Elucidating The Biological Role Of i-Motif DNA Structures In Vertebrate Genomes

The i-motif is a non-standard DNA structure, very different to the typical Watson-Crick double-helix shape. It was previously believed that the i-motif structure was not biologically relevant because it appeared to demand an acidic environment not normally seen inside cells. However, recent evidence shows that some DNA sequences form i-motif structures at intra-cellular pHs and that they can be directly visualised in genomes of living cells.

Whilst a sequence-specific prediction of i-motifs has been proposed, this clearly over-estimates the number of i-motifs present in cells and misses potential i-motif forming sequences that do not follow this pattern precisely. In order to fully understand the role played by the i-motif, we must first define a complete map of their real distribution in the genome and the conditions under which they exist in living organisms.

This research project will generate the first experimentally validated map of i-motifs by making use of an antibody that specifically recognises i-motifs over other DNA structures. We will use a developing organism (the frog, Xenopus tropicalis) and link this to specific DNA features such as promoters and enhancers of genes, the aim being to define the functional context of these structures. We will explore the stability of i-motif structures at different stages of a growing tadpole, where during development DNA undergoes significant changes of a type known to affect i-motif formation.

Whilst the common structure of DNA is the Watson/Crick double-helix, it is now known that DNA can form alternative structures, each with an essential role in normal biological activity. Recent research has shown that the four-stranded ‘i-motif’ DNA structure can form under normal conditions in living cells including those of humans. However, the biological context in which i-motifs form and their biological role remains to be understood. To answer these questions, this project will explore this new level of genomic organisation in order to identify the role that i-motifs play in the mechanisms of gene regulation and other DNA transactions.