Molecular insights into nucleic acid triplexes: methodologies and biological applications

Triplex DNA structures form through the binding of a third oligonucleotide strand to the major groove of canonical double-stranded DNA at sites of extended polypurine sequence. Although they are known to be favored with certain sequence specificity and cellular conditions, including decreased pH and the presence of multivalent cations, there remains ambiguity in the structures and extent to which they form in vivo. Therefore, despite their biological relevance and many potential applications, the use of DNA triplexes in biotechnology has been limited to date. The focus of this review is to explore the intricacies of DNA triplex formation, as well as the current state of research into their functions and applications in molecular cell biology. The range of analytical, computational and synthetic chemistry techniques employed to investigate and enhance the stability of triplex assemblies is also reviewed. Understanding the structural properties that underpin triplex formation and activity, coupled with computational and synthetic methodologies to expand their utility, can unlock the potential of various triplex-forming oligonucleotides as a contemporary tool for regulating gene expression.