Effects of benzo[a]pyrene-deoxyguanosine lesions on DNA methylation catalyzed by EcoRII DNA methyltransferase and on DNA cleavage effected by EcoRII restriction endonuclease

Vladimir B. Baskunov, Fedor V. Subach, Alexandr Kolbanovskiy, Marina Kolbanovskiy, Sergei A. Eremin, Francis Johnson, Radha Bonala, Nicholas Geacintov, Elizaveta S. Gromova

Research output: Contribution to journalArticle

Abstract

DNA methylation is an important cellular mechanism for controlling gene expression. Whereas the mutagenic properties of many DNA adducts, e.g., those arising from polycyclic aromatic hydrocarbons, have been widely studied, little is known about their influence on DNA methylation. We have constructed site-specifically modified 18-mer oligodeoxynucleotide duplexes containing a pair of stereoisomeric adducts derived from a benzo[a]pyrene-derived diol epoxide [(+)- and (-)-r7,t8-dihydroxy-t9,10-epoxy-7,8,9,-10-tetrahydrobenzo[a] pyrene, or B[a]PDE] bound to the exocyclic amino group of guanine. The adducts, either (+)- or (-)-trans-anti-B[a]P-N2-dG (G*), positioned either at the 5′-side or the 3′-side deoxyguanosine residue in the recognition sequence of EcoRII restriction-modification enzymes (5′-...CCA/ TGG...) were incorporated into 18-mer oligodeoxynucleotide duplexes. The effects of these lesions on complex formation and the catalytic activity of the EcoRII DNA methyltransferase (M.EcoRII) and EcoRII restriction endonuclease (R.EcoRII) were investigated. The M.EcoRII catalyzes the transfer of a methyl group to the C5 position of the 3′-side cytosine of each strand of the recognition sequence, whereas R.EcoRII catalyzes cleavage of both strands. The binding of R.EcoRII to the oligodeoxynucleotide duplexes and the catalytic cleavage were completely abolished when G* was positioned at the 3′-side dG position (5′-...CCTGG*...). When G* was at the 5′-side dG position, binding was moderately diminished, but cleavage was completely blocked. In the case of M.EcoRII, binding is diminished by factors of 5-30 but the catalytic activity was either abolished or reduced 4-80-fold when the adducts were located at either position. Somewhat smaller effects were observed with hemimethylated oligodeoxynucleotide duplexes. These findings suggest that epigenetic effects, in addition to genotoxic effects, need to be considered in chemical carcinogenesis initiated by B[a]PDE, since the inhibition of methylation may allow the expression of genes that promote tumor development.

Original languageEnglish (US)
Pages (from-to)1054-1066
Number of pages13
JournalBiochemistry
Volume44
Issue number3
DOIs
StatePublished - Jan 25 2005

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DNA Cleavage
Deoxyguanosine
Oligodeoxyribonucleotides
Benzo(a)pyrene
DNA Restriction Enzymes
Methyltransferases
DNA Methylation
DNA
Catalyst activity
Gene Expression
Methylation
DNA Adducts
Cytosine
Polycyclic Aromatic Hydrocarbons
Epoxy Compounds
Guanine
Epigenomics
Gene expression
Tumors
Carcinogenesis

ASJC Scopus subject areas

  • Biochemistry

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Effects of benzo[a]pyrene-deoxyguanosine lesions on DNA methylation catalyzed by EcoRII DNA methyltransferase and on DNA cleavage effected by EcoRII restriction endonuclease. / Baskunov, Vladimir B.; Subach, Fedor V.; Kolbanovskiy, Alexandr; Kolbanovskiy, Marina; Eremin, Sergei A.; Johnson, Francis; Bonala, Radha; Geacintov, Nicholas; Gromova, Elizaveta S.

In: Biochemistry, Vol. 44, No. 3, 25.01.2005, p. 1054-1066.

Research output: Contribution to journalArticle

Baskunov, Vladimir B. ; Subach, Fedor V. ; Kolbanovskiy, Alexandr ; Kolbanovskiy, Marina ; Eremin, Sergei A. ; Johnson, Francis ; Bonala, Radha ; Geacintov, Nicholas ; Gromova, Elizaveta S. / Effects of benzo[a]pyrene-deoxyguanosine lesions on DNA methylation catalyzed by EcoRII DNA methyltransferase and on DNA cleavage effected by EcoRII restriction endonuclease. In: Biochemistry. 2005 ; Vol. 44, No. 3. pp. 1054-1066.
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title = "Effects of benzo[a]pyrene-deoxyguanosine lesions on DNA methylation catalyzed by EcoRII DNA methyltransferase and on DNA cleavage effected by EcoRII restriction endonuclease",
abstract = "DNA methylation is an important cellular mechanism for controlling gene expression. Whereas the mutagenic properties of many DNA adducts, e.g., those arising from polycyclic aromatic hydrocarbons, have been widely studied, little is known about their influence on DNA methylation. We have constructed site-specifically modified 18-mer oligodeoxynucleotide duplexes containing a pair of stereoisomeric adducts derived from a benzo[a]pyrene-derived diol epoxide [(+)- and (-)-r7,t8-dihydroxy-t9,10-epoxy-7,8,9,-10-tetrahydrobenzo[a] pyrene, or B[a]PDE] bound to the exocyclic amino group of guanine. The adducts, either (+)- or (-)-trans-anti-B[a]P-N2-dG (G*), positioned either at the 5′-side or the 3′-side deoxyguanosine residue in the recognition sequence of EcoRII restriction-modification enzymes (5′-...CCA/ TGG...) were incorporated into 18-mer oligodeoxynucleotide duplexes. The effects of these lesions on complex formation and the catalytic activity of the EcoRII DNA methyltransferase (M.EcoRII) and EcoRII restriction endonuclease (R.EcoRII) were investigated. The M.EcoRII catalyzes the transfer of a methyl group to the C5 position of the 3′-side cytosine of each strand of the recognition sequence, whereas R.EcoRII catalyzes cleavage of both strands. The binding of R.EcoRII to the oligodeoxynucleotide duplexes and the catalytic cleavage were completely abolished when G* was positioned at the 3′-side dG position (5′-...CCTGG*...). When G* was at the 5′-side dG position, binding was moderately diminished, but cleavage was completely blocked. In the case of M.EcoRII, binding is diminished by factors of 5-30 but the catalytic activity was either abolished or reduced 4-80-fold when the adducts were located at either position. Somewhat smaller effects were observed with hemimethylated oligodeoxynucleotide duplexes. These findings suggest that epigenetic effects, in addition to genotoxic effects, need to be considered in chemical carcinogenesis initiated by B[a]PDE, since the inhibition of methylation may allow the expression of genes that promote tumor development.",
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AU - Baskunov, Vladimir B.

