Differential impacts of DNA methylation on pH dependent i-motifs formation in rice [Bisulfite-seq]
I-motifs (iMs) are four-stranded non-B DNA structures containing C-rich DNA sequences, which can be formed under various pH conditions. DNA methylation exhibits complex impacts on iM formation in vitro. However, how DNA methylation differentially affects pH dependent iM formation on a genome wide scale is completely uncharacterized in both humans and plants. To this end, we conducted iM-IP-seq under pH 5.5 and 7.0 conditions using CK and hyper/hypomethylated DNA in combination with BS-seq in rice. We found that pH 7.0 and 5.5 biased iMs had distinct genomic features and differential DNA methylation levels, the former having higher DNA methylation levels than the latter. Moreover, DNA demethylation and hyper methylation had more impacts on a subset of iM formation under pH 7.0 and 5.5, respectively, indicating that DNA de/hyper-methylation exhibits pH dependent impacts on iM formation. Importantly, we found that CG hypo-DMRs and CHH hyper-DMRs alone or coordinated with CG/CHG hyper-DMRs may play determinant roles in the regulation of iM formation under pH 5.5 and 7.0. Thus, our study shows that intrinsic DNA sequences alone or in combination with DNA methylation play vital roles in determining pH-dependent formation of iMs. It will contribute to in-depth understanding of DNA methylation in the modulation of pH dependent dynamics of iM conformation, therefore broadening its biological implications and practical application ranges. Overall design: M. Cvip I-(hypermethylation)/zebularine (demethylation)-treated genomic DNA and CK (untreatment) were used for bisulfite treatment