Dietary restriction (DR) increases lifespan in a range of evolutionarily distinct species. The polyphenol resveratrol may be a dietary mimetic of some effects of DR. The pivotal role of the mammalian histone deacetylase (HDAC) Sirt1, and its homologue in other organisms, in mediating the effects of both DR and resveratrol on lifespan/ageing suggests it may be the common conduit through which these dietary interventions influence ageing. We propose the novel hypothesis that effects of DR relevant to lifespan extension include maintenance of DNA methylation patterns through Sirt1-mediated epigenetic effects, and proffer the view that dietary components, including resveratrol, may mimic these actions.

Epigenetic changes may be causal in the ageing process and may be influenced by diet, providing opportunities to improve health in later life. The aim of this review is to provide an overview of several areas of research relevant to this topic and to explore a hypothesis relating to a possible role of epigenetic effects, mediated by sirtuin 1, in the beneficial effects of dietary restriction, including increased lifespan. Epigenetic features of ageing include changes in DNA methylation, both globally and at specific loci, which differ between individuals. A major focus of research on dietary influences on epigenetic status has been on nutrition in utero, because the epigenome is probably particularly malleable during this life-course window and because epigenetic marking by early exposures is a compelling mechanism underlying effects on lifelong health. We explore the potential of diet during adulthood, including the practice of dietary restriction, to affect the epigenetic architecture. We report progress with respect to deriving data to support our hypothesis that sirtuin 1 may mediate some of the effects of dietary restriction through effects on DNA methylation and note observations that resveratrol affects DNA methylation and other epigenetic features. Disentangling cause and effect in the context of epigenetic change and ageing is a challenge and requires better understanding of the underlying mechanisms and also the development of more refined experimental tools to manipulate the epigenetic architecture, to facilitate hypothesis-driven research to elucidate these links and thus to exploit them to improve health across the full life-course through dietary measures.

Changes in DNA methylation across the life course may contribute to the ageing process. We hypothesised that some effects of dietary restriction to extend lifespan and/or mitigate against features of ageing result from changes in DNA methylation, so we determined if genes that respond to dietary restriction also show age-related changes in DNA methylation. In support of our hypothesis, the intersection of lists of genes compiled from published sources that (1) were differentially expressed in response to dietary restriction and (2) showed altered methylation with increased age was greater than expected. We also hypothesised that some effects of Sirt1, which may play a pivotal role in beneficial effects of dietary restriction, are mediated through DNA methylation. We thus measured effects of Sirt1 overexpression and knockdown in a human cell line on DNA methylation and expression of a panel of eight genes that respond to dietary restriction and show altered methylation with age. Six genes were affected at the level of DNA methylation, and for six expressions were affected. In further support of our hypothesis, we observed by DNA microarray analysis that genes showing differential expression in response to Sirt1 knockdown were over-represented in the complied list of genes that respond to dietary restriction. The findings reveal that

BackgroundSIRT1 is likely to play a role in the extension in healthspan induced by dietary restriction. Actions of SIRT1 are pleiotropic, and effects on healthspan may include effects on DNA methylation. Polycomb group protein target genes (PCGTs) are suppressed by epigenetic mechanisms in stem cells, partly through the actions of the polycomb repressive complexes (PRCs), and have been shown previously to correspond with loci particularly susceptible to age-related changes in DNA methylation. We hypothesised that SIRT1 would affect DNA methylation particularly at PCGTs. To map the sites in the genome where SIRT1 affects DNA methylation, we altered SIRT1 expression in human intestinal (Caco-2) and vascular endothelial (HuVEC) cells by transient transfection with an expression construct or with siRNA. DNA was enriched for the methylated fraction then sequenced (HuVEC) or hybridised to a human promoter microarray (Caco-2).ResultsThe profile of genes where SIRT1 manipulation affected DNA methylation was enriched for PCGTs in both cell lines, thus supporting our hypothesis. SIRT1 knockdown affected the mRNA for none of seven PRC components nor for DNMT1 or DNMT3b. We thus find no evidence that SIRT1 affects DNA methylation at PCGTs by affecting the expression of these gene transcripts. EZH2, a component of PRC2 that can affect DNA methylation through association with DNA methyltransferases (DNMTs), did not co-immunoprecipitate with SIRT1, and SIRT1 knockdown did not affect the expression of EZH2 protein. Thus, it is unlikely that the effects of SIRT1 on DNA methylation at PCGTs are mediated through direct intermolecular association with EZH2 or through effects in its expression.ConclusionsSIRT1 affects DNA methylation across the genome, but particularly at PCGTs. Although the mechanism through which SIRT1 has these effects is yet to be uncovered, this action is likely to contribute to extended healthspan, for example under conditions of dietary restriction.Electronic supplementary materialThe online version of this article (doi:10.1186/s40246-015-0036-0) contains supplementary material, which is available to authorized users.

Dietary restriction (DR) remains one of the most robust dietary interventions proved effective to increase lifespan across evolutionarily distinct species, from yeast to rodents (1) . The NAD-dependent (class III) histone deacetylase Sirt1 in mammals, and its orthologue in other species, may be pivotal in mediating the effect of DR to increase lifespan. Ageing is associated with changes in genome methylation (e.g.(2) ), which may be causative in the ageing process. Such observations, in view of the activity of Sirt1 at the level of the epigenome, form the basis for our hypothesis that epigenetic effects of Sirt1 activity mediate some of the beneficial effects of DR that contribute to lifespan extension