Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:5.99.1.3 (topoisomerase)
9,911 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The Saccharomyces cerevisiae SGS1 gene is homologous to Escherichia coli RecQ and the human BLM and WRN proteins that are defective in the cancer-prone disorder Bloom's syndrome and the premature aging disorder Werner's syndrome, respectively. While recQ mutants are deficient in conjugational recombination and DNA repair, Bloom's syndrome cell lines show hyperrecombination. Bloom's and Werner's syndrome cell lines both exhibit chromosomal instability, sgs1 delta strains show mitotic hyperrecombination, as do Bloom's cells. This was manifested as an increase in the frequency of interchromosomal homologous recombination, intrachromosomal excision recombination, and ectopic recombination. Hyperrecombination was partially independent of both RAD52 and RAD1. Meiotic recombination was not increased in sgs1 delta mutants, although meiosis I chromosome missegregation has been shown to be elevated sgs1 delta suppresses the slow growth of a top3 delta strain lacking topoisomerase III. Although there was an increase in subtelomeric Y' instability in sgs1 delta strains due to hyperrecombination, no evidence was found for an increase in the instability of terminal telomeric sequences in a top3 delta or a sgs1 delta strain. This contrasts with the telomere maintenance defects of Werner's patients. We conclude that the SGS1 gene product is involved in the maintenance of genome stability in S. cerevisiae.
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PMID:SGS1, a homologue of the Bloom's and Werner's syndrome genes, is required for maintenance of genome stability in Saccharomyces cerevisiae. 891 39

Werner syndrome (WS) is an autosomal recessive disorder characterized by genomic instability and the premature onset of a number of age-related diseases. The gene responsible for WS encodes a member of the RecQ-like subfamily of DNA helicases. Here we show that its murine homologue maps to murine chromosome 8 in a region syntenic with the human WRN gene. We have deleted a segment of this gene and created Wrn-deficient embryonic stem (ES) cells and WS mice. While displaying reduced embryonic survival, live-born WS mice otherwise appear normal during their first year of life. Nonetheless, although several DNA repair systems are apparently intact in homozygous WS ES cells, such cells display a higher mutation rate and are significantly more sensitive to topoisomerase inhibitors (especially camptothecin) than are wild-type ES cells. Furthermore, mouse embryo fibroblasts derived from homozygous WS embryos show premature loss of proliferative capacity. At the molecular level, wild-type, but not mutant, WS protein copurifies through a series of centrifugation and chromatography steps with a multiprotein DNA replication complex.
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PMID:A deletion within the murine Werner syndrome helicase induces sensitivity to inhibitors of topoisomerase and loss of cellular proliferative capacity. 978 47

Mutations in the WRN gene result in Werner syndrome, an autosomal recessive disease in which many characteristics of aging are accelerated. A probable role in some aspect of DNA metabolism is suggested by the primary sequence of the WRN gene product. A recombinant His-tagged WRN protein (WRNp) was overproduced in insect cells using the baculovirus system and purified to near homogeneity by several chromatographic steps. This purification scheme removes both nuclease and topoisomerase contaminants that persist following a single Ni(2+)affinity chromatography step and allows for unambiguous interpretation of WRNp enzymatic activities on DNA substrates. Purified WRNp has DNA-dependent ATPase and helicase activities consistent with its homology to the RecQ subfamily of proteins. The protein also binds with higher affinity to single-stranded DNA than to double-stranded DNA. However, WRNp has no higher affinity for various types of DNA damage, including adducts formed during 4NQO treatment, than for undamaged DNA. Our results confirm that WRNp has a role in DNA metabolism, although this role does not appear to be the specific recognition of damage in DNA.
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PMID:Enzymatic and DNA binding properties of purified WRN protein: high affinity binding to single-stranded DNA but not to DNA damage induced by 4NQO. 1044 47

