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International Journal of Molecular... Nov 2017DNA Topoisomerase IIα (Topo IIα) is a ubiquitous enzyme in eukaryotes that performs the strand passage reaction where a double helix of DNA is passed through a second... (Review)
Review
DNA Topoisomerase IIα (Topo IIα) is a ubiquitous enzyme in eukaryotes that performs the strand passage reaction where a double helix of DNA is passed through a second double helix. This unique reaction is critical for numerous cellular processes. However, the enzyme also possesses a C-terminal domain (CTD) that is largely dispensable for the strand passage reaction but is nevertheless important for the fidelity of cell division. Recent studies have expanded our understanding of the roles of the Topo IIα CTD, in particular in mitotic mechanisms where the CTD is modified by Small Ubiquitin-like Modifier (SUMO), which in turn provides binding sites for key regulators of mitosis.
Topics: Adaptor Proteins, Signal Transducing; Aurora Kinase B; Catalytic Domain; Centromere; Chromatin; Cysteine Endopeptidases; DNA Topoisomerases, Type II; DNA-Binding Proteins; Humans; Intracellular Signaling Peptides and Proteins; Mitosis; Protein Serine-Threonine Kinases; Saccharomyces cerevisiae
PubMed: 29149026
DOI: 10.3390/ijms18112438 -
Molecular Cell Jul 2022Zhao et al. (2022) demonstrate that HIV Tat-specific factor 1, an RPA PARylation reader, recruits Topoisomerase IIβ-binding protein 1 to double-strand break sites...
Zhao et al. (2022) demonstrate that HIV Tat-specific factor 1, an RPA PARylation reader, recruits Topoisomerase IIβ-binding protein 1 to double-strand break sites specifically in the S phase of the cell cycle to promote homologous recombination.
Topics: DNA Breaks, Double-Stranded; DNA Repair; DNA Topoisomerases, Type II; Homologous Recombination; Poly ADP Ribosylation; S Phase
PubMed: 35868254
DOI: 10.1016/j.molcel.2022.06.038 -
Cell Death & Disease Jan 2024DNA topoisomerase II (TOP2) is an enzyme that performs a critical function in manipulating DNA topology during replication, transcription, and chromosomal compaction by...
DNA topoisomerase II (TOP2) is an enzyme that performs a critical function in manipulating DNA topology during replication, transcription, and chromosomal compaction by forming a vital intermediate known as the TOP2-DNA cleavage complex (TOP2cc). Although the TOP2cc is often transient, stabilization can be achieved by TOP2 poisons, a family of anti-cancer chemotherapeutic agents targeting TOP2, such as etoposide (VP-16), and then induce double-strand breaks (DSBs) in cellular DNA. TOP2cc first needs to be proteolyzed before it can be processed by TDP2 for the removal of these protein adducts and to produce clean DNA ends necessary for proper repair. However, the mechanism by which TOP2βcc is proteolyzed has not been thoroughly studied. In this study, we report that after exposure to VP-16, MDM2, a RING-type E3 ubiquitin ligase, attaches to TOP2β and initiates polyubiquitination and proteasomal degradation. Mechanistically, during exposure to VP-16, TOP2β binds to DNA to form TOP2βcc, which promotes MDM2 binding and subsequent TOP2β ubiquitination and degradation, and results in a decrease in TOP2βcc levels. Biologically, MDM2 inactivation abrogates TOP2β degradation, stabilizes TOP2βcc, and subsequently increases the number of TOP2β-concealed DSBs, resulting in the rapid death of cancer cells via the apoptotic process. Furthermore, we demonstrate the combination activity of VP-16 and RG7112, an MDM2 inhibitor, in the xenograft tumor model and in situ lung cancer mouse model. Taken together, the results of our research reveal an underlying mechanism by which MDM2 promotes cancer cell survival in the presence of TOP2 poisons by activating proteolysis of TOP2βcc in a p53-independent manner, and provides a rationale for the combination of MDM2 inhibitors with TOP2 poisons for cancer therapy.
Topics: Animals; Humans; Mice; Disease Models, Animal; DNA; DNA Topoisomerases, Type II; DNA-Binding Proteins; Etoposide; Phosphoric Diester Hydrolases; Proteolysis; Proto-Oncogene Proteins c-mdm2; Tumor Suppressor Protein p53
PubMed: 38263255
DOI: 10.1038/s41419-024-06474-3 -
Nucleic Acids Research Jan 2018In vivo DNA molecules are narrowly folded within chromatin fibers and self-interacting chromatin domains. Therefore, intra-molecular DNA entanglements (knots) might...
