scholarly journals Dynamics of human DNA topoisomerases IIα and IIβ in living cells

2002 ◽  
Vol 157 (1) ◽  
pp. 31-44 ◽  
Author(s):  
Morten O. Christensen ◽  
Morten K. Larsen ◽  
Hans Ullrich Barthelmes ◽  
Robert Hock ◽  
Claus L. Andersen ◽  
...  
Keyword(s):  
Structural Component ◽  
Mitotic Chromosome ◽  
Proliferating Cells ◽  
Dna Topoisomerase ◽  
Genomic Changes ◽  
Human Dna ◽  
Catalytic Dna ◽  
Topo Ii ◽  
Chromosome Scaffold

DNA topoisomerase (topo) II catalyses topological genomic changes essential for many DNA metabolic processes. It is also regarded as a structural component of the nuclear matrix in interphase and the mitotic chromosome scaffold. Mammals have two isoforms (α and β) with similar properties in vitro. Here, we investigated their properties in living and proliferating cells, stably expressing biofluorescent chimera of the human isozymes. Topo IIα and IIβ behaved similarly in interphase but differently in mitosis, where only topo IIα was chromosome associated to a major part. During interphase, both isozymes joined in nucleolar reassembly and accumulated in nucleoli, which seemed not to involve catalytic DNA turnover because treatment with teniposide (stabilizing covalent catalytic DNA intermediates of topo II) relocated the bulk of the enzymes from the nucleoli to nucleoplasmic granules. Photobleaching revealed that the entire complement of both isozymes was completely mobile and free to exchange between nuclear subcompartments in interphase. In chromosomes, topo IIα was also completely mobile and had a uniform distribution. However, hypotonic cell lysis triggered an axial pattern. These observations suggest that topo II is not an immobile, structural component of the chromosomal scaffold or the interphase karyoskeleton, but rather a dynamic interaction partner of such structures.

scholarly journals Linker histone H1.8 inhibits chromatin-binding of condensins and DNA topoisomerase II to tune chromosome length and individualization

eLife ◽  
2021 ◽  
Vol 10 ◽  
Author(s):  
Pavan Choppakatla ◽  
Bastiaan Dekker ◽  
Erin E Cutts ◽  
Alessandro Vannini ◽  
Job Dekker ◽  
...  
Keyword(s):  
Topoisomerase Ii ◽  
Mitotic Chromosome ◽  
Chromosome Length ◽  
Chromosome Morphology ◽  
Linker Histone ◽  
Dna Topoisomerase Ii ◽  
Dna Amount ◽  
Dna Topoisomerase ◽  
Topo Ii

DNA loop extrusion by condensins and decatenation by DNA topoisomerase II (topo II) are thought to drive mitotic chromosome compaction and individualization. Here, we reveal that the linker histone H1.8 antagonizes condensins and topo II to shape mitotic chromosome organization. In vitro chromatin reconstitution experiments demonstrate that H1.8 inhibits binding of condensins and topo II to nucleosome arrays. Accordingly, H1.8 depletion in Xenopus egg extracts increased condensins and topo II levels on mitotic chromatin. Chromosome morphology and Hi-C analyses suggest that H1.8 depletion makes chromosomes thinner and longer through shortening the average loop size and reducing the DNA amount in each layer of mitotic loops. Furthermore, excess loading of condensins and topo II to chromosomes by H1.8 depletion causes hyper-chromosome individualization and dispersion. We propose that condensins and topo II are essential for chromosome individualization, but their functions are tuned by the linker histone to keep chromosomes together until anaphase.

scholarly journals In Vitro and In Silico Characterization of an Antimalarial Compound with Antitumor Activity Targeting Human DNA Topoisomerase IB

