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International Journal of Molecular... Apr 2022The damage and repair of DNA is a continuous process required to maintain genomic integrity. DNA double-strand breaks (DSBs) are the most lethal type of DNA damage and... (Review)
Review
The damage and repair of DNA is a continuous process required to maintain genomic integrity. DNA double-strand breaks (DSBs) are the most lethal type of DNA damage and require timely repair by dedicated machinery. DSB repair is uniquely important to nondividing, post-mitotic cells of the central nervous system (CNS). These long-lived cells must rely on the intact genome for a lifetime while maintaining high metabolic activity. When these mechanisms fail, the loss of certain neuronal populations upset delicate neural networks required for higher cognition and disrupt vital motor functions. Mammalian cells engage with several different strategies to recognize and repair chromosomal DSBs based on the cellular context and cell cycle phase, including homologous recombination (HR)/homology-directed repair (HDR), microhomology-mediated end-joining (MMEJ), and the classic non-homologous end-joining (NHEJ). In addition to these repair pathways, a growing body of evidence has emphasized the importance of DNA damage response (DDR) signaling, and the involvement of heterogeneous nuclear ribonucleoprotein (hnRNP) family proteins in the repair of neuronal DSBs, many of which are linked to age-associated neurological disorders. In this review, we describe contemporary research characterizing the mechanistic roles of these non-canonical proteins in neuronal DSB repair, as well as their contributions to the etiopathogenesis of selected common neurological diseases.
Topics: Animals; DNA; DNA Breaks, Double-Stranded; DNA End-Joining Repair; DNA Repair; Mammals; Nervous System Diseases; Recombinational DNA Repair
PubMed: 35563044
DOI: 10.3390/ijms23094653 -
Journal of Visualized Experiments : JoVE Jun 2018The repair of double-stranded breaks (DSBs) in DNA is a highly coordinated process, necessitating the formation and resolution of multi-protein repair complexes. This...
The repair of double-stranded breaks (DSBs) in DNA is a highly coordinated process, necessitating the formation and resolution of multi-protein repair complexes. This process is regulated by a myriad of proteins that promote the association and disassociation of proteins to these lesions. Thanks in large part to the ability to perform functional screens of a vast library of proteins, there is a greater appreciation of the genes necessary for the double-strand DNA break repair. Often knockout or chemical inhibitor screens identify proteins involved in repair processes by using increased toxicity as a marker for a protein that is required for DSB repair. Although useful for identifying novel cellular proteins involved in maintaining genome fidelity, functional analysis requires the determination of whether the protein of interest promotes localization, formation, or resolution of repair complexes. The accumulation of repair proteins can be readily detected as distinct nuclear foci by immunofluorescence microscopy. Thus, association and disassociation of these proteins at sites of DNA damage can be accessed by observing these nuclear foci at representative intervals after the induction of double-strand DNA breaks. This approach can also identify mis-localized repair factor proteins, if repair defects do not simultaneously occur with incomplete delays in repair. In this scenario, long-lasting double-strand DNA breaks can be engineered by expressing a rare cutting endonuclease (e.g., I-SceI) in cells where the recognition site for the said enzyme has been integrated into the cellular genome. The resulting lesion is particularly hard to resolve as faithful repair will reintroduce the enzyme's recognition site, prompting another round of cleavage. As a result, differences in the kinetics of repair are eliminated. If repair complexes are not formed, localization has been impeded. This protocol describes the methodology necessary to identify changes in repair kinetics as well as repair protein localization.
Topics: DNA Breaks, Double-Stranded; DNA Repair; DNA-Binding Proteins; Humans; Microscopy, Fluorescence
PubMed: 29939192
DOI: 10.3791/57653 -
Progress in Biophysics and Molecular... Jan 2024The proteins and protein assemblies involved in DNA repair have been the focus of a multitude of structural studies for the past few decades. Historically, the... (Review)
Review
The proteins and protein assemblies involved in DNA repair have been the focus of a multitude of structural studies for the past few decades. Historically, the structures of these protein complexes have been resolved by X-ray crystallography. However, more recently with the advancements in cryo-electron microscopy (cryo-EM) ranging from optimising the methodology for sample preparation to the development of improved electron detectors, the focus has shifted from X-ray crystallography to cryo-EM. This methodological transition has allowed for the structural determination of larger, more complex protein assemblies involved in DNA repair pathways and has subsequently led to a deeper understanding of the mechanisms utilised by these fascinating molecular machines. Here, we review some of the key structural advancements that have been gained in the study of non-homologous end joining (NHEJ) by the use of cryo-EM, with a focus on assemblies composed of DNA-PKcs and Ku70/80 (Ku) and the various methodologies utilised to obtain these structures.
