In vitro transcription analysis of DNA adducts induced by cyanomorpholinoadriamycin.
The reaction of cyanomorpholinoadriamycin (CMA) with DNA results in the formation of sequence-specific complexes with DNA. These complexes were revealed as blocked transcripts in an in vitro transcription assay--of 14 high-intensity blockages detected in the 120 bp probed in this assay, 12 were prior to GpG or CpC sequences. Slow read-through past the first few sites exhibited first-order kinetics, with half-lives of 25-200 min. Bidirectional transcription footprinting revealed nine high-intensity sites, eight of which were defined by a GpG element (nontemplate strand). Reaction of CMA with single-strand DNA, followed by a primer-extension assay, revealed four major blockages all of which were at GpG sites on the initial single-strand DNA. From a combination of these three experimental approaches, it appears that CMA yields dominantly intrastrand cross-links between adjacent guanine residues. Since CMA is also known to form interstrand cross-links, these appear to occur at GpC sequences but are minor in comparison to the extent of formation of intrastrand cross-links.
Cullinane
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Department of Biochemistry, La Trobe University, Bundoora, Victoria, Australia.
Phillips
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D000478:Alkylation; D001483:Base Sequence; D003432:Cross-Linking Reagents / Q000494:pharmacology*; D004247:DNA / Q000737:chemistry / Q000187:drug effects* / Q000235:genetics; D016172:DNA Fingerprinting; D004277:DNA, Single-Stranded / Q000737:chemistry; D015226:Dinucleoside Phosphates / Q000737:chemistry; D004317:Doxorubicin / Q000031:analogs & derivatives* / Q000494:pharmacology; D007700:Kinetics; D008969:Molecular Sequence Data; D014158:Transcription, Genetic
D003432:Cross-Linking Reagents; D004277:DNA, Single-Stranded; D015226:Dinucleoside Phosphates; C015101:cytidylyl-(3'-5')-cytidine; C018309:guanylyl-(3'-5')-guanosine; D004317:Doxorubicin; C038555:3'-deamino-3'-(3-cyano-4-morpholinyl)doxorubicin; D004247:DNA