Folding double-stranded DNA into designed shapes with triplex-forming oligonucleotides

Abstract

The folding of double-stranded DNA around histones is a central mechanism in eukaryotic cells for compacting the genetic information into chromosomes. Very few artificial methods are available for controlling the shape of dsDNA at any level, whereas several artificial methods have been developed to efficiently organize single-stranded DNA and RNA into a variety of well-defined nanostructures by programmed self-assembly. Here, we show how long double-stranded DNA sequences can be spatially organized by triplex-forming oligonucleotides (TFOs), which bridge two or more encoded polypurine domains. The linearized or plasmid dsDNA is compacted into antiparallel folds, which enables the formation of raster-filled 2D shapes and 3D structures with either square or hexagonal organizations. Contrary to ssDNA, dsDNA has inherent rigidity which alleviates the requirement to saturate a structure with TFO strands, yet the TFOs are still able to bend the dsDNA controllably and steeply up to 180° over 6 bp. The majority of structures investigated here are formed by Hoogsteen interactions which require pH = 5-6, however, the methodology is also applied with reverse Hoogsteen interactions at physiological pH. In both cases, the DNA triplexes render pure polypurine scaffolded structures resistant to DNase I.

Integration of DNA–RNA‐triplex‐based regulation of transcription into molecular logic gates

“…In the toolbox of synthetic biology, the formation of alternative, non‐duplex nucleic acid structures represents a valuable approach for the control of biochemical processes [1–3]. This is due to a number of reasons, including their involvement with biological targets such as gene promoters and endonuclease restriction sites [4,5], their tunability using specific stabilizing agents [6], and their compatibility with other natural or artificial single‐ and double‐stranded nucleic acid structures [7,8]. In addition, non‐canonical nucleic acid structures seem to play a role in cellular differentiation and cancer development via the regulation of gene expression [9].…”