Role Of Topoisomerase Research Articles

Genomic Insights into the Role of TOP Gene Family in Soft-Tissue Sarcomas: Implications for Prognosis and Therapy.

This study focuses on the role of topoisomerases (TOPs) in sarcomas (SARCs), highlighting TOPs' influence on sarcoma prognosis through mRNA expression, genetic mutations, immune infiltration, and DNA methylation analysis using transcriptase sequencing and other techniques. The findings indicate that TOP gene mutations correlate with increased inflammation, immune cell infiltration, DNA repair abnormalities, and mitochondrial fusion genes alterations, all of which negatively affect sarcoma prognosis. Abnormal TOP expression may independently affect sarcoma patients' survival. Cutting-edge genomic tools such as Oncomine, gene expression profiling interactive analysis (GEPIA), and cBio Cancer Genomics Portal (cBioPortal) are utilizedto explore the TOP gene family (TOP1/1MT/2A/2B/3A/3B) in soft-tissue sarcomas (STSs). This in-depth analysis reveals a notable upregulation of TOP mRNA in STS patients arcoss various SARC subtypes, French Federation Nationale des Centres de Lutte Contre le Cancer classification (FNCLCC) grades, and specific molecular profiles correlating with poorer clinical outcomes. Furthermore, this investigation identifies distinct patterns of immune cell infiltration, genetic mutations, and somatic copy number variations linked to TOP genes that inversely affect patient survival rates. These findings underscore the diagnostic and therapeutic relevance of the TOP gene suite in STSs.

Expanding the type IIB DNA topoisomerase family: identification of new topoisomerase and topoisomerase-like proteins in mobile genetic elements.

The control of DNA topology by DNA topoisomerases is essential for virtually all DNA transactions in the cell. These enzymes, present in every organism, exist as several non-homologous families. We previously identified a small group of atypical type IIB topoisomerases, called Topo VIII, mainly encoded by plasmids. Here, taking advantage of the rapid expansion of sequence databases, we identified new putative Topo VIII homologs. Our analyses confirm the exclusivity of the corresponding genes to mobile genetic elements (MGE) and extend their distribution to nine different bacterial phyla and one archaeal superphylum. Notably, we discovered another subfamily of topoisomerases, dubbed ‘Mini-A’, including distant homologs of type IIB topoisomerases and encoded by extrachromosomal and integrated bacterial and archaeal viruses. Interestingly, a short, functionally uncharacterized motif at the C-terminal extremity of type IIB topoisomerases appears sufficient to discriminate between Mini-A, Topo VI and Topo VIII subfamilies. This motif could be a key element for understanding the differences between the three subfamilies. Collectively, this work leads to an updated model for the origin and evolution of the type IIB topoisomerase family and raises questions regarding the role of topoisomerases during replication of MGE in bacteria and archaea.

Roles of Topoisomerases in Heterochromatin, Aging, and Diseases.

Heterochromatin is a transcriptionally repressive chromatin architecture that has a low abundance of genes but an enrichment of transposons. Defects in heterochromatin can cause the de-repression of genes and transposons, leading to deleterious physiological changes such as aging, cancer, and neurological disorders. While the roles of topoisomerases in many DNA-based processes have been investigated and reviewed, their roles in heterochromatin formation and function are only beginning to be understood. In this review, we discuss recent findings on how topoisomerases can promote heterochromatin organization and impact the transcription of genes and transposons. We will focus on two topoisomerases: Top2α, which catenates and decatenates double-stranded DNA, and Top3β, which can change the topology of not only DNA, but also RNA. Both enzymes are required for normal heterochromatin formation and function, as the inactivation of either protein by genetic mutations or chemical inhibitors can result in defective heterochromatin formation and the de-silencing of transposons. These defects may contribute to the shortened lifespan and neurological disorders observed in individuals carrying mutations of Top3β. We propose that topological stress may be generated in both DNA and RNA during heterochromatin formation and function, which depend on multiple topoisomerases to resolve.

Effects of an unusual poison identify a lifespan role for Topoisomerase 2 in Saccharomyces cerevisiae.

