Characterisation of DNA i-motif structures in the human genome and the role of non-canonical nucleic acid structures

Genomic research stands as a cornerstone of biological sciences, constantly improving our understanding of the essential mechanisms in life. Genomic structural architecture, fundamentally constructed by nucleic acids, has been observed in diverse forms. Varying in nucleic acid composition, such as DNA and RNA, or by structure, such as linear or circular structures. In the complex variety of nucleic acid forms, non-canonical nucleic acid structures have emerged as a research frontier, hence their unique properties compared to their more stable double-helix or single-stranded counterparts. These non-canonical structures include triplexes, quadruplexes, or Z-DNA, to name a few, which have been observed to affect biological roles. Understanding these structures and harnessing their unique properties is a promising field not only in basic genomic research but also in synthetic biotechnology. DNA quadruplexes, also known as tetraplexes, are structures rising from the interaction of four nucleic acid units. The two known DNA tetraplexes are G-quadruplexes and i-Motifs. G-quadruplexes rise from the interaction of four guanine bases and multiple tetrads stacked on top of each other, and i-Motifs rise from the intercalated interaction of cytosine base pairs, essentially binding four parts of a single strand together. By utilising the newly developed high-affinity antibody and conducting immunoprecipitation followed by sequencing to map i-motifs in the genomic DNA, we have mapped the possible forming locations. We validated these immunoprecipitated sequences, mapped locations by biophysical forms and conducted comparative analyses in diverse environmental and experimental conditions. We also screened for human nuclear proteins interacting with i-motifs to catalogue their interactome. We showcased the effects of the stabilisation quadruplexes in cell culture to investigate their regulatory roles in gene transcription. Additionally, we did not only venture into the observation of non-canonical nucleic acids. Through the identification and use of unique antibodies targeting quadruplexes, we characterised an artificially modified xeno-nucleic acid aptamer that distinctively recognises HEL and contains a G-quadruplex-like structure in the core. This novel structural study highlights the unique properties of non-canonical nucleic acid structures, paving the way for novel designs and applications.

Grantor dc:publisher

UNSW, Sydney

Year dc:date

2025

Author dc:creator

• Pena Martinez, Cristian

• DNA
• nucleic acid
• human genome
• i-motif
• non-canonical nucleic acids
• anzsrc-for: 310509 Genomics
• anzsrc-for: 320505 Medical biochemistry - nucleic acids
• anzsrc-for: 310606 Industrial molecular engineering of nucleic acids and proteins
• anzsrc-for: 320603 Medical molecular engineering of nucleic acids and proteins
• anzsrc-for: 320402 Applied immunology (incl. antibody engineering, xenotransplantation and t-cell therapies)

Statement dc:rights

• embargoed access
• CC BY 4.0

Licence

• http://purl.org/coar/access_right/c_f1cf
• https://creativecommons.org/licenses/by/4.0/

Language dc:language

en

Identifier

https://doi.org/10.26190/unsworks/30924

OAI identifier oai:identifier

oai:unsworks.library.unsw.edu.au:1959.4/104271