In vivo DNA molecules are narrowly folded within chromatin fibers and self-interacting chromatin domains. Therefore, intra-molecular DNA entanglements (knots) might occur via DNA strand passage activity of topoisomerase II. Here, we assessed the presence of such DNA knots in a variety of yeast circular minichromosomes. We found that small steady state fractions of DNA knots are common in intracellular chromatin. These knots occur irrespective of DNA replication and cell proliferation, though their abundance is reduced during DNA transcription. We found also that in vivo DNA knotting probability does not scale proportionately with chromatin length: it reaches a value of ∼0.025 in domains of ∼20 nucleosomes but tends to level off in longer chromatin fibers. These figures suggest that, while high flexibility of nucleosomal fibers and clustering of nearby nucleosomes facilitate DNA knotting locally, some mechanism minimizes the scaling of DNA knot formation throughout intracellular chromatin. We postulate that regulation of topoisomerase II activity and the fractal architecture of chromatin might be crucial to prevent a potentially massive and harmful self-entanglement of DNA molecules in vivo.
Topics: Cell Division; Chromatin; DNA Replication; DNA Topoisomerases, Type II; DNA, Fungal; DNA, Superhelical; Models, Molecular; Nucleic Acid Conformation; Protein Binding; Saccharomyces cerevisiae
PubMed: 29149297
DOI: 10.1093/nar/gkx1137 -
Chemical Research in Toxicology Apr 20211,2-Naphthoquinone, a secondary metabolite of naphthalene, is an environmental pollutant found in diesel exhaust particles that displays cytotoxic and genotoxic...
1,2-Naphthoquinone, a secondary metabolite of naphthalene, is an environmental pollutant found in diesel exhaust particles that displays cytotoxic and genotoxic properties. Because many quinones have been shown to act as topoisomerase II poisons, the effects of this compound on DNA cleavage mediated by human topoisomerase IIα and IIβ were examined. The compound increased the levels of double-stranded DNA breaks generated by both enzyme isoforms and did so better than a series of naphthoquinone derivatives. Furthermore, 1,2-naphthoquinone was a more efficacious poison against topoisomerase IIα than IIβ. Topoisomerase II poisons can be classified as interfacial (which interact noncovalently at the enzyme-DNA interface and increase DNA cleavage by blocking ligation) or covalent (which adduct the protein and increase DNA cleavage by closing the N-terminal gate of the enzyme). Therefore, experiments were performed to determine the mechanistic basis for the actions of 1,2-naphthoquinone. In contrast to results with etoposide (an interfacial poison), the activity of 1,2-naphthoquinone against topoisomerase IIα was abrogated in the presence of sulfhydryl and reducing agents. Moreover, the compound inhibited cleavage activity when incubated with the enzyme prior to the addition of DNA and induced virtually no cleavage with the catalytic core of the enzyme. It also induced stable covalent topoisomerase IIα-DNA cleavage complexes and was a partial inhibitor of DNA ligation. Findings were also consistent with 1,2-naphthoquinone acting as a covalent poison of topoisomerase IIβ; however, mechanistic studies with this isoform were less conclusive. Whereas the activity of 1,2-naphthoquinone was blocked in the presence of a sulfhydryl reagent, it was much less sensitive to the presence of a reducing agent. Furthermore, the reduced form of 1,2-naphthoquinone, 1,2-dihydroxynaphthalene, displayed high activity against the β isoform. Taken together, results suggest that 1,2-naphthoquinone increases topoisomerase II-mediated double-stranded DNA scission (at least in part) by acting as a covalent poison of the human type II enzymes.
Topics: DNA Cleavage; DNA Topoisomerases, Type II; Humans; Molecular Structure; Naphthoquinones; Recombinant Proteins; Topoisomerase II Inhibitors
PubMed: 33760604
DOI: 10.1021/acs.chemrestox.0c00492 -
Archives of Toxicology Jul 2020Sertraline, an antidepressant, is commonly used to manage mental health symptoms related to depression, anxiety disorders, and obsessive-compulsive disorder. The use of...