AU - Subach, Fedor V.

AU - Kolbanovskiy, Alexandr

AU - Kolbanovskiy, Marina

AU - Eremin, Sergei A.

AU - Johnson, Francis

AU - Bonala, Radha

AU - Geacintov, Nicholas

AU - Gromova, Elizaveta S.

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N2 - DNA methylation is an important cellular mechanism for controlling gene expression. Whereas the mutagenic properties of many DNA adducts, e.g., those arising from polycyclic aromatic hydrocarbons, have been widely studied, little is known about their influence on DNA methylation. We have constructed site-specifically modified 18-mer oligodeoxynucleotide duplexes containing a pair of stereoisomeric adducts derived from a benzo[a]pyrene-derived diol epoxide [(+)- and (-)-r7,t8-dihydroxy-t9,10-epoxy-7,8,9,-10-tetrahydrobenzo[a] pyrene, or B[a]PDE] bound to the exocyclic amino group of guanine. The adducts, either (+)- or (-)-trans-anti-B[a]P-N2-dG (G*), positioned either at the 5′-side or the 3′-side deoxyguanosine residue in the recognition sequence of EcoRII restriction-modification enzymes (5′-...CCA/ TGG...) were incorporated into 18-mer oligodeoxynucleotide duplexes. The effects of these lesions on complex formation and the catalytic activity of the EcoRII DNA methyltransferase (M.EcoRII) and EcoRII restriction endonuclease (R.EcoRII) were investigated. The M.EcoRII catalyzes the transfer of a methyl group to the C5 position of the 3′-side cytosine of each strand of the recognition sequence, whereas R.EcoRII catalyzes cleavage of both strands. The binding of R.EcoRII to the oligodeoxynucleotide duplexes and the catalytic cleavage were completely abolished when G* was positioned at the 3′-side dG position (5′-...CCTGG*...). When G* was at the 5′-side dG position, binding was moderately diminished, but cleavage was completely blocked. In the case of M.EcoRII, binding is diminished by factors of 5-30 but the catalytic activity was either abolished or reduced 4-80-fold when the adducts were located at either position. Somewhat smaller effects were observed with hemimethylated oligodeoxynucleotide duplexes. These findings suggest that epigenetic effects, in addition to genotoxic effects, need to be considered in chemical carcinogenesis initiated by B[a]PDE, since the inhibition of methylation may allow the expression of genes that promote tumor development.

AB - DNA methylation is an important cellular mechanism for controlling gene expression. Whereas the mutagenic properties of many DNA adducts, e.g., those arising from polycyclic aromatic hydrocarbons, have been widely studied, little is known about their influence on DNA methylation. We have constructed site-specifically modified 18-mer oligodeoxynucleotide duplexes containing a pair of stereoisomeric adducts derived from a benzo[a]pyrene-derived diol epoxide [(+)- and (-)-r7,t8-dihydroxy-t9,10-epoxy-7,8,9,-10-tetrahydrobenzo[a] pyrene, or B[a]PDE] bound to the exocyclic amino group of guanine. The adducts, either (+)- or (-)-trans-anti-B[a]P-N2-dG (G*), positioned either at the 5′-side or the 3′-side deoxyguanosine residue in the recognition sequence of EcoRII restriction-modification enzymes (5′-...CCA/ TGG...) were incorporated into 18-mer oligodeoxynucleotide duplexes. The effects of these lesions on complex formation and the catalytic activity of the EcoRII DNA methyltransferase (M.EcoRII) and EcoRII restriction endonuclease (R.EcoRII) were investigated. The M.EcoRII catalyzes the transfer of a methyl group to the C5 position of the 3′-side cytosine of each strand of the recognition sequence, whereas R.EcoRII catalyzes cleavage of both strands. The binding of R.EcoRII to the oligodeoxynucleotide duplexes and the catalytic cleavage were completely abolished when G* was positioned at the 3′-side dG position (5′-...CCTGG*...). When G* was at the 5′-side dG position, binding was moderately diminished, but cleavage was completely blocked. In the case of M.EcoRII, binding is diminished by factors of 5-30 but the catalytic activity was either abolished or reduced 4-80-fold when the adducts were located at either position. Somewhat smaller effects were observed with hemimethylated oligodeoxynucleotide duplexes. These findings suggest that epigenetic effects, in addition to genotoxic effects, need to be considered in chemical carcinogenesis initiated by B[a]PDE, since the inhibition of methylation may allow the expression of genes that promote tumor development.

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