The RecQ helicase superfamily has been implicated in DNA repair and recombination. At least five human RecQ-related genes exist: RecQ1, BLM, WRN, RecQ4 and RecQ5. Mutations in BLM, WRN and RecQ4 are associated with Bloom, Werner and Rothmund-Thomson syndromes, respectively, involving a predisposition to malignancies and a cellular phenotype that includes increased chromosome instability. RecQ5 is small, containing only a core part of the RecQ helicase, but three isomer transcripts code for small RecQ5alpha (corresponding to the original RecQ5 with 410 amino acids), new large RecQ5beta (991 amino acids) and small RecQ5gamma (435 amino acids) proteins that contain the core helicase motifs. By determining the genomic structure, we found that the three isoforms are generated by differential splicing from the RecQ5 gene that contains at least 19 exons. Northern blot analysis using a RecQ5beta-specific probe indicates that RecQ5beta mRNA is expressed strongly in the testis. Immunocytochemical staining of three N-terminally tagged RecQ5 isomers expressed in 293EBNA cells showed that RecQ5beta migrates to the nucleus and exists exclusively in the nucleoplasm, while the small RecQ5alpha and RecQ5gamma proteins stay in the cytoplasm. Immunoprecipitation and an extended cytochemical experiment suggested that the nucleoplasmic RecQ5beta, like yeast Sgs1 DNA helicase, binds to topoisomerases 3alpha and 3beta, but not to topoisomerase 1. These results predict that RecQ5beta may have an important role in DNA metabolism and may also be related to a distinct genetic disease.
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PMID:Human RecQ5beta, a large isomer of RecQ5 DNA helicase, localizes in the nucleoplasm and interacts with topoisomerases 3alpha and 3beta. 1071 Apr 32

Werner's syndrome (WS) is a rare autosomal recessive human disorder and the patients exhibit many symptoms of accelerated ageing in their early adulthood. The gene (WRN) responsible for WS has been biochemically characterised as a 3'-5' helicase and is homologous to a number of RecQ superfamily of helicases. The yeast SGS1 helicase is considered as a human WRN homologue and SGS1 physically interacts with topoisomerases II and III. In view of this, it has been hypothesised that the WRN gene may also interact with topoisomerases II and III. The purpose of this study is to determine whether the loss of function of WRN protein alters the sensitivity of WS cells to agents that block the action of topoisomerase II. This study deals with the comparison of the chromosomal damage induced by the two anti-topoisomerase II drugs, VP-16 and amsacrine, in both G1 and G2 phases of the cell cycle, in lymphoblastoid cells from WS patients and from a healthy donor. Our results show that the WS cell lines are hypersensitive to chromosome damage induced by VP-16 and amsacrine only in the G2 phase of the cell cycle. No difference either in the yield of the induced aberrations or SCEs was found after treatment of cells at G1 stage. These data might suggest that in WS cells, because of the mutation of the WRN protein, the inhibition of topoisomerase II activity results in a higher rate of misrepair, probably due to some compromised G2 phase processes involving the WRN protein.
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PMID:Werner's syndrome lymphoblastoid cells are hypersensitive to topoisomerase II inhibitors in the G2 phase of the cell cycle. 1072 63

The Saccharomyces cerevisiae gene SGS1 encodes a DNA helicase that shows homology to the Escherichia coli protein RecQ and the products of the BLM and WRN genes in humans, which are defective in Bloom's and Werner's syndrome, respectively. Recently, it has been proposed that this helicase is involved in maintaining the integrity of the rDNA and that loss of Sgs1 function leads to accelerated aging. Sgs1 has been isolated on the basis of its genetic interaction with both topoisomerase I and topoisomerase III, as well as in a two-hybrid screen for proteins that interact with the C-terminal portion of topoisomerase II. We have defined the minimal structural elements of Sgs1 required for its interactions with the three topoisomerases, and demonstrate that the complex phenotypes associated with sgs1 mutants are a consequence of a dysfunctional Sgs1-Top3 complex. We also report that the synthetic relationship between mutations in SGS1 and SRS2, which encodes another helicase implicated in recombinational repair, likewise result from a dysfunctional Sgs1-Top3 interaction. Our findings indicate that Sgs1 may act on different DNA structures depending on the activity of topoisomerase I, Srs2 and topoisomerase III.
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PMID:Genetic analysis of the Saccharomyces cerevisiae Sgs1 helicase defines an essential function for the Sgs1-Top3 complex in the absence of SRS2 or TOP1. 1101 37