Sertraline, an antidepressant, is commonly used to manage mental health symptoms related to depression, anxiety disorders, and obsessive-compulsive disorder. The use of sertraline has been associated with rare but severe hepatotoxicity. Previous research demonstrated that mitochondrial dysfunction, apoptosis, and endoplasmic reticulum stress were involved in sertraline-associated cytotoxicity. In this study, we reported that after a 24-h treatment in HepG2 cells, sertraline caused cytotoxicity, suppressed topoisomerase I and IIα, and damaged DNA in a concentration-dependent manner. We also investigated the role of cytochrome P450 (CYP)-mediated metabolism in sertraline-induced toxicity using our previously established HepG2 cell lines individually expressing 14 CYPs (1A1, 1A2, 1B1, 2A6, 2B6, 2C8, 2C9, 2C18, 2C19, 2D6, 2E1, 3A4, 3A5, and 3A7). We demonstrated that CYP2D6, 2C19, 2B6, and 2C9 metabolize sertraline, and sertraline-induced cytotoxicity was significantly decreased in the cells expressing these CYPs. Western blot analysis demonstrated that the induction of ɣH2A.X (a hallmark of DNA damage) and topoisomerase inhibition were partially reversed in CYP2D6-, 2C19-, 2B6-, and 2C9-overexpressing HepG2 cells. These data indicate that DNA damage and topoisomerase inhibition are involved in sertraline-induced cytotoxicity and that CYPs-mediated metabolism plays a role in decreasing the toxicity of sertraline.
Topics: Antidepressive Agents; Chemical and Drug Induced Liver Injury; Cytochrome P-450 Enzyme System; DNA Damage; DNA Topoisomerases, Type I; DNA Topoisomerases, Type II; G2 Phase Cell Cycle Checkpoints; Hep G2 Cells; Hepatocytes; Humans; Isoenzymes; Liver; Metabolic Detoxication, Phase I; Poly-ADP-Ribose Binding Proteins; Selective Serotonin Reuptake Inhibitors; Sertraline
PubMed: 32372212
DOI: 10.1007/s00204-020-02753-y -
Genes Oct 2019DNA topoisomerase II (TOP2) plays a critical role in many processes such as replication and transcription, where it resolves DNA structures and relieves torsional... (Review)
Review
DNA topoisomerase II (TOP2) plays a critical role in many processes such as replication and transcription, where it resolves DNA structures and relieves torsional stress. Recent evidence demonstrated the association of TOP2 with topologically associated domains (TAD) boundaries and CCCTC-binding factor (CTCF) binding sites. At these sites, TOP2 promotes interactions between enhancers and gene promoters, and relieves torsional stress that accumulates at these physical barriers. Interestingly, in executing its enzymatic function, TOP2 contributes to DNA fragility through re-ligation failure, which results in persistent DNA breaks when unrepaired or illegitimately repaired. Here, we discuss the biological processes for which TOP2 is required and the steps at which it can introduce DNA breaks. We describe the repair processes that follow removal of TOP2 adducts and the resultant broken DNA ends, and present how these processes can contribute to disease-associated mutations. Furthermore, we examine the involvement of TOP2-induced breaks in the formation of oncogenic translocations of leukemia and papillary thyroid cancer, as well as the role of TOP2 and proteins which repair TOP2 adducts in other diseases. The participation of TOP2 in generating persistent DNA breaks and leading to diseases such as cancer, could have an impact on disease treatment and prevention.
Topics: CCCTC-Binding Factor; Chromatin; DNA; DNA Breaks, Double-Stranded; DNA Repair; DNA Topoisomerases, Type II; Humans; Leukemia, Myeloid, Acute; Thyroid Cancer, Papillary; Topoisomerase II Inhibitors; Torsion, Mechanical
PubMed: 31614754
DOI: 10.3390/genes10100791 -
International Journal of Molecular... May 2023DNA topoisomerases are important enzymes that stabilize DNA supercoiling and resolve entanglements. There are two main types of topoisomerases in all cells: type I,... (Review)
Review
DNA topoisomerases are important enzymes that stabilize DNA supercoiling and resolve entanglements. There are two main types of topoisomerases in all cells: type I, which causes single-stranded DNA breaks, and type II, which cuts double-stranded DNA. Topoisomerase activity is particularly increased in rapidly dividing cells, such as cancer cells. Topoisomerase inhibitors have been an effective chemotherapeutic option for the treatment of several cancers. In addition, combination cancer therapy with topoisomerase inhibitors may increase therapeutic efficacy and decrease resistance or side effects. Topoisomerase inhibitors are currently being used worldwide, including in the United States, and clinical trials on the combination of topoisomerase inhibitors with other drugs are currently underway. The primary objective of this review was to comprehensively analyze the current clinical landscape concerning the combined application of irinotecan, an extensively investigated type I topoisomerase inhibitor for colorectal cancer, and doxorubicin, an extensively researched type II topoisomerase inhibitor for breast cancer, while presenting a novel approach for cancer therapy.
