USE OF OLIGONUCLEOTIDES TO DEFINE THE SITE OF INTERSTRAND CROSS-LINKS INDUCED BY ADRIAMYCIN

It has been known for several years that Adriamycin forms adducts and interstrand cross-links when reacted for long periods of time with bacterial and mammalian DNA in vitro, with the cross-link being restricted to 2 bp elements containing GpC sequences, The self-complementary 20mer deoxyoligon ucleotide TA(4)T(4)GCA(4)T(4)A has been used in this study as a model of the apparent G-G cross-linking site at GpC sequences, The rate of formation of cross-links, as well as the dependence on both Adriamycin and Fe(III) concentration, were similar with this oligonucleotide as compared with calf thymus DNA, The cross-linking was demonstrated on both denaturing and non-denaturing sequencing gels, The half-life of the G-G cross-link was 40 h, consistent with that implied with high molecular weight, heterogeneous sequence DNA. Exonuclease III digests of adducts formed with 20mer deoxyoligonucleotides containing single, central G-G, G-I and I-I potential cross-links revealed that a guanine residue is required at both ends of the cross-link. No cross-linking was observed with a similar oligonucleotide containing only a single central (G . C) bp.