Topics: Cryoelectron Microscopy; DNA Repair; DNA End-Joining Repair; DNA-Activated Protein Kinase; Crystallography, X-Ray; DNA
PubMed: 38036101
DOI: 10.1016/j.pbiomolbio.2023.11.007 -
Cells Jun 2021Protection of genome integrity is vital for all living organisms, particularly when DNA double-strand breaks (DSBs) occur. Eukaryotes have developed two main pathways,... (Review)
Review
Protection of genome integrity is vital for all living organisms, particularly when DNA double-strand breaks (DSBs) occur. Eukaryotes have developed two main pathways, namely Non-Homologous End Joining (NHEJ) and Homologous Recombination (HR), to repair DSBs. While most of the current research is focused on the role of key protein players in the functional regulation of DSB repair pathways, accumulating evidence has uncovered a novel class of regulating factors termed non-coding RNAs. Non-coding RNAs have been found to hold a pivotal role in the activation of DSB repair mechanisms, thereby safeguarding genomic stability. In particular, long non-coding RNAs (lncRNAs) have begun to emerge as new players with vast therapeutic potential. This review summarizes important advances in the field of lncRNAs, including characterization of recently identified lncRNAs, and their implication in DSB repair pathways in the context of tumorigenesis.
Topics: Animals; DNA; DNA Breaks, Double-Stranded; DNA Damage; DNA End-Joining Repair; DNA Repair; Genomic Instability; Humans; RNA, Long Noncoding; Recombinational DNA Repair
PubMed: 34203749
DOI: 10.3390/cells10061506 -
DNA Repair Oct 2023DNA double strand breaks (DSBs) are common lesions whose misrepair are drivers of oncogenic transformations. The non-homologous end joining (NHEJ) pathway repairs the... (Review)
Review
DNA double strand breaks (DSBs) are common lesions whose misrepair are drivers of oncogenic transformations. The non-homologous end joining (NHEJ) pathway repairs the majority of these breaks in vertebrates by directly ligating DNA ends back together. Upon formation of a DSB, a multiprotein complex is assembled on DNA ends which tethers them together within a synaptic complex. Synapsis is a critical step of the NHEJ pathway as loss of synapsis can result in mispairing of DNA ends and chromosome translocations. As DNA ends are commonly incompatible for ligation, the NHEJ machinery must also process ends to enable rejoining. This review describes how recent progress in single-molecule approaches and cryo-EM have advanced our molecular understanding of DNA end synapsis during NHEJ and how synapsis is coordinated with end processing to determine the fidelity of repair.
Topics: Animals; DNA End-Joining Repair; DNA; DNA-Binding Proteins; DNA Breaks, Double-Stranded; Chromosome Pairing; DNA Repair
PubMed: 37572577
DOI: 10.1016/j.dnarep.2023.103553 -
International Journal of Molecular... Dec 2016As a result of various stresses, lesions caused by DNA-damaging agents occur constantly in each cell of the human body. Generally, DNA damage is recognized and repaired... (Review)
Review
As a result of various stresses, lesions caused by DNA-damaging agents occur constantly in each cell of the human body. Generally, DNA damage is recognized and repaired by the DNA damage response (DDR) machinery, and the cells survive. When repair fails, the genomic integrity of the cell is disrupted-a hallmark of cancer. In addition, the DDR plays a dual role in cancer development and therapy. Cancer radiotherapy and chemotherapy are designed to eliminate cancer cells by inducing DNA damage, which in turn can promote tumorigenesis. Over the past two decades, an increasing number of microRNAs (miRNAs), small noncoding RNAs, have been identified as participating in the processes regulating tumorigenesis and responses to cancer treatment with radiation therapy or genotoxic chemotherapies, by modulating the DDR. The purpose of this review is to summarize the recent findings on how miRNAs regulate the DDR and discuss the therapeutic functions of miRNAs in cancer in the context of DDR regulation.
Topics: Antineoplastic Agents; DNA Damage; DNA Repair; Gene Expression Regulation, Neoplastic; Humans; MicroRNAs; Neoplasms
PubMed: 27973455
DOI: 10.3390/ijms17122087 -
Yonsei Medical Journal May 2010Mammalian cells are frequently at risk of DNA damage from both endogenous and exogenous sources. Accordingly, cells have evolved the DNA damage response (DDR) pathways... (Review)
Review
Mammalian cells are frequently at risk of DNA damage from both endogenous and exogenous sources. Accordingly, cells have evolved the DNA damage response (DDR) pathways to monitor and assure the integrity of their genome. In cells, the intact and effective DDR is essential for the maintenance of genomic stability and it acts as a critical barrier to suppress the development of cancer in humans. Two central kinases for the DDR pathway are ATM and ATR, which can phosphorylate and activate many downstream proteins for cell cycle arrest, DNA repair, or apoptosis if the damages are irreparable. In the last several years, we and others have made significant progress to this field by identifying BRIT1 (also known as MCPH1) as a novel key regulator in the DDR pathway. BRIT1 protein contains 3 breast cancer carboxyl terminal (BRCT) domains which are conserved in BRCA1, MDC1, 53BP1, and other important molecules involved in DNA damage signaling, DNA repair, and tumor suppression. Our in vitro studies revealed BRIT1 to be a chromatinbinding protein required for recruitment of many important DDR proteins (ATM, MDC1, NBS1, RAD51, BRCA2) to the DNA damage sites. We recently also generated the BRIT1 knockout mice and demonstrated its essential roles in homologous recombination DNA repair and in maintaining genomic stability in vivo. In humans, BRIT1 is located on chromosome 8p23.1, where loss of hetero-zigosity is very common in many types of cancer. In this review, we will summarize the novel roles of BRIT1 in DDR, describe the relationship of BRIT1 deficiency with cancer development, and also discuss the use of synthetic lethality approach to target cancers with HR defects due to BRIT1 deficiency.