Topics: Humans; Female; Topoisomerase I Inhibitors; Breast Neoplasms; Topoisomerase II Inhibitors; Drug Therapy, Combination; Colorectal Neoplasms; DNA Topoisomerases, Type II; DNA Topoisomerases, Type I
PubMed: 37176164
DOI: 10.3390/ijms24098457 -
Proceedings of the National Academy of... Jan 2022Topoisomerases nick and reseal DNA to relieve torsional stress associated with transcription and replication and to resolve structures such as knots and catenanes....
Topoisomerases nick and reseal DNA to relieve torsional stress associated with transcription and replication and to resolve structures such as knots and catenanes. Stabilization of the yeast Top2 cleavage intermediates is mutagenic in yeast, but whether this extends to higher eukaryotes is less clear. Chemotherapeutic topoisomerase poisons also elevate cleavage, resulting in mutagenesis. Here, we describe p.K743N mutations in human topoisomerase hTOP2α and link them to a previously undescribed mutator phenotype in cancer. Overexpression of the orthologous mutant protein in yeast generated a characteristic pattern of 2- to 4-base pair (bp) duplications resembling those in tumors with p.K743N. Using mutant strains and biochemical analysis, we determined the genetic requirements of this mutagenic process and showed that it results from trapping of the mutant yeast yTop2 cleavage complex. In addition to 2- to 4-bp duplications, hTOP2α p.K743N is also associated with deletions that are absent in yeast. We call the combined pattern of duplications and deletions ID_TOP2α. All seven tumors carrying the hTOP2α p.K743N mutation showed ID_TOP2α, while it was absent from all other tumors examined ( = 12,269). Each tumor with the ID_TOP2α signature had indels in several known cancer genes, which included frameshift mutations in tumor suppressors PTEN and TP53 and an activating insertion in BRAF. Sequence motifs found at ID_TOP2α mutations were present at 80% of indels in cancer-driver genes, suggesting that ID_TOP2α mutagenesis may contribute to tumorigenesis. The results reported here shed further light on the role of topoisomerase II in genome instability.
Topics: Alleles; Amino Acid Substitution; Base Sequence; Cell Survival; DNA Damage; DNA Mutational Analysis; DNA Topoisomerases, Type II; Gene Duplication; Gene Rearrangement; Genetic Predisposition to Disease; Genotype; Humans; INDEL Mutation; Mutagenesis; Mutation; Neoplasms; Oncogenes; Phenotype; Poly-ADP-Ribose Binding Proteins; Sequence Deletion
PubMed: 35058360
DOI: 10.1073/pnas.2114024119 -
Journal of Virology Nov 2023African swine fever virus (ASFV) is a highly fatal swine disease that severely affects the pig industry. Although ASFV has been prevalent for more than 100 years,...
African swine fever virus (ASFV) is a highly fatal swine disease that severely affects the pig industry. Although ASFV has been prevalent for more than 100 years, effective vaccines or antiviral strategies are still lacking. In this study, we identified four strains that inhibited ASFV proliferation . Pigs fed with liquid biologics or powders derived from four strains mixed with pellet feed showed reduced morbidity and mortality when challenged with ASFV. Further analysis showed that the antiviral activity of was based on its metabolites arctiin and genistein interfering with the function of viral topoisomerase II. Our findings offer a promising new strategy for the prevention and control of ASFV that may significantly alleviate the economic losses in the pig industry.
Topics: Animals; African Swine Fever; African Swine Fever Virus; Antiviral Agents; Bacillus subtilis; DNA Topoisomerases, Type II; Genistein; Swine
PubMed: 37929962
DOI: 10.1128/jvi.00719-23