Werner's syndrome (WS) is a recessive human genetic disorder associated with an elevated incidence of many types of cancer. The WS gene product, WRNp, belongs to the RecQ family of DNA helicases and is required for the maintenance of genomic stability in human cells. A possible interaction between helicases and topoisomerases that could co-operate in many aspects of DNA metabolism such as progression of the replication forks, recombination and repair has been recently suggested. In addition, sgs1 gene product in yeast, homologous to WS gene, has been shown to physically interact with topoisomerase types I and II. Earlier data from our laboratory suggested that WRN helicase might play a role in a G2 recombinational pathway of double strand breaks (DSBs) repair, co-operating with topoisomerase II. In this work, the effect of the topoisomerase I inhibitor camptothecin in WS cells has been investigated at the chromosomal level. The data from the present work suggest that the inhibition of topoisomerase I activity by camptothecin results in a higher induction of chromosomal damage in WS cell lines in the G2-phase and in the S-phase of the cell cycle compared to normal cells, perhaps associated with the defects in DNA replication synthesis.
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PMID:Werner's syndrome cell lines are hypersensitive to camptothecin-induced chromosomal damage. 1108 95

Deficiency in a helicase of the RecQ family is found in at least three human genetic disorders associated with cancer predisposition and/or premature ageing. The RecQ helicases encoded by the BLM, WRN and RECQ4 genes are defective in Bloom's, Werner's and Rothmund-Thomson syndromes, respectively. Cells derived from individuals with these disorders in each case show inherent genomic instability. Recent studies have demonstrated direct interactions between these RecQ helicases and human nuclear proteins required for several aspects of chromosome maintenance, including p53, BRCA1, topoisomerase III, replication protein A and DNA polymerase delta. Here, we review this network of protein interactions, and the clues that they present regarding the potential roles of RecQ family members in DNA repair, replication and/or recombination pathways.
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PMID:DNA helicase deficiencies associated with cancer predisposition and premature ageing disorders. 1125 7

The Saccharomyces cerevisiae SGS1 gene is a member of the RecQ family of ATP-dependent DNA helicases, which includes the human WRN, BLM and RECQ4 genes. Mutations in the WRN gene cause the human premature ageing disorder, Werner's syndrome. Deletion of the SGS1 gene also causes premature ageing in yeast, suggesting that the molecular mechanisms of cellular ageing may be evolutionarily conserved. To investigate the role of the RecQ helicase domain in ageing, a point mutation (SGS1 K(706)-->A) known to eliminate the DNA helicase activity of Sgs1p was constructed. This mutant allele failed to rescue the premature ageing of the sgs1Delta strain, demonstrating that Sgs1p DNA helicase activity is required for a normal lifespan. In contrast, the SGS1 K(706)-->A allele was sufficient to rescue the hypersensitivity of the sgs1Delta strain to topoisomerase inhibitors, but not other genotoxic agents. These findings support the idea that Sgs1p fulfils multiple cellular functions, and that DNA helicase activity is dispensable for some of these (e.g. functional interaction with topoisomerases), but essential for others (e.g. longevity).
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PMID:The DNA helicase activity of yeast Sgs1p is essential for normal lifespan but not for resistance to topoisomerase inhibitors. 1138 27

It has been proposed that cells monitor chromatid catenation status after DNA replication and inhibit progression into mitosis until chromatids are correctly decatenated by topoisomerase II (TopoII). Studies in yeast have suggested that TopoII may interact with RecQ helicases during this process. Using ICRF187, a TopoII catalytic inhibitor that prevents chromatid decatenation without producing DNA strand breaks, we demonstrated that cells deficient of WRN, a human RecQ helicase, displayed a defect in decatenation checkpoint activation, which was corrected by ectopic expression of wild-type WRN. We also provide evidence that BRCA1 is phosphorylated in an ATR-dependent manner in response to decatenation checkpoint activation and that this phosphorylation is not detectable in Werner syndrome cells. Furthermore, ICRF187 treatment resulted in coimmunoprecipitation of WRN and TopoII. Finally, we demonstrated that override of the decatenation checkpoint resulted in enhanced chromosomal damage and apoptosis only in the absence of WRN, but not in normal cells. Our findings suggest that WRN plays a role in the activation of G(2) decatenation checkpoint and that the abortive function of this pathway itself does not appear to be sufficient to cause genomic instability but rather predisposes to genomic instability and apoptotic cell death in the absence of other "caretaker" genes, such as WRN.
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PMID:The G2-phase decatenation checkpoint is defective in Werner syndrome cells. 1281 Jun 61


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