XRCC1 - Strategies for coordinating and assembling a versatile DNA damage response.
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ABSTRACT:
SUBMITTER: London RE
PROVIDER: S-EPMC7587305 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:The DNA damage response (DDR) has an essential function in maintaining genomic stability. Ataxia telangiectasia-mutated (ATM)-checkpoint kinase 2 (Chk2) and ATM- and Rad3-related (ATR)-Chk1, triggered, respectively, by DNA double-strand breaks and blocked replication forks, are two major DDRs processing structurally complicated DNA damage. In contrast, damage repaired by base excision repair (BER) is structurally simple, but whether, and how, the DDR is involved in repairing this damage is unclear. Here, we demonstrated that ATM-Chk2 was activated in the early response to oxidative and alkylation damage, known to be repaired by BER. Furthermore, Chk2 formed a complex with XRCC1, the BER scaffold protein, and phosphorylated XRCC1 in vivo and in vitro at Thr(284). A mutated XRCC1 lacking Thr(284) phosphorylation was linked to increased accumulation of unrepaired BER intermediate, reduced DNA repair capacity, and higher sensitivity to alkylation damage. In addition, a phosphorylation-mimic form of XRCC1 showed increased interaction with glycosylases, but not other BER proteins. Our results are consistent with the phosphorylation of XRCC1 by ATM-Chk2 facilitating recruitment of downstream BER proteins to the initial damage recognition/excision step to promote BER.
| S-EPMC2599873 | biostudies-other
Project description:On induction of DNA damage with 405-nm laser light, proteins involved in base excision repair (BER) are recruited to DNA lesions. We find that the dynamics of factors typical of either short-patch (XRCC1) or long-patch (PCNA) BER are altered by chemicals that perturb actin or tubulin polymerization in human cells. Whereas the destabilization of actin filaments by latrunculin B, cytochalasin B, or Jasplakinolide decreases BER factor accumulation at laser-induced damage, inhibition of tubulin polymerization by nocodazole increases it. We detect no recruitment of actin to sites of laser-induced DNA damage, yet the depolymerization of cytoplasmic actin filaments elevates both actin and tubulin signals in the nucleus. While published evidence suggested a positive role for F-actin in double-strand break repair in mammals, the enrichment of actin in budding yeast nuclei interferes with BER, augmenting sensitivity to Zeocin. Our quantitative imaging results suggest that the depolymerization of cytoplasmic actin may compromise BER efficiency in mammals not only due to elevated levels of nuclear actin but also of tubulin, linking cytoskeletal integrity to BER.
| S-EPMC8684753 | biostudies-literature
Project description:Poly(ADP-ribose) polymerase-1 (PARP-1) binds intermediates of base excision repair (BER) and becomes activated for poly(ADP-ribose) (PAR) synthesis. PAR mediates recruitment and functions of the key BER factors XRCC1 and DNA polymerase β (pol β) that in turn regulate PAR. Yet, the molecular mechanism and implications of coordination between XRCC1 and pol β in regulating the level of PAR are poorly understood. A complex of PARP-1, XRCC1 and pol β is found in vivo, and it is known that pol β and XRCC1 interact through a redox-sensitive binding interface in the N-terminal domain of XRCC1. We confirmed here that both oxidized and reduced forms of XRCC1 are present in mouse fibroblasts. To further understand the importance of the C12-C20 oxidized form of XRCC1 and the interaction with pol β, we characterized cell lines representing stable transfectants in Xrcc1(-/-) mouse fibroblasts of wild-type XRCC1 and two mutants of XRCC1, a novel reduced form with the C12-C20 disulfide bond blocked (C12A) and a reference mutant that is unable to bind pol β (V88R). XRCC1-deficient mouse fibroblasts are extremely hypersensitive to methyl methanesulfonate (MMS), and transfected wild-type and C12A mutant XRCC1 proteins similarly reversed MMS hypersensitivity. However, after MMS exposure the cellular PAR level was found to increase to a much greater extent in cells expressing the C12A mutant than in cells expressing wild-type XRCC1. PARP inhibition resulted in very strong MMS sensitization in cells expressing wild-type XRCC1, but this sensitization was much less in cells expressing the C12A mutant. The results suggest a role for the oxidized form of XRCC1 in the interaction with pol β in (1) controlling the PAR level after MMS exposure and (2) enabling the extreme cytotoxicity of PARP inhibition during the MMS DNA damage response.