2021 ◽  
Vol 22 (14) ◽  
pp. 7455
Author(s):  
Bini Chhetri Soren ◽  
Jagadish Babu Dasari ◽  
Alessio Ottaviani ◽  
Beatrice Messina ◽  
Giada Andreotti ◽  
...  
Keyword(s):  
Enzyme Reaction ◽  
Dna Topoisomerase ◽  
Human Dna ◽  
Topoisomerase Ib ◽  
Topological State ◽  
Cleavage Step ◽  
Dynamics Simulations ◽  
In Silico Characterization ◽  
Dna Complex

Human DNA topoisomerase IB controls the topological state of supercoiled DNA through a complex catalytic cycle that consists of cleavage and religation reactions, allowing the progression of fundamental DNA metabolism. The catalytic steps of human DNA topoisomerase IB were analyzed in the presence of a drug, obtained by the open-access drug bank Medicines for Malaria Venture. The experiments indicate that the compound strongly and irreversibly inhibits the cleavage step of the enzyme reaction and reduces the cell viability of three different cancer cell lines. Molecular docking and molecular dynamics simulations suggest that the drug binds to the human DNA topoisomerase IB-DNA complex sitting inside the catalytic site of the enzyme, providing a molecular explanation for the cleavage-inhibition effect. For all these reasons, the aforementioned drug could be a possible lead compound for the development of an efficient anti-tumor molecule targeting human DNA topoisomerase IB.

Synthesis, crystal structure, DNA interaction and in vitro anticancer activity of a Cu(ii) complex of purpurin: dual poison for human DNA topoisomerase I and II

RSC Advances ◽  
2014 ◽  
Vol 4 (103) ◽  
pp. 59344-59357 ◽  
Author(s):  
Piyal Das ◽  
Chetan Kumar Jain ◽  
Sanjoy K. Dey ◽  
Rajat Saha ◽  
Abhishek Dutta Chowdhury ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Topoisomerase I ◽  
Dna Interaction ◽  
Dna Topoisomerase I ◽  
Oxygen Species ◽  
Dna Topoisomerase ◽  
Human Dna ◽  
Anti Tumor Activity ◽  
Human Dna Topoisomerase I

Although generation of reactive oxygen species (ROS) by anthracycline anticancer drugs is essential for anti-tumor activity, they make these drugs cardiotoxic.

Bioassays and In Silico Methods in the Identification of Human DNA Topoisomerase IIα Inhibitors

2018 ◽  
Vol 25 (28) ◽  
pp. 3286-3318 ◽  
Author(s):  
Kaja Bergant ◽  
Matej Janezic ◽  
Andrej Perdih
Keyword(s):  
In Silico ◽  
In Vivo Studies ◽  
Topoisomerase Iiα ◽  
Dna Topoisomerase ◽  
Human Dna ◽  
Practical Guidelines ◽  
In Silico Methods ◽  
Dna Topoisomerase Iiα

Background: The family of DNA topoisomerases comprises a group of enzymes that catalyse the induction of topological changes to DNA. These enzymes play a role in the cell replication machinery and are, therefore, important targets for anticancer drugs - with human DNA topoisomerase IIα being one of the most prominent. Active compounds targeting this enzyme are classified into two groups with diverse mechanisms of action: DNA poisons act by stabilizing a covalent cleavage complex between DNA and the topoisomerase enzyme, transforming it into a cellular toxin, while the second diverse group of catalytic inhibitors, provides novel inhibition avenues for tackling this enzyme due to frequent occurrence of side effects observed during the DNA poison therapy. Methods: Based on a comprehensive literature search we present an overview of available bioassays and in silico methods in the identification of human DNA topoisomerase IIα inhibitors. Results and Conclusion: A comprehensive outline of the available methods and approaches that explore in detail the in vitro mechanistic and functional aspects of the topoisomerase IIα inhibition of both topo IIα inhibitor groups is presented. The utilized in vitro cell-based assays and in vivo studies to further explore the validated topo IIα inhibitors in subsequent preclinical stages of the drug discovery are discussed. The potential of in silico methods in topoisomerase IIα inhibitor discovery is outlined. A list of practical guidelines was compiled to aid new as well experienced researchers in how to optimally approach the design of targeted inhibitors and validation in the preclinical drug development stages.

