Identification of multiple genomic DNA sequences which form i-motif structures at neutral pH.
i-Motifs are alternative DNA secondary structures formed in cytosine-rich sequences. Particular examples of these structures, traditionally assumed to be stable only at acidic pH, have been found to form under near-physiological conditions. To determine the potential impact of these structures on physiological processes, investigation of sequences with the capacity to fold under physiological conditions is required. Here we describe a systematic study of cytosine-rich DNA sequences, with varying numbers of consecutive cytosines, to gain insights into i-motif DNA sequence and structure stability. i-Motif formation was assessed using ultraviolet spectroscopy, circular dichroism and native gel electrophoresis. We found that increasing cytosine tract lengths resulted in increased thermal stability; sequences with at least five cytosines per tract folded into i-motif at room temperature and neutral pH. Using these results, we postulated a folding rule for i-motif formation, analogous to (but different from) that for G-quadruplexes. This indicated that thousands of cytosine-rich sequences in the human genome may fold into i-motif structures under physiological conditions. Many of these were found in locations where structure formation is likely to influence gene expression. Characterization of a selection of these identified i-motif forming sequences uncovered 17 genomic i-motif forming sequence examples which were stable at neutral pH.
School of Pharmacy, University of East Anglia, Norwich Research Park, Norwich NR4 7TJ, UK.
Elisé P
EP
Wright
Intellectual Forum, Jesus College, University of Cambridge, Cambridge CB5 8BL, UK.
Julian L
JL
Huppert
School of Pharmacy, University of East Anglia, Norwich Research Park, Norwich NR4 7TJ, UK.|Centre for Molecular and Structural Biochemistry, University of East Anglia, Norwich Research Park, Norwich NR4 7TJ, UK.
Zoë A E
ZAE
Waller
D001483:Base Sequence; D003596:Cytosine / Q000737:chemistry; D004247:DNA / Q000737:chemistry*; D015894:Genome, Human; D006801:Humans; D006863:Hydrogen-Ion Concentration; D009690:Nucleic Acid Conformation; D016615:Telomere / Q000737:chemistry; D013696:Temperature
D003596:Cytosine; D004247:DNA