| S-EPMC3924596 | biostudies-literature
Project description:Alternative reading frame (ARF) is a tumor suppressor protein that senses oncogenic and other stressogenic signals. It can trigger p53-dependent and -independent responses with cell cycle arrest and apoptosis induction being the most prominent ones. Other ARF activities, particularly p53-independent ones, that could help in understanding cancer development and provide potential therapeutic exploitation are underrated. Although ARF is generally not expressed in normal tissues, it is essential for ocular and male germ cells development. The underlying mechanism(s) in these processes, while not clearly defined, point toward a functional link between ARF, DNA damage and angiogenesis. Based on a recent study from our group demonstrating a functional interplay between ataxia-telangiectasia mutated (ATM) and ARF during carcinogenesis, we discuss the role of ARF at the crossroads of cancer and developmental processes.
| S-EPMC4106421 | biostudies-literature
Project description:DNA and RNA have emerged as a material for nanotechnology applications that take advantage of the nucleic acids' ability to encode folding and programmable self-assembly through mainly base pairing. The two types of nucleic acid have rarely been used in combination to enhance structural diversity or for partitioning of functional and architectural roles. Here, we report a design and screening strategy to integrate combinations of RNA motifs as architectural joints and DNA building blocks as functional modules for programmable self-assembly of a versatile toolkit of polygonal nucleic acid nanoshapes. Clean incorporation of diverse DNA modules with various topologies attest to the extraordinary robustness of the RNA-DNA hybrid framework. The design and screening strategy enables systematic development of RNA-DNA hybrid nanoshapes as programmable platforms for applications in molecular recognition, sensor and catalyst development as well as protein interaction studies.
| S-EPMC6363791 | biostudies-literature
Project description:Oxidative damage to DNA has important implications for human health and has been identified as a key factor in the onset and development of numerous diseases. Thus, it is evident that preventing DNA from oxidative damage is crucial for humans and for any living organism. Melatonin is an astonishingly versatile molecule in this context. It can offer both direct and indirect protection against a wide variety of damaging agents and through multiple pathways, which may (or may not) take place simultaneously. They include direct antioxidative protection, which is mediated by melatonin's free radical scavenging activity, and also indirect ways of action. The latter include, at least: (i) inhibition of metal-induced DNA damage; (ii) protection against non-radical triggers of oxidative DNA damage; (iii) continuous protection after being metabolized; (iv) activation of antioxidative enzymes; (v) inhibition of pro-oxidative enzymes; and (vi) boosting of the DNA repair machinery. The rather unique capability of melatonin to exhibit multiple neutralizing actions against diverse threatening factors, together with its low toxicity and its ability to cross biological barriers, are all significant to its efficiency for preventing oxidative damage to DNA.
| S-EPMC6017920 | biostudies-literature
Project description:Maintenance of genomic stability is a critical determinant of cell survival and relies on the coordinated action of the DNA damage response (DDR), which orchestrates a network of cellular processes, including DNA replication, DNA repair and cell-cycle progression. In cancer, the critical balance between the loss of genomic stability in malignant cells and the DDR provides exciting therapeutic opportunities. Drugs targeting DDR pathways taking advantage of clinical synthetic lethality have already shown therapeutic benefit - for example, the PARP inhibitor olaparib has shown benefit in BRCA-mutant ovarian and breast cancer. Olaparib has also shown benefit in metastatic prostate cancer in DDR-defective patients, expanding the potential biomarker of response beyond BRCA. Other agents and combinations aiming to block the DDR while pushing damaged DNA through the cell cycle, including PARP, ATR, ATM, CHK and DNA-PK inhibitors, are in development. Emerging work is also uncovering how the DDR interacts intimately with the host immune response, including by activating the innate immune response, further suggesting that clinical applications together with immunotherapy may be beneficial. Here, we review recent considerations related to the DDR from a clinical standpoint, providing a framework to address future directions and clinical opportunities.
