Genome-wide methylation is modified by caloric restriction in Daphnia magna

Abstract

Background: The degradation of epigenetic control with age is associated with progressive diseases of ageing, including cancers, immunodeficiency and diabetes. Reduced caloric intake slows the effects of ageing and age-related disease in vertebrates and invertebrates, a process potentially mediated by the impact of caloric restriction on epigenetic factors such as DNA methylation. We used whole genome bisulphite sequencing to study how DNA methylation patterns change with diet in a small invertebrate, the crustacean Daphnia magna. Daphnia show the classic response of longer life under caloric restriction (CR), and they reproduce clonally, which permits the study of epigenetic changes in the absence of genetic variation.

Results: Global cytosine followed by guanine (CpG) methylation was 0.7-0.9%, and there was no difference in overall methylation levels between normal and calorie restricted replicates. However, 333 differentially methylated regions (DMRs) were evident between the normally fed and CR replicates post-filtering. Of these 65% were hypomethylated in the CR group, and 35% were hypermethylated in the CR group.

Conclusions: Our results demonstrate an effect of CR on the genome-wide methylation profile. This adds to a growing body of research in Daphnia magna that demonstrate an epigenomic response to environmental stimuli. Specifically, gene Ontology (GO) term enrichment of genes associated with hyper and hypo-methylated DMRs showed significant enrichment for methylation and acyl-CoA dehydrogenase activity, which are linked to current understanding of their roles in CR in invertebrate model organisms.

Keywords: Bisulphite sequencing; Caloric restriction; DNA methylation; Daphnia; Differential methylation; Epigenetics; Genomics; Nutrition.

Fig. 1

Dendrogram showing replicates cluster by mother and not by treatment in global CpG similarity. Mother is indicated by number, blue H: normal food and red L: calorie restricted replicate. Node support values in green were assessed by 10,000 bootstraps in pvclust, bootstrap probability and approximately unbiased methods had identical values. Dendrogram created using ward.D2 method in methylKit and pvclust. Number refers to mother from which that replicate was derived; H for normal food diet, L for caloric restriction

Fig. 2

Volcano plot of per-region FDR corrected p-values against difference in mean methylation between caloric restricted and normal food conditions. Red dots are DMRs that passed t-statistic filtering and had a region p-value below < 0.05, these were retained for further analysis. Pink dots are DMRs that passed t-statistic filtering but were not significant by corrected p-value. Black dots are none-significant DMRs by the chosen criteria, dashed grey vertical lines indicate the difference in means (0.1, − 0.1) required to accept a DMR along with a t-statistic of − 4.6 and 4.6 and 3 CpG sites, black dots with a p-value less than 0.05 and greater than [0.1] difference in means failed one of these two conditions. Horizontal grey dashed line indicates a p-value threshold of 0.05