A SHORT PURINE OLIGONUCLEOTIDE FORMS A HIGHLY STABLE TRIPLE-HELIX WITH THE PROMOTER OF THE MURINE C-PIM-1 PROTOONCOGENE

A homopurine - homopyrimidine region of murine c-pim-1 proto-oncogene was chosen as a target for triple-helix-forming oligonucleotide. Oligo nucleotide 5'-GGGGAGGGGGAGG-3' was shown to bind to its target sequenc e in the presence of 50 mM Na+ or K+, 10 mM MgCl2 and 20 mM Tris-aceta te, pH 7.5. This oligonucleotide is bound in an antiparallel orientati on with respect to the homopurine sequence. As was shown by co-migrati on assay the tripler is stable up to 65 degrees C. At 37 degrees C it was practically irreversible: after 24 hours of co-migration assay the re was no traces of tripler dissociation. The rate of tripler formatio n was highly accelerated with increase of temperature and Mg2+ concent ration. This rate was higher for superhelical DNA when compared to the linear and circular ones and the preference was dependent from temper ature and Mg2+ concentration. The precision of this interaction is ext remely high: sequences in c-pim-l promoter region with only one substi tution when compared to the target gave negligible tripler formation u nder investigated conditions. These data suppose that natural tripler structures could play an important role in eukaryotic gene regulation and/or chromatin structure formation.