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

2020 ◽  
Author(s):  
Pavan Choppakatla ◽  
Bastiaan Dekker ◽  
Erin E. Cutts ◽  
Alessandro Vannini ◽  
Job Dekker ◽  
...  
Keyword(s):  
Topoisomerase Ii ◽  
Chromosome Morphology ◽  
Linker Histone ◽  
Dna Topoisomerase Ii ◽  
Dna Amount ◽  
Dna Topoisomerase ◽  
Dna Loop ◽  
Egg Extracts ◽  
Topo Ii

SummaryDNA loop extrusion by condensins and decatenation by DNA topoisomerase II (topo II) drive mitotic chromosome compaction and individualization. Here, we reveal that the linker histone H1.8 regulates chromatin levels of condensins and topo II. 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 likely through shortening the average loop size and reducing DNA amount in each layer of mitotic loops. Furthermore, H1.8-mediated suppression of condensins and topo II binding to chromatin limits chromosome individualization by preventing resolution of interchromosomal linkages. While linker histones locally compact DNA by clustering nucleosomes, we propose that H1.8 controls chromosome morphology and topological organization through restricting the loading of condensins and topo II on chromatin.

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.

The in vivo distribution and dynamics of DNA topoisomerase II in Drosophila embryonic nuclei and chromosomes

1993 ◽  
Vol 51 ◽  
pp. 74-75
Author(s):  
Jason R. Swedlow ◽  
Neil Osheroff ◽  
Tim Karr ◽  
John W. Sedat ◽  
David A. Agard
Keyword(s):  
Topoisomerase Ii ◽  
Biochemical Characterization ◽  
Developmental Cycle ◽  
Dna Topoisomerase Ii ◽  
Chromosomal Distribution ◽  
Dna Topoisomerase ◽  
Ideal System ◽  
Dnase Treatment

DNA topoisomerase II is an ATP-dependent double-stranded DNA strand-passing enzyme that is necessary for full condensation of chromosomes and for complete segregation of sister chromatids at mitosis in vivo and in vitro. Biochemical characterization of chromosomes or nuclei after extraction with high-salt or detergents and DNAse treatment showed that topoisomerase II was a major component of this remnant, termed the chromosome scaffold. The scaffold has been hypothesized to be the structural backbone of the chromosome, so the localization of topoisomerase II to die scaffold suggested that the enzyme might play a structural role in the chromosome. However, topoisomerase II has not been studied in nuclei or chromosomes in vivo. We have monitored the chromosomal distribution of topoisomerase II in vivo during mitosis in the Drosophila embryo. This embryo forms a multi-nucleated syncytial blastoderm early in its developmental cycle. During this time, the embryonic nuclei synchronously progress through 13 mitotic cycles, so this is an ideal system to follow nuclear and chromosomal dynamics.

The C-terminal domain of Saccharomyces cerevisiae DNA topoisomerase II

1994 ◽  
Vol 14 (5) ◽  
pp. 3197-3207
Author(s):  
P R Caron ◽  
P Watt ◽  
J C Wang
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Nuclear Localization ◽  
Topoisomerase Ii ◽  
Dna Topoisomerase Ii ◽  
Dna Topoisomerase ◽  
Mutant Proteins ◽  
Terminal Domain ◽  
Catalytically Active

A set of carboxy-terminal deletion mutants of Saccharomyces cerevisiae DNA topoisomerase II were constructed for studying the functions of the carboxyl domain in vitro and in vivo. The wild-type yeast enzyme is a homodimer with 1,429 amino acid residues in each of the two polypeptides; truncation of the C terminus to Ile-1220 has little effect on the function of the enzyme in vitro or in vivo, whereas truncations extending beyond Gln-1138 yield completely inactive proteins. Several mutant enzymes with C termini in between these two residues were found to be catalytically active but unable to complement a top2-4 temperature-sensitive mutation. Immunomicroscopy results suggest that the removal of a nuclear localization signal in the C-terminal domain is likely to contribute to the physiological dysfunction of these proteins; the ability of these mutant proteins to relax supercoiled DNA in vivo shows, however, that at least some of the mutant proteins are present in the nuclei in a catalytically active form. In contrast to the ability of the catalytically active mutant proteins to relax supercoiled intracellular DNA, all mutants that do not complement the temperature-dependent lethality and high frequency of chromosomal nondisjunction of top2-4 were found to lack decatenation activity in vivo. The plausible roles of the DNA topoisomerase II C-terminal domain, in addition to providing a signal for nuclear localization, are discussed in the light of these results.

scholarly journals Rearrangements of the MLL gene in therapy-related acute myeloid leukemia in patients previously treated with agents targeting DNA- topoisomerase II

Blood ◽  
1993 ◽  
Vol 82 (12) ◽  
pp. 3705-3711 ◽  
Author(s):  
HJ Super ◽  
NR McCabe ◽  
MJ Thirman ◽  
RA Larson ◽  
MM Le Beau ◽  
...  
Keyword(s):  
Acute Myeloid Leukemia ◽  
Myeloid Leukemia ◽  
Topoisomerase Ii ◽  
De Novo ◽  
Chromosome Band ◽  
Dna Topoisomerase Ii ◽  
Dna Topoisomerase ◽  
Mll Gene ◽  
Topo Ii ◽  
Acute Myeloid

Chromosome band 11q23 is frequently involved in acute myeloid leukemia (AML) and acute lymphoblastic leukemia (ALL) de novo, as well as in myelodysplastic syndromes (MDS) and lymphoma. Five percent to 15% of patients treated with chemotherapy for a primary neoplasm develop therapy-related AML (t-AML) that may show rearrangements, usually translocations involving band 11q23 or, less often, 21q22. These leukemias develop after a relatively short latent period and often follow the use of drugs that inhibit the activity of DNA-topoisomerase II (topo II). We previously identified a gene, MLL (myeloid-lymphoid leukemia or mixed-lineage leukemia), at 11q23 that is involved in the de novo leukemias. We have studied 17 patients with t-MDS/t-AML, 12 of whom had cytogenetically detectable 11q23 rearrangements. Ten of the 12 t-AML patients had received topo II inhibitors and 9 of these, all with balanced translocations of 11q23, had MLL rearrangements on Southern blot analysis. None of the patients who had not received topo II inhibitors showed an MLL rearrangement. Of the 5 patients lacking 11q23 rearrangements, some of whom had monoblastic features, none had an MLL rearrangement, although 4 had received topo II inhibitors. Our study indicates that the MLL gene rearrangements are similar both in AML that develops de novo and in t-AML. The association of exposure to topo II- reactive chemotherapy with 11q23 rearrangements involving the MLL gene in t-AML suggests that topo II may play a role in the aberrant recombination events that occur in this region both in AML de novo and in t-AML.

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