Inhibition of gene transcription by purine rich triplex forming oligodeoxyribonucleotides

Roy, C.

Nucleic Acids Res

1993-06-25 / vol 21 / pages 2845-52


Several oligodeoxynucleotides (ODNs) were designed in order to interact with the purine rich element of the IRE (Interferon Responsive Element) of the 6-16 gene by triplex formation. An ODN of 21 bases, the sequence being identical to that of the purine strand of the IRE (48% G), but in reverse orientation, was able to interact with the IRE (KD: 20 nM). The binding was Mg2+ dependent. The two purine strands of the triplex were oriented antiparallel as confirmed by DNAase I and copper-phenanthroline footprinting experiments. An ODN in which A were replaced by T, also interacted with the same target, but with a lower affinity. Exonuclease III action indicated that the two IRE repeats of the 6-16 promoter interacted with each other through Hoogsteen base pairing, the third strand being parallel to the paired Watson-Crick strand. This led to a potential H-DNA structure which could be destabilized by adding ODNs able to form a triplex structure. 6-16 IRE driven-reporter gene constructs lost their interferon stimulability when co-transfected with triplex forming ODNs. The range of effective ODN concentrations was compatible with the affinity determined when measuring their direct interactions with the DNA.

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Molecular Sequence Data; Base Sequence; Binding Sites; Structure-Activity Relationship; Transfection; *Nucleic Acid Conformation; Interferons/*pharmacology; Plasmids; Transcription, Genetic/*drug effects; *Purines/chemistry; Deoxyribonuclease I; DNA/chemistry/genetics/*metabolism; Magnesium/pharmacology; Oligodeoxyribonucleotides/chemistry/metabolism/*pharmacology; Phenanthrolines

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