| S-EPMC6047242 | biostudies-literature
Project description:Transcription-coupled nucleotide excision repair factor Cockayne syndrome protein B (CSB) was suggested to function in the repair of oxidative DNA damage. However thus far, no clear role for CSB in base excision repair (BER), the dedicated pathway to remove abundant oxidative DNA damage, could be established. Using live cell imaging with a laser-assisted procedure to locally induce 8-oxo-7,8-dihydroguanine (8-oxoG) lesions, we previously showed that CSB is recruited to these lesions in a transcription-dependent but NER-independent fashion. Here we showed that recruitment of the preferred 8-oxoG-glycosylase 1 (OGG1) is independent of CSB or active transcription. In contrast, recruitment of the BER-scaffolding protein, X-ray repair cross-complementing protein 1 (XRCC1), to 8-oxoG lesions is stimulated by CSB and transcription. Remarkably, recruitment of XRCC1 to BER-unrelated single strand breaks (SSBs) does not require CSB or transcription. Together, our results suggest a specific transcription-dependent role for CSB in recruiting XRCC1 to BER-generated SSBs, whereas XRCC1 recruitment to SSBs generated independently of BER relies predominantly on PARP activation. Based on our results, we propose a model in which CSB plays a role in facilitating BER progression at transcribed genes, probably to allow XRCC1 recruitment to BER-intermediates masked by RNA polymerase II complexes stalled at these intermediates.
| S-EPMC6125634 | biostudies-literature
Project description:Repair of two major forms of DNA damage, single strand breaks and base modifications, are dependent on XRCC1. XRCC1 orchestrates these repair processes by temporally and spatially coordinating interactions between several other repair proteins. Here we show that XRCC1 contains a central DNA binding domain (CDB, residues 219-415) encompassing its first BRCT domain. In contrast to the N-terminal domain of XRCC1, which has been reported to mediate damage sensing in vitro, we demonstrate that the DNA binding module identified here lacks binding specificity towards DNA containing nicks or gaps. Alanine substitution of residues within the CDB of XRCC1 disrupt DNA binding in vitro and lead to a significant reduction in XRCC1 retention at DNA damage sites without affecting initial recruitment. Interestingly, reduced retention at sites of DNA damage is associated with an increased rate of repair. These findings suggest that DNA binding activity of XRCC1 plays a significant role in retention at sites of damage and the rate at which damage is repaired.
| S-EPMC6395731 | biostudies-literature
Project description:The members of the Poly(ADP-ribose) polymerase (PARP) superfamily are involved in several biological processes and, in particular, in the DNA damage response (DDR). The most studied members, PARP1, PARP2 and PARP3, act as sensors of DNA damages, in order to activate different intracellular repair pathways, including single-strand repair, homologous recombination, conventional and alternative non-homologous end joining. This review recapitulates the functional role of PARPs in the DDR pathways, also in relationship with the cell cycle phases, which drives our knowledge of the mechanisms of action of PARP inhibitors (PARPi), encompassing inhibition of single-strand breaks and base excision repair, PARP trapping and sensitization to antileukemia immune responses. Several studies have demonstrated a preclinical activity of the current available PARPi, olaparib, rucaparib, niraparib, veliparib and talazoparib, as single agent and/or in combination with cytotoxic, hypomethylating or targeted drugs in acute leukemia, thus encouraging the development of clinical trials. We here summarize the most recent preclinical and clinical findings and discuss the synthetic lethal interactions of PARPi in acute myeloid leukemia (AML) and acute lymphoblastic leukemia (ALL). Despite the low frequency of genomic alterations of PARP and other DDR-related genes in acute leukemia, selective vulnerabilities have been reported in several disease subgroups, along with a "BRCAness phenotype." AML carrying the RUNX1-RUNX1T1 or PML-RARA fusion genes or mutations in signaling genes (FLT3-ITD in combination with TET2 or TET2 and DNMT3A deficiency), cohesin complex members (STAG2), TP53 and BCOR as co-occurring lesions, IDH1/2 and ALL cases expressing the TCF3-HLF chimera or TET1 was highly sensitive to PARPi in preclinical studies. These data, along with the warning coming from the observation of cases of therapy-related myeloid malignancies among patients receiving PARPi for solid tumors treatment, indicate that PARPi represents a promising strategy in a personalized medicine setting. The characterization of the clonal and subclonal genetic background and of the DDR functionality is crucial to select acute leukemia patients that will likely benefit of PARPi-based therapeutic regimens.
| S-EPMC8783444 | biostudies-literature