A progressive loss of genome maintenance has been implicated as both a cause and consequence of aging. Here we present evidence supporting the hypothesis that an age-associated decay in genome maintenance promotes aging in Saccharomyces cerevisiae (yeast) due to an inability to sense or repair DNA damage by topoisomerase 2 (yTop2). We describe the characterization of LS1, identified in a high throughput screen for small molecules that shorten the replicative lifespan of yeast. LS1 accelerates aging without affecting proliferative growth or viability. Genetic and biochemical criteria reveal LS1 to be a weak Top2 poison. Top2 poisons induce the accumulation of covalent Top2-linked DNA double strand breaks that, if left unrepaired, lead to genome instability and death. LS1 is toxic to cells deficient in homologous recombination, suggesting that the damage it induces is normally mitigated by genome maintenance systems. The essential roles of yTop2 in proliferating cells may come with a fitness trade-off in older cells that are less able to sense or repair yTop2-mediated DNA damage. Consistent with this idea, cells live longer when yTop2 expression levels are reduced. These results identify intrinsic yTop2-mediated DNA damage as a potentially manageable cause of aging.

The role of topoisomerase I in suppressing genome instability associated with a highly transcribed guanine-rich sequence is not restricted to preventing RNA:DNA hybrid accumulation.

Highly transcribed guanine-run containing sequences, in Saccharomyces cerevisiae, become unstable when topoisomerase I (Top1) is disrupted. Topological changes, such as the formation of extended RNA:DNA hybrids or R-loops or non-canonical DNA structures including G-quadruplexes has been proposed as the major underlying cause of the transcription-linked genome instability. Here, we report that R-loop accumulation at a guanine-rich sequence, which is capable of assembling into the four-stranded G4 DNA structure, is dependent on the level and the orientation of transcription. In the absence of Top1 or RNase Hs, R-loops accumulated to substantially higher extent when guanine-runs were located on the non-transcribed strand. This coincides with the orientation where higher genome instability was observed. However, we further report that there are significant differences between the disruption of RNase Hs and Top1 in regards to the orientation-specific elevation in genome instability at the guanine-rich sequence. Additionally, genome instability in Top1-deficient yeasts is not completely suppressed by removal of negative supercoils and further aggravated by expression of mutant Top1. Together, our data provide a strong support for a function of Top1 in suppressing genome instability at the guanine-run containing sequence that goes beyond preventing the transcription-associated RNA:DNA hybrid formation.

Role of topoisomerase I and thymidylate synthase expression in sporadic colorectal cancer: Associations with clinicopathological and molecular features

Topoisomerase I (Topo I) and thymidylate synthase (TS) are essential enzymes for the replication, transcription and repair of DNA, and are potential biomarkers in colorectal cancer (CRC). The aim of the study was to correlate the tissue expression of Topo I and TS in sporadic CRCs with relevant pathological and molecular features and patients’ outcome.Topo I and TS expression was assessed by immunostaining in 112 consecutive primary CRCs. Increased expression of Topo I was found in 36% of tumors, preferentially rectal (50%) and with not otherwise specified (NOS) histology (44%). Topo I expression was associated with 18q allelic loss (LOH), (p=0.013), microsatellite stable phenotype (p=0.002) and normal expression of mismatch proteins hMLH1 and hMSH2 (p=0.0012 and p=0.02, respectively). High TS expression was found in 60% of tumors, more frequently in distal sites (62%) and with NOS histology (66%); no association with microsatellite instability was observed.Topo I seems to be involved in the chromosomal instability pathway of sporadic CRCs. Conversely, high TS expression is unlikely to affect the clinical behavior of microsatellite unstable CRCs.

Role of Topoisomerases in Pediatric High Grade Osteosarcomas: TOP2A Gene Is One of the Unique Molecular Biomarkers of Chemoresponse

