I-motif DNA structures are formed in the nuclei of human cells

Abstract

Human genome function is underpinned by the primary storage of genetic information in canonical B-form DNA, with a second layer of DNA structure providing regulatory control. I-motif structures are thought to form in cytosine-rich regions of the genome and to have regulatory functions; however, in vivo evidence for the existence of such structures has so far remained elusive. Here we report the generation and characterization of an antibody fragment (iMab) that recognizes i-motif structures with high selectivity and affinity, enabling the detection of i-motifs in the nuclei of human cells. We demonstrate that the in vivo formation of such structures is cell-cycle and pH dependent. Furthermore, we provide evidence that i-motif structures are formed in regulatory regions of the human genome, including promoters and telomeric regions. Our results support the notion that i-motif structures provide key regulatory roles in the genome.

Analysis of Interactions between Telomeric i‐Motif DNA and a Cyclic Tetraoxazole Compound

“…The complementary sequence of a G4, thus enriched in cytosines, is known to form the i‐motif structure under acidic conditions in vitro, through the interaction of hemiprotonated cytosine–cytosine (C−C) base pairs (Scheme A and B) . Although this structure was first recognised in 1993, its biological functions are still poorly understood, partly because of the lack of selective and potent ligands . In the first study of a small molecule binding to i‐motif in 2000, the group of Hurley examined the interaction between the telomeric i‐motif and TMPyP4, which is a porphyrin‐type G4‐stabilising ligand .…”

On the nature of ion‐stabilized cytosine pairs in DNA i‐motifs: The importance of charge transfer processes

“…Specifically cytosine‐ or guanine‐rich sequences form distinct and well‐defined higher‐order structures, which significantly differ from common helical B‐DNA conformations . A typical example are DNA i‐motifs with hemi‐protonated cytosine (C:H:C) + pairs (Figure ). In contrast to standard double‐stranded B‐DNA, cytosine pairs in i‐motifs at pH values around 4‐6 form a stabilizing third hydrogen bond due to the intercalation of a proton .…”

Dynamic DNA Structures

“…DNA strands with such capability are designated as DNA aptamers (Figure d) . Certain special structural motifs are reconfigurable as well, such as guanine‐ and cytosine‐rich sequences that can fold into stable quadruplexes in the presence of specific metal ions and low pH (Figure e) . Base stacking between blunt ends of DNA helices are ion‐ and temperature‐sensitive (Figure f), and are, therefore, able to reconfigure upon stimulation .…”