Genome-wide DNA methylation patterns in Daphnia magna are not significantly associated with age

Background: DNA methylation plays a crucial role in gene regulation and epigenetic inheritance across diverse organisms. Daphnia magna, a model organism in ecological and evolutionary research, has been widely used to study environmental responses, pharmaceutical toxicity, and developmental plasticity. However, its DNA methylation landscape and age-related epigenetic changes remain incompletely understood. Results: In this study, we characterized DNA methyltransferases (DNMTs) and mapped DNA methylation across the D. magna genome using whole-genome bisulfite sequencing. Our analysis identified three DNMTs: a highly expressed but nonfunctional de novo methyltransferase (DNMT3.1), alongside lowly expressed yet functional de novo methyltransferase (DNMT3.2) and maintenance methyltransferase (DNMT1). D. magna exhibits overall low DNA methylation, targeting primarily CpG dinucleotides. Methylation is sparse at promoters but elevated in the first exons downstream of transcription start sites, with these exons showing hypermethylation relative to adjacent introns. To examine age-associated DNA methylation changes, we analyzed D. magna individuals across multiple life stages. Our results showed no significant global differences in DNA methylation levels between young, mature, and old individuals, nor any age-related clustering in dimensionality reduction analyses. Attempts to construct an epigenetic clock using machine learning models did not yield accurate age predictions, likely due to the overall low DNA methylation levels and lack of robust age-associated methylation changes. Conclusions: This study provides a comprehensive characterization of D. magna’s DNA methylation landscape and DNMT enzymes, highlighting a distinct pattern of exon-biased CpG methylation. Contrary to prior studies, we found no strong evidence supporting age-associated epigenetic changes, suggesting that DNA methylation may have a limited role in aging in D. magna. These findings enhance our understanding of invertebrate epigenetics and emphasize the need for further research into the interplay between DNA methylation, environmental factors, and gene regulation in D. magna.