AC-motif: a DNA motif containing adenine and cytosine repeat plays a role in gene regulation

I-motif or C4 is a four-stranded DNA structure with a protonated cytosine:cytosine base pair (C+:C) found in cytosine-rich sequences. We have found that oligodeoxynucleotides containing adenine and cytosine repeats form a stable secondary structure at a physiological pH with magnesium ion, which is similar to i-motif structure, and have named this structure ‘adenine:cytosine-motif (AC-motif)’. AC-motif contains C(+):C base pairs intercalated with putative A(+):C base pairs between protonated adenine and cytosine. By investigation of the AC-motif present in the CDKL3 promoter (AC-motif(CDKL3)), one of AC-motifs found in the genome, we confirmed that AC-motif(CDKL3) has a key role in regulating CDKL3 gene expression in response to magnesium. This is further supported by confirming that genome-edited mutant cell lines, lacking the AC-motif formation, lost this regulation effect. Our results verify that adenine-cytosine repeats commonly present in the genome can form a stable non-canonical secondary structure with a non-Watson–Crick base pair and have regulatory roles in cells, which expand non-canonical DNA repertoires.