Topics: Animals; Cell Cycle Proteins; Chromosomal Proteins, Non-Histone; Cytoskeletal Proteins; DNA Damage; DNA Repair; Humans; Mice; Models, Biological; Neoplasms; Nerve Tissue Proteins
PubMed: 20376879
DOI: 10.3349/ymj.2010.51.3.295 -
Carcinogenesis Jan 2010The reaction of DNA-damaging agents with the genome results in a plethora of lesions, commonly referred to as adducts. Adducts may cause DNA to mutate, they may... (Review)
Review
The reaction of DNA-damaging agents with the genome results in a plethora of lesions, commonly referred to as adducts. Adducts may cause DNA to mutate, they may represent the chemical precursors of lethal events and they can disrupt expression of genes. Determination of which adduct is responsible for each of these biological endpoints is difficult, but this task has been accomplished for some carcinogenic DNA-damaging agents. Here, we describe the respective contributions of specific DNA lesions to the biological effects of low molecular weight alkylating agents.
Topics: Alkylation; DNA; DNA Damage; DNA Repair; Mutagenesis
PubMed: 19875697
DOI: 10.1093/carcin/bgp262 -
Toxicology and Applied Pharmacology Jan 2016Uranium has radiological and non-radiological effects within biological systems and there is increasing evidence for genotoxic and carcinogenic properties attributable...
Uranium has radiological and non-radiological effects within biological systems and there is increasing evidence for genotoxic and carcinogenic properties attributable to uranium through its heavy metal properties. In this study, we report that low concentrations of uranium (as uranyl acetate; <10 μM) is not cytotoxic to human embryonic kidney cells or normal human keratinocytes; however, uranium exacerbates DNA damage and cytotoxicity induced by hydrogen peroxide, suggesting that uranium may inhibit DNA repair processes. Concentrations of uranyl acetate in the low micromolar range inhibited the zinc finger DNA repair protein poly(ADP-ribose) polymerase (PARP)-1 and caused zinc loss from PARP-1 protein. Uranyl acetate exposure also led to zinc loss from the zinc finger DNA repair proteins Xeroderma Pigmentosum, Complementation Group A (XPA) and aprataxin (APTX). In keeping with the observed inhibition of zinc finger function of DNA repair proteins, exposure to uranyl acetate enhanced retention of induced DNA damage. Co-incubation of uranyl acetate with zinc largely overcame the impact of uranium on PARP-1 activity and DNA damage. These findings present evidence that low concentrations of uranium can inhibit DNA repair through disruption of zinc finger domains of specific target DNA repair proteins. This may provide a mechanistic basis to account for the published observations that uranium exposure is associated with DNA repair deficiency in exposed human populations.
Topics: Cell Survival; DNA Repair; Dose-Response Relationship, Drug; HEK293 Cells; Humans; Poly (ADP-Ribose) Polymerase-1; Poly(ADP-ribose) Polymerase Inhibitors; Poly(ADP-ribose) Polymerases; Uranium
PubMed: 26627003
DOI: 10.1016/j.taap.2015.11.017 -
Antioxidants & Redox Signaling May 2011Stroke is a common cause of death and serious long-term adult disability. Oxidative DNA damage is a severe consequence of oxidative stress associated with ischemic... (Review)
Review
Stroke is a common cause of death and serious long-term adult disability. Oxidative DNA damage is a severe consequence of oxidative stress associated with ischemic stroke. The accumulation of DNA lesions, including oxidative base modifications and strand breaks, triggers cell death in neurons and other vulnerable cell populations in the ischemic brain. DNA repair systems, particularly base excision repair, are endogenous defense mechanisms that combat oxidative DNA damage. The capacity for DNA repair may affect the susceptibility of neurons to ischemic stress and influence the pathological outcome of stroke. This article reviews the accumulated understanding of molecular pathways by which oxidative DNA damage is triggered and repaired in ischemic cells, and the potential impact of these pathways on ischemic neuronal cell death/survival. Genetic or pharmacological strategies that target the signaling molecules in DNA repair responses are promising for potential clinically effective treatment. Further understanding of mechanisms for oxidative DNA damage and its repair processes may lead to new avenues for stroke management.
Topics: Animals; Brain Ischemia; DNA Damage; DNA Repair; Humans; Models, Biological; Signal Transduction
PubMed: 20677909
DOI: 10.1089/ars.2010.3451