scholarly journals Human DNA topoisomerase I-mediated cleavage and recombination of duck hepatitis B virus DNA in vitro

1999 ◽  
Vol 27 (8) ◽  
pp. 1919-1925 ◽  
Author(s):  
P. Pourquier ◽  
Y. Pommier ◽  
A. D. Jensen ◽  
S. S. Gong ◽  
C. E. Rogler
Keyword(s):  
Hepatitis B Virus ◽  
Hepatitis B ◽  
Topoisomerase I ◽  
Dna Topoisomerase I ◽  
Dna Topoisomerase ◽  
Duck Hepatitis ◽  
Human Dna ◽  
B Virus ◽  
Human Dna Topoisomerase I

scholarly journals Topoisomerase II does not play a scaffolding role in the organization of mitotic chromosomes assembled in Xenopus egg extracts.

1993 ◽  
Vol 120 (3) ◽  
pp. 601-612 ◽  
Author(s):  
T Hirano ◽  
T J Mitchison
Keyword(s):  
Topoisomerase Ii ◽  
Mitotic Chromosome ◽  
Chromosome Morphology ◽  
Sperm Chromatin ◽  
Mitotic Chromosomes ◽  
Egg Extracts ◽  
Xenopus Egg ◽  
Topo Ii ◽  
Xenopus Egg Extracts

We have investigated the role of topoisomerase II (topo II) in mitotic chromosome assembly and organization in vitro using Xenopus egg extracts. When sperm chromatin was incubated with mitotic extracts, the highly compact chromatin rapidly swelled and concomitantly underwent local condensation. Further incubation induced the formation of entangled thin chromatin fibers that eventually resolved into highly condensed individual chromosomes. This in vitro system made it possible to manipulate mitotic chromosomes in their assembly condition without any isolation or stabilization steps. Two complementary approaches, immunodepletion and antibody blocking, demonstrated that topo II activity is required for chromosome assembly and condensation. Once condensation was completed, however, blocking of topo II activity had little effect on the chromosome morphology. Immunofluorescent studies showed that topo II was uniformly distributed throughout the condensed chromosomes and was not restricted to the chromosomal axis. Surprisingly, all detectable topo II molecules were easily extracted from the chromosomes under mild conditions where the shape of chromosomes was well preserved. Our results show that topo II is essential for mitotic chromosome assembly, but does not play a scaffolding role in the structural maintenance of chromosomes assembled in vitro. We also present evidence that changes of DNA topology affect the distribution of topo II in mitotic chromosomes in our system.

scholarly journals Guiding functions of the C-terminal domain of topoisomerase IIα advance mitotic chromosome assembly

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Keishi Shintomi ◽  
Tatsuya Hirano
Keyword(s):  
Topoisomerase Ii ◽  
Mitotic Chromosome ◽  
Fine Tuning ◽  
Terminal Domain ◽  
Nucleosomal Dna ◽  
Egg Extracts ◽  
Guiding Functions ◽  
Topo Ii ◽  
Crowded Environments

AbstractTopoisomerase II (topo II) is one of the six proteins essential for mitotic chromatid reconstitution in vitro. It is not fully understood, however, mechanistically how this enzyme regulates this process. In an attempt to further refine the reconstitution assay, we have found that chromosomal binding of Xenopus laevis topo IIα is sensitive to buffer conditions and depends on its C-terminal domain (CTD). Enzymological assays using circular DNA substrates supports the idea that topo IIα first resolves inter-chromatid entanglements to drive individualization and then generates intra-chromatid entanglements to promote thickening. Importantly, only the latter process requires the CTD. By using frog egg extracts, we also show that the CTD contributes to proper formation of nucleosome-depleted chromatids by competing with a linker histone for non-nucleosomal DNA. Our results demonstrate that topo IIα utilizes its CTD to deliver the enzymatic core to crowded environments created during mitotic chromatid assembly, thereby fine-tuning this process.

Sign in / Sign up

Export Citation Format

Share Document