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The Impact of the Human DNA Topoisomerase II C-Terminal Domain on Activity

BACKGROUND: Type II DNA topoisomerases (topos) are essential enzymes needed for the resolution of topological problems that occur during DNA metabolic processes. Topos carry out an ATP-dependent strand passage reaction whereby one double helix is passed through a transient break in another. Humans h...

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Autores principales: Meczes, Emma L., Gilroy, Kathryn L., West, Katherine L., Austin, Caroline A.
Formato: Texto
Lenguaje:English
Publicado: Public Library of Science 2008
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2262138/
https://www.ncbi.nlm.nih.gov/pubmed/18335031
http://dx.doi.org/10.1371/journal.pone.0001754
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author Meczes, Emma L.
Gilroy, Kathryn L.
West, Katherine L.
Austin, Caroline A.
author_facet Meczes, Emma L.
Gilroy, Kathryn L.
West, Katherine L.
Austin, Caroline A.
author_sort Meczes, Emma L.
collection PubMed
description BACKGROUND: Type II DNA topoisomerases (topos) are essential enzymes needed for the resolution of topological problems that occur during DNA metabolic processes. Topos carry out an ATP-dependent strand passage reaction whereby one double helix is passed through a transient break in another. Humans have two topoII isoforms, α and β, which while enzymatically similar are differentially expressed and regulated, and are thought to have different cellular roles. The C-terminal domain (CTD) of the enzyme has the most diversity, and has been implicated in regulation. We sought to investigate the impact of the CTD domain on activity. METHODOLOGY/PRINCIPLE FINDINGS: We have investigated the role of the human topoII C-terminal domain by creating constructs encoding C-terminally truncated recombinant topoIIα and β and topoIIα+β-tail and topoIIβ+α-tail chimeric proteins. We then investigated function in vivo in a yeast system, and in vitro in activity assays. We find that the C-terminal domain of human topoII isoforms is needed for in vivo function of the enzyme, but not needed for cleavage activity. C-terminally truncated enzymes had similar strand passage activity to full length enzymes, but the presence of the opposite C-terminal domain had a large effect, with the topoIIα-CTD increasing activity, and the topoIIβ-CTD decreasing activity. CONCLUSIONS/SIGNIFICANCE: In vivo complementation data show that the topoIIα C-terminal domain is needed for growth, but the topoIIβ isoform is able to support low levels of growth without a C-terminal domain. This may indicate that topoIIβ has an additional localisation signal. In vitro data suggest that, while the lack of any C-terminal domain has little effect on activity, the presence of either the topoIIα or β C-terminal domain can affect strand passage activity. Data indicates that the topoIIβ-CTD may be a negative regulator. This is the first report of in vitro data with chimeric human topoIIs.
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spelling pubmed-22621382008-03-12 The Impact of the Human DNA Topoisomerase II C-Terminal Domain on Activity Meczes, Emma L. Gilroy, Kathryn L. West, Katherine L. Austin, Caroline A. PLoS One Research Article BACKGROUND: Type II DNA topoisomerases (topos) are essential enzymes needed for the resolution of topological problems that occur during DNA metabolic processes. Topos carry out an ATP-dependent strand passage reaction whereby one double helix is passed through a transient break in another. Humans have two topoII isoforms, α and β, which while enzymatically similar are differentially expressed and regulated, and are thought to have different cellular roles. The C-terminal domain (CTD) of the enzyme has the most diversity, and has been implicated in regulation. We sought to investigate the impact of the CTD domain on activity. METHODOLOGY/PRINCIPLE FINDINGS: We have investigated the role of the human topoII C-terminal domain by creating constructs encoding C-terminally truncated recombinant topoIIα and β and topoIIα+β-tail and topoIIβ+α-tail chimeric proteins. We then investigated function in vivo in a yeast system, and in vitro in activity assays. We find that the C-terminal domain of human topoII isoforms is needed for in vivo function of the enzyme, but not needed for cleavage activity. C-terminally truncated enzymes had similar strand passage activity to full length enzymes, but the presence of the opposite C-terminal domain had a large effect, with the topoIIα-CTD increasing activity, and the topoIIβ-CTD decreasing activity. CONCLUSIONS/SIGNIFICANCE: In vivo complementation data show that the topoIIα C-terminal domain is needed for growth, but the topoIIβ isoform is able to support low levels of growth without a C-terminal domain. This may indicate that topoIIβ has an additional localisation signal. In vitro data suggest that, while the lack of any C-terminal domain has little effect on activity, the presence of either the topoIIα or β C-terminal domain can affect strand passage activity. Data indicates that the topoIIβ-CTD may be a negative regulator. This is the first report of in vitro data with chimeric human topoIIs. Public Library of Science 2008-03-12 /pmc/articles/PMC2262138/ /pubmed/18335031 http://dx.doi.org/10.1371/journal.pone.0001754 Text en Meczes et al. http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are properly credited.
spellingShingle Research Article
Meczes, Emma L.
Gilroy, Kathryn L.
West, Katherine L.
Austin, Caroline A.
The Impact of the Human DNA Topoisomerase II C-Terminal Domain on Activity
title The Impact of the Human DNA Topoisomerase II C-Terminal Domain on Activity
title_full The Impact of the Human DNA Topoisomerase II C-Terminal Domain on Activity
title_fullStr The Impact of the Human DNA Topoisomerase II C-Terminal Domain on Activity
title_full_unstemmed The Impact of the Human DNA Topoisomerase II C-Terminal Domain on Activity
title_short The Impact of the Human DNA Topoisomerase II C-Terminal Domain on Activity
title_sort impact of the human dna topoisomerase ii c-terminal domain on activity
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2262138/
https://www.ncbi.nlm.nih.gov/pubmed/18335031
http://dx.doi.org/10.1371/journal.pone.0001754
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