Triple‐helix formation interferes with the transcription and hinged DNA structure of the interferon‐inducible 6–16 gene promoter

Abstract

The interferon responsive element (IRE) of the 6–16 gene lies within two 39‐bp elements in tandem. A purine‐rich oligodeoxynucleotide, oligo(dN), was found to be able to pair with the purinerich strand of the IRE in an antiparallel orientation which led to triple‐helix formation with Mg2+ being necessary for triplex stability. Footprinting analysis confirmed these results. The interaction between the IRE and the oligo(dN) was reversible and had a Kd equal to 20 nM. The two repeats of the 6–16 gene IRE can form a hinged DNA structure through pairing of their purine‐rich regions; exonuclease III experiments support this model. The hybrid DNA structure leads to a parallel pairing of the purine strands of the 6–16 gene IRE and this conformation was shown to be destabilized by triplex formation. When co‐transfected with a reporter gene whose promoter was under the control of the 6–16 gene IRE, the triple‐helix‐forming oligo(dN)s inhibit the interferon‐induced stimulation of the reporter gene with complete inhibition being obtained with 1 μM oligo(dN) at the time of transfection. When added to the cell culture medium after transfection, the concentrations of oligo(dN) needed to obtain 50% inhibition of the interferon effect on gene transcription must be 50–100 times higher. Besides the existence of a peculiar structure for the 6–16 gene IRE, the possibility of interfering with gene expression by means of oligo(dN)s is demonstrated. Copyright © 1994, Wiley Blackwell. All rights reserved