DNA methylation impacts the cleavage activity of chlorella virus topoisomerase II

Jennifer S. Dickey, James L. Van Etten, Neil Osheroff

Research output: Contribution to journalArticle

4 Citations (Scopus)

Abstract

Topoisomerase II from Paramecium bursaria chlorella virus-1 (PBCV-1) and chlorella virus Marburg-1 (CVM-1) displays an extraordinarily high in vitro DNA cleavage activity that is 30-50 times higher than that of human topoisomerase IIα. This remarkable scission activity may reflect a unique role played by the type II enzyme during the viral life cycle that extends beyond the normal control of DNA topology. Alternatively, but not mutually exclusively, it may reflect an adaptation to some aspect of the viral environment that differs from the in vitro conditions. To this point, the genomes of many chlorella viruses contain high levels of N6-methyladenine (6mA) and 5-methylcytosine (5mC), but the DNA employed in vitro is unmodified. Therefore, to determine whether methylation impacts the ability of chlorella virus topoisomerase II to cleave DNA, the effects of 6mA and 5mC on the PBCV-1 and CVM-1 enzymes were examined. Results indicate that 6mA strongly inhibits DNA scission mediated by both enzymes, while 5mC has relatively little effect. At levels of 6mA and 5mC methylation comparable to those found in the CVM-1 genome (10% 6mA and 42% 5mC), the level of DNA cleavage decreased ∼4-fold. As determined using a novel rapid quench pre-equilibrium DNA cleavage system in conjunction with oligonucleotide binding and ligation assays, this decrease appears to be caused primarily by a slower forward rate of DNA scission. These findings suggest that the high DNA cleavage activity of chlorella virus topoisomerase II on unmodified nucleic acid substrates may reflect, at least in part, an adaptation to act on methylated genomic DNA.

Original languageEnglish (US)
Pages (from-to)15378-15386
Number of pages9
JournalBiochemistry
Volume44
Issue number46
DOIs
StatePublished - Nov 22 2005

Fingerprint

Chlorella
Type II DNA Topoisomerase
DNA Methylation
Viruses
5-Methylcytosine
DNA Cleavage
Marburgvirus
DNA
Paramecium
Methylation
Enzymes
Genome
Genes
Life Cycle Stages
Oligonucleotides
Nucleic Acids
Ligation
Life cycle
Assays
Topology

ASJC Scopus subject areas

  • Biochemistry

Cite this

DNA methylation impacts the cleavage activity of chlorella virus topoisomerase II. / Dickey, Jennifer S.; Van Etten, James L.; Osheroff, Neil.

In: Biochemistry, Vol. 44, No. 46, 22.11.2005, p. 15378-15386.

Research output: Contribution to journalArticle

Dickey, Jennifer S. ; Van Etten, James L. ; Osheroff, Neil. / DNA methylation impacts the cleavage activity of chlorella virus topoisomerase II. In: Biochemistry. 2005 ; Vol. 44, No. 46. pp. 15378-15386.
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abstract = "Topoisomerase II from Paramecium bursaria chlorella virus-1 (PBCV-1) and chlorella virus Marburg-1 (CVM-1) displays an extraordinarily high in vitro DNA cleavage activity that is 30-50 times higher than that of human topoisomerase IIα. This remarkable scission activity may reflect a unique role played by the type II enzyme during the viral life cycle that extends beyond the normal control of DNA topology. Alternatively, but not mutually exclusively, it may reflect an adaptation to some aspect of the viral environment that differs from the in vitro conditions. To this point, the genomes of many chlorella viruses contain high levels of N6-methyladenine (6mA) and 5-methylcytosine (5mC), but the DNA employed in vitro is unmodified. Therefore, to determine whether methylation impacts the ability of chlorella virus topoisomerase II to cleave DNA, the effects of 6mA and 5mC on the PBCV-1 and CVM-1 enzymes were examined. Results indicate that 6mA strongly inhibits DNA scission mediated by both enzymes, while 5mC has relatively little effect. At levels of 6mA and 5mC methylation comparable to those found in the CVM-1 genome (10{\%} 6mA and 42{\%} 5mC), the level of DNA cleavage decreased ∼4-fold. As determined using a novel rapid quench pre-equilibrium DNA cleavage system in conjunction with oligonucleotide binding and ligation assays, this decrease appears to be caused primarily by a slower forward rate of DNA scission. These findings suggest that the high DNA cleavage activity of chlorella virus topoisomerase II on unmodified nucleic acid substrates may reflect, at least in part, an adaptation to act on methylated genomic DNA.",
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