No abstract

Monocrotaline is a pyrrolizidine alkaloid known to cause toxicity in humans and animals. Its mechanism of biological action is still unclear although DNA crosslinking has been suggested to a play a role in its activity. In this study we found that an active metabolite of monocrotaline, dehydromonocrotaline (DHM), alkylates guanines at the N7 position of DNA with a preference for 5'-GG and 5'-GA sequences. In addition, it generates piperidine- and heat-resistant multiple DNA crosslinks, as confirmed by electrophoresis and electron microscopy. On the basis of these findings, we propose that DHM undergoes rapid polymerization to a structure which is able to crosslink several fragments of DNA.

Bracken fern (Pteridjum spp.) causes cancer of the oesophagus and the urinary bladder in cattle and sheep. Ptaquiloside (PT) is believed to b e the carcinogenic principle which alkylates DNA when activated to its unstable dienone form (APT) under alkaline conditions. In this report w e present evidence for the presence of PT-DNA adducts in the ileum of bracken fern-fed calves using the 3PP-postlabelling assay. H-ras mutations were also observed in the ileum using single strand conformation polymorphism (SSCP) technique. Mutations corresponding to adenine to pyrimidine transversions in the codon 61 of H-ras were identified b y the cycle sequencing method. In vitro DNA alkylation studies showed that APT alkylated H-ras primarily at the adenines. In addition, the rate of depurination of alkylated adenine was sequence dependent. Investigation of DNA template activity using a plasmrd DNA showed that DNA synthesis by T7 DNA polymerase was terminated b y the presence of all alkylated bases but certain apurinic sites allowed the DNAsynthesis to continue. These results suggest that initial alkylation of adenine b y PT in codon 61 followed by depurination and error in DNA synthesis lead to activation of H-ras proto-oncogene. o 1996 W~lq-L~ss, Inc