Homeostasis of DNA Hemi‐Methylation in Arabidopsis through Methylation Maintenance, DNA Replication, and Nucleosome Positioning Mechanisms

Abstract The DNA methylome in eukaryotes is an equilibrium contributed by maintenance methylation, de novo methylation, demethylation, and DNA replication processes. The asymmetric hemi‐methylation status is thought of as a transient intermediate state en route to full‐methylation. However, the recent studies in mammalian cells suggest that hemi‐methylated CG dyads in certain regions can be stably maintained and may serve as epigenetic marks. Compared to mammals, plants have more diversified methylomes often including non‐CG dyads. In this work, hairpin BS‐seq is performed to acquire dyad‐resolution methylomes from Arabidopsis wildtype and methylation‐deficient mutant lines. Analyses of methylomes across multiple lines reveal how different DNA methyltransferases influence the equilibrium of hemi‐ and full‐methylation in vivo. The results suggest that nucleosomes may protect hemi‐methylated dyads from being further methylated, resulting in relatively higher hemi‐methylation frequencies in nucleosome‐occupied than ‐depleted regions. Adjacent hemi‐methylated dyads tend to have a strong strand‐specificity, indicating that DNA replication is a driving force for the homeostasis of hemi‐methylation. Overall, comprehensive multi‐line and dyad‐resolved DNA methylation maps in Arabidopsis is presented to reveal that the homeostasis of DNA hemi‐methylation in plants is achieved through diverse methylation regulatory and chromatin‐related activities.