Currently, the treatment of pediatric high-grade osteosarcomas systematically includes one topoisomerase IIα inhibitor. This chemotherapy is usually adapted to the response to the neo-adjuvant therapy after surgery. The current and unique marker of chemoresponsiveness is the percentage of viable residual cells in the surgical resection. This late patient management marker has to be evaluated earlier in the therapeutic history of the patients on initial biopsy. Therefore, new biomarkers, especially those involved in the topoisomerase IIα inhibitor response might be good candidates. Therefore, our study was designed to target TOP1, TOP2A and TOP2B genes in 105 fresh-frozen diagnostic biopsies by allelotyping and real-time quantitative PCR. Our analyses in those pediatric osteosarcomas, homogeneously treated, highlighted the frequent involvement of topo-isomerase genes. The main and most important observation was the statistical link between the presence of TOP2A amplification and the good response to neo-adjuvant chemotherapy. Compared to adult cancers, the 17q21 amplicon, including TOP2A and ERBB2 genes, seems to be differentially implicated in the osteosarcoma chemoresponsiveness. Surprisingly, there is no ERBB2 gene co-amplification and the patients harboring TOP2A amplification tend to show a worse survival, so TOP2A analyses remain a preliminary, but a good molecular approach for the evaluation at diagnosis of pediatric osteosarcoma chemoresponsiveness.

ERK1/2 signaling plays an important role in topoisomerase II poison-induced G2/M checkpoint activation.

Topo II poisons, which target topoisomerase II (topo II) to generate enzyme mediated DNA damage, have been commonly used for anti-cancer treatment. While clinical evidence demonstrate a capability of topo II poisons in inducing apoptosis in cancer cells, accumulating evidence also show that topo II poison treatment frequently results in cell cycle arrest in cancer cells, which was associated with subsequent resistance to these treatments. Results in this report indicate that treatment of MCF-7 and T47D breast cancer cells with topo II poisons resulted in an increased phosphorylation of extracellular signal-regulated kinase 1 and 2 (ERK1/2) and an subsequent induction of G2/M cell cycle arrest. Furthermore, inhibition of ERK1/2 activation using specific inhibitors markedly attenuated the topo II poison-induced G2/M arrest and diminished the topo II poison-induced activation of ATR and Chk1 kinases. Moreover, decreased expression of ATR by specific shRNA diminished topo II poison-induced G2/M arrest but had no effect on topo II poison-induced ERK1/2 activation. In contrast, inhibition of ERK1/2 signaling had little, if any, effect on topo II poison-induced ATM activation. In addition, ATM inhibition by either incubation of cells with ATM specific inhibitor or transfection of cells with ATM specific siRNA did not block topo II poison-induced G2/M arrest. Ultimately, inhibition of ERK1/2 signaling greatly enhanced topo II poison-induced apoptosis. These results implicate a critical role for ERK1/2 signaling in the activation of G2/M checkpoint response following topo II poison treatment, which protects cells from topo II poison-induced apoptosis.

Topoisomerase expression in oral squamous cell carcinoma: relationship with cancer stem cells profiles and lymph node metastasis

The relationship between predictive proteins and tumors presenting cancer stem cells (CSCs) profiles in oral tumors is still poorly understood. This study aims to identify the relationship between topoisomerases I, IIα, and IIIα and putative CSCs immunophenotype in oral squamous cell carcinoma (OSCC) and determine its influence on prognosis. The following data were retrieved from 127 patients: age, gender, primary anatomic site, smoking and alcohol intake, recurrence, metastases, histologic classification, treatment, and survival. An immunohistochemical study for topoisomerases I, IIα, and IIIα was performed in a tissue microarray containing 127 paraffin blocks of OSCCs. In univariate analysis, topoisomerases expression showed significant differences according to CSCs profiles and p53 immunoexpression, but not with survival. Topoisomerases IIα and IIIα also showed significant relationship with lymph node metastasis. The multivariate test confirmed these associations. The results that all topoisomerases correlates with OSCC CSCs may indicate a role for topoisomerases in head and neck carcinogenesis. Notwithstanding, it is plausible that other members of topoisomerases family could represent novel therapeutical targets in oral squamous cell carcinoma.

Target Identification and Intervention Strategies against Kinetoplastid Protozoan Parasites

The past few decades have been marked by numerous admirable research efforts and promising technological advancements in the field of research on protozoan parasites. The parasites of this genre cause some devastating diseases that pose alarming threat to the mankind. Though several intervention strategies have been developed to get rid of these parasites, they always seem to frustrate the efforts of the scientific community sooner or later. The intervention strategies include identification of novel drug targets, development of target-based therapy, and development of vaccines. that provide significant impetus in the field of research pertaining to these parasites. In this context, several reviews have appeared in the past few years elucidating different drug targets in these parasites. For example, Das et al. [1], Balana-Fouce et al. [2], and others have described the role of topoisomerases as potential drug targets in these kinetoplastid protozoa. Urbina [3] has described the lipid biosynthetic pathway as a possible chemotherapeutic target whereas McConville [4] has elucidated the potential of parasite surface glycoconjugates as possible drug targets. Other targets include cysteine peptidases [5] and histone deacetylases [6] of the trypanosomatid parasites.

