Echinomycin, a bis-intercalating agent, induces C-->T mutations via cytosine deamination.
Echinomycin, a bis-intercalating, antitumor drug, has been studied for its ability to induce the deamination of cytosine to uracil (C-->U) in double-stranded DNA. We have employed a sensitive lacZ alpha-complementation reversion assay to detect G.C-->A.T mutations at a number of sites in M13mp2 DNA to determine the extent to which distortions of DNA structure induced by echinomycin may affect C-->U rates. When double-stranded M13mp2 DNA with a 12-base target containing a CpG site was incubated at 37 degrees C, the reversion frequency of the echinomycin-treated DNA increased linearly over time, with a rate constant 3-fold greater than DNA incubated without echinomycin. Of the 11 ways that blue pseudo-revertants can occur in the target, 96% of the observed revertants arose from C-->T and tandem CC-->TT transitions, with 78% attributable to single-base C-->T changes at three sites. Transfection into ung+ cells decreased the reversion frequencies by 85% to near background levels, indicating that the increase in C-->T mutations was due to deamination of C to U. The cytosine deamination rate constants for the entire target at pH 6.0 and 37 degrees C were 1.2 x 10(-11) sec-1 for untreated DNA and 3.5 x 10(-11) sec-1 for echinomycin-treated DNA. The increase in C-->T mutation rates occurred at cytosines both proximal and distal to a CpG echinomycin-binding site. We hypothesize that this increase in deamination rate is due to a more open or single-stranded DNA structure caused by the echinomycin: DNA interaction.
Moyer
R
R
Department of Chemistry, Duke University, Durham, NC 27708-0346.
Briley
D
D
Johnsen
A
A
Stewart
U
U
Shaw
B R
BR
D001483:Base Sequence; D002460:Cell Line; D003596:Cytosine / Q000378:metabolism*; D003641:Deamination; D004448:Echinomycin / Q000494:pharmacology*; D007364:Intercalating Agents / Q000494:pharmacology*; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D013696:Temperature; D014162:Transfection
D007364:Intercalating Agents; D004448:Echinomycin; D003596:Cytosine