Journal Article

DNA Cross-Linking by Dehydromonocrotaline Lacks Apparent Base Sequence Preference

W. Kurt Rieben and Roger A. Coulombe

in Toxicological Sciences

Volume 82, issue 2, pages 497-503
Published in print December 2004 | ISSN: 1096-6080
Published online August 2004 | e-ISSN: 1096-0929 | DOI: https://dx.doi.org/10.1093/toxsci/kfh262
DNA Cross-Linking by Dehydromonocrotaline Lacks Apparent Base Sequence Preference

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Pyrrolizidine alkaloids (PAs) are ubiquitous plant toxins, many of which, upon oxidation by hepatic mixed-function oxidases, become reactive bifunctional pyrrolic electrophiles that form DNA-DNA and DNA-protein cross-links. The anti-mitotic, toxic, and carcinogenic action of PAs is thought to be caused, at least in part, by these cross-links. We wished to determine whether the activated PA pyrrole dehydromonocrotaline (DHMO) exhibits base sequence preferences when cross-linked to a set of model duplex poly A-T 14-mer oligonucleotides with varying internal and/or end 5′-d(CG), 5′-d(GC), 5′-d(TA), 5′-d(CGCG), or 5′-d(GCGC) sequences. DHMO-DNA cross-links were assessed by electrophoretic mobility shift assay (EMSA) of 32P endlabeled oligonucleotides and by HPLC analysis of cross-linked DNAs enzymatically digested to their constituent deoxynucleosides. The degree of DNA cross-links depended upon the concentration of the pyrrole, but not on the base sequence of the oligonucleotide target. Likewise, HPLC chromatograms of cross-linked and digested DNAs showed no discernible sequence preference for any nucleotide. Added glutathione, tyrosine, cysteine, and aspartic acid, but not phenylalanine, threonine, serine, lysine, or methionine competed with DNA as alternate nucleophiles for cross-linking by DHMO. From these data it appears that DHMO exhibits no strong base preference when forming cross-links with DNA, and that some cellular nucleophiles can inhibit DNA cross-link formation.

Keywords: dehydromonocrotaline; DNA cross-linking; sequence specificity

Journal Article.  4232 words.  Illustrated.

Subjects: Medical Toxicology ; Toxicology (Non-medical)

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