Role for topoisomerase 1 in transcription-associated mutagenesis in yeast

High levels of transcription in Saccharomyces cerevisiae are associated with increased genetic instability, which has been linked to DNA damage. Here, we describe a pGAL-CAN1 forward mutation assay for studying transcription-associated mutagenesis (TAM) in yeast. In a wild-type background with no alterations in DNA repair capacity, ≈50% of forward mutations that arise in the CAN1 gene under high-transcription conditions are deletions of 2-5 bp. Furthermore, the deletions characteristic of TAM localize to discrete hotspots that coincide with 2-4 copies of a tandem repeat. Although the signature deletions of TAM are not affected by the loss of error-free or error-prone lesion bypass pathways, they are completely eliminated by deletion of the TOP1 gene, which encodes the yeast type IB topoisomerase. Hotspots can be transposed into the context of a frameshift reversion assay, which is sensitive enough to detect Top1-dependent deletions even in the absence of high transcription. We suggest that the accumulation of Top1 cleavage complexes is related to the level of transcription and that their removal leads to the signature deletions. Given the high degree of conservation between DNA metabolic processes, the links established here among transcription, Top1, and mutagenesis are likely to extend beyond the yeast system.

On the role of topoisomerase I in mediating the cytotoxicity of 9-aminoacridine-based anticancer agents

The cytotoxicity and mechanism of action of a series of substituted 9-aminoacridines is evaluated using topoisomerase I and cancer cell growth inhibition assays. In previous work, compounds of this type were shown to catalytically inhibit topoisomerase II, leading to a G1-S phase arrest of the cell cycle and apoptosis in pancreatic cancer cells in vitro and in vivo. The present study expands the potential utility of these compounds in the development of cancer therapeutics by showing that these compounds inhibit proliferation of cell lines derived from the nine most common human cancers. Further results show that at least one of the compounds effectively stabilizes topoisomerase I–DNA adduct formation in intact cells. RNA interference experiments, however, indicate that this interaction does not contribute to the drug-induced killing of cancer cells indicating the compounds may be non-lethal poisons of topoisomerase I.

A New Role for a Long-Studied DNA-Wrangling Enzyme

A New Role for a Long-Studied DNA-Wrangling Enzyme

DNA Topoisomerase I Inhibitory Alkaloids from Corydalis saxicola

Chemical studies of the Chinese herb Corydalis saxicola Bunting led to the isolation and identification of 14 alkaloids, 1-14. Seven of these compounds, 4-9 and 11, were obtained from this plant for the first time. Feruloylagmatine (7) is the first guanidine-type alkaloid to be identified in the family Papaveraceae and in dicotyledonous plants. All of the isolated compounds were assayed for inhibitory activity against human DNA topoisomerase I. A DNA cleavage assay demonstrated that these alkaloids specifically inhibit topoisomerase through stabilization of the enzyme-DNA complex. Among the isolated alkaloids, (-)-pallidine (8) and (-)-scoulerine (11) showed strong inhibitory activities toward topoisomerase I that were comparable to camptothecin, a typical topoisomerase I inhibitor. A preliminary structure-activity relationship study suggested that the quaternary ammonium ion might play an important role in topoisomerase I inhibition by the isoquinoline alkaloids. These data indicated that DNA topoisomerase I inhibition represents probably one of the anticarcinogenic mechanisms of C. saxicola.

