REPAIR OF TRIPLE-HELIX DIRECTED PSORALEN ADDUCTS IN HUMAN-CELLS

Triple helix forming oligonucleotides can direct DNA damaging agents a t specific sites in an intact double helix. In our study, triple helix formation was demonstrated in a SV40 based shuttle vector treated wit h psoralen linked to a 22-mer purine rich oligonucleotide. UVA irradia tion caused a covalent linkage of the oligonucleotide through the psor alen to the mutational supF marker gene of the plasmid. After passage in the Jurkat human cell line the recovered vector was analysed in an indicator bacterial strain and mutants were collected. The presence of adducts in the target sequence did not reduce the yield of replicated progeny vector molecules, indicating repair of triple helix associate d monoadducts and cross-links. Mutations were highly targeted to a six nucleotide long region of the target sequence. The number of target s equence mutants obtained after triple helix directed psoralen treatmen t was approximately 160 times higher than with free psoralen. A furthe r investigation of the exact mechanism of the mutational process could make triple helix directed mutagenesis a more useful tool in gene the rapy, antiviral therapy, and in studies on DNA repair and genome organ isation.