidine triad (HIT) family of nucleotide hydrolases and transferases3, and inactivating mutations are largely confined to this HIT domain. Aprataxin associates with the DNA repair proteins XRCC1 and XRCC4, which are partners of DNA ligase III and ligase IV, respectively4, 5, 6, 7, suggestive of a role in DNA repair. Consistent with this, APTX-defective cell lines are sensitive to agents that cause single-strand breaks and exhibit an increased incidence of induced chromosomal aberrations4, 5, 8. It is not, however, known whether aprataxin has a direct or indirect role in DNA repair, or what the physiological substrate of aprataxin might be. Here we show, using purified aprataxin protein and extracts derived from either APTX-defective chicken DT40 cells or Aptx-/- mouse primary neural cells, that aprataxin resolves abortive DNA ligation intermediates. Specifically, aprataxin catalyses the nucleophilic release of adenylate groups covalently linked to 5'-phosphate termini at single-strand nicks and gaps, resulting in the production of 5'-phosphate termini that can be efficiently rejoined. These data indicate that neurological disorders associated with APTX mutations may be caused by the gradual accumulation of unrepaired DNA strand breaks resulting from abortive DNA ligation events.

Spinocerebellar ataxia with axonal neuropathy-1 (SCAN1) is a neurodegenerative disease that results from mutation of tyrosyl phosphodiesterase 1 (TDP1). In lower eukaryotes, Tdp1 removes topoisomerase 1 (top1) peptide from DNA termini during the repair of double-strand breaks created by collision of replication forks with top1 cleavage complexes in proliferating cells. Although TDP1 most probably fulfils a similar function in human cells, this role is unlikely to account for the clinical phenotype of SCAN1, which is associated with progressive degeneration of post-mitotic neurons. In addition, this role is redundant in lower eukaryotes, and Tdp1 mutations alone confer little phenotype. Moreover, defects in processing or preventing double-strand breaks during DNA replication are most probably associated with increased genetic instability and cancer, phenotypes not observed in SCAN1 (ref. 8). Here we show that in human cells TDP1 is required for repair of chromosomal single-strand breaks arising independently of DNA replication from abortive top1 activity or oxidative stress. We report that TDP1 is sequestered into multi-protein single-strand break repair (SSBR) complexes by direct interaction with DNA ligase IIIalpha and that these complexes are catalytically inactive in SCAN1 cells. These data identify a defect in SSBR in a neurodegenerative disease, and implicate this process in the maintenance of genetic integrity in post-mitotic neurons.

Breaking and sealing one strand of DNA is an inherent feature of chromosome metabolism to overcome torsional barriers. Failure to reseal broken DNA strands results in protein-linked DNA breaks, causing neurodegeneration in humans. This is typified by defects in tyrosyl DNA phosphodiesterase 1 (TDP1), which removes stalled topoisomerase 1 peptides from DNA termini. Here we show that TDP1 is a substrate for modification by the small ubiquitin-like modifier SUMO. We purify SUMOylated TDP1 from mammalian cells and identify the SUMOylation site as lysine 111. While SUMOylation exhibits no impact on TDP1 catalytic activity, it promotes its accumulation at sites of DNA damage. A TDP1 SUMOylation-deficient mutant displays a reduced rate of repair of chromosomal single-strand breaks arising from transcription-associated topoisomerase 1 activity or oxidative stress. These data identify a role for SUMO during single-strand break repair, and suggest a mechanism for protecting the nervous system from genotoxic stress., Tyrosyl DNA phosphodiesterase 1 (TDP1) repairs DNA breaks and is mutated in the disease Spinocerebellar Ataxia with Axonal Neuropathy. Here TDP1 is shown to be post-translationally modified by sumoylation of lysine 111, and cells carrying a mutation at this residue are inefficient at single-strand DNA break repair.