Cells Lacking DNA Topoisomerase IIβ are Resistant to Genistein

Evidence suggests that DNA topoisomerases (topos) may be involved in the anticancer and carcinogenic properties attributed to flavonoids. Using the cell-based assay TARDIS, the dietary flavonoids genistein (1) and luteolin (2) have been evaluated as topo I and topo II poisons and catalytic inhibitors in K562 leukemia cells. Both flavonoids induced topo II-DNA complexes, but they did not induce significant levels of topo I-DNA complexes. Genistein decreased the topo II-DNA complexes induced by the topo II poison etoposide, suggestive of a catalytic inhibition of topo II, and luteolin decreased the topo I-DNA complexes induced by the topo I poison camptothecin, indicative of a catalytic inhibition of topo I. Murine transgenic cells lacking topo II beta were resistant to genistein-induced cell growth inhibition (XTT assays) and cytotoxicity (clonogenic assay). High levels of topo II beta-DNA complexes were also observed in K562 cells exposed to genistein. These data suggest that topo II beta has an important function in genistein-induced cell growth inhibition and cell death. The possible role of topoisomerases in the putative anticancer and carcinogenic properties of genistein and luteolin is discussed.

302 POSTER Processing of 1-nitroacridine-induced DNA-DNA cross-links by topoisomerase I is associated with enhanced cellular survival: a possible role of topoisomerase I in the removal of DNA cross-links

302 POSTER Processing of 1-nitroacridine-induced DNA-DNA cross-links by topoisomerase I is associated with enhanced cellular survival: a possible role of topoisomerase I in the removal of DNA cross-links

Topoisomerase II beta levels are a determinant of melphalan-induced DNA crosslinks and sensitivity to cell death

The role of topoisomerase (topo) II in DNA repair has yet to be fully elucidated. Current evidence suggesting a role for topo II in the repair of DNA damage has been obtained by using in vitro model systems or inferred from correlative data in drug resistant cell lines. In this study we directly examined the role of topo IIα and β in mediating the repair of melphalan-induced crosslinks in cellular DNA. To accomplish this, we used siRNA technology to knock down either topo IIα or β in human chronic myelogenous leukemia K562 and histiocytic lymphoma U937 cell line. Our data demonstrate that topo IIβ levels, (but not α), are a determinant of melphalan-induced crosslinks and sensitivity to melphalan. Furthermore, we show that knocking down topo IIβ inhibits the repair of melphalan-induced crosslinks in K562 cells. These studies represent the first direct evidence that topo IIβ participates in the repair of DNA damage induced by an alkylating agent in cellular DNA. Finally, these results suggest non-redundant roles for these two isoforms in mediating repair of DNA crosslinks.

507 Role of topoisomerase I inhibition in the cytotoxic action of synthetic derivatives of the anticancer marine alkaloid lamellarin D

507 Role of topoisomerase I inhibition in the cytotoxic action of synthetic derivatives of the anticancer marine alkaloid lamellarin D

Plant DNA Topoisomerases: Structure, Function, and Cellular Roles in Plant Development

Regulation of the topological state of DNA is critical for cellular viability. DNA topoisomerases alter DNA topology and participate in nearly all events related to DNA metabolism such as replication, transcription, recombination, and chromosome segregation. Despite significant progress in our understanding of the structure, mechanism, and the role of topoisomerases in diverse cellular processes in bacteria, yeast, and animal systems, the roles of plant topoisomerases are poorly understood. In recent years expression studies, characterization of recombinant topoisomerases, and gene knockout studies have highlighted the importance of topoisomerases in plant DNA replication, mitosis, cell proliferation, organelle division, meiotic recombination, morphogenesis, and plant development. We have focused this review specifically on plant topoisomerases, with an emphasis on their biological role in plant development.

Transcription from the perspective of the DNA: twists and bumps in the road.

Although nature's design of the DNA double helix is ingenious for the storage of information, this design creates topological problems for the processes that occur on the DNA. Cellular processes, such as transcription, replication, chromosomal segregation, and chromosomal condensation, are all complicated by the double helix. The problem is compounded in cells since the DNA is in chromatin. Topoisomerases relax positive and negative superhelical turns in DNA, and thereby topoisomerases have long been recognized as key components of the DNA replication and chromosome segregation and condensation machinery. A role for topoisomerases in the transcription process has also been noted in living cells, but only recently has such a role been recapitulated in the test tube for transcription reactions. New data are discussed that demonstrate that for in vitro transcription reactions, topoisomerases are dispensable when the template is naked DNA, but when the template is reconstituted into chromatin, topoisomerases are required for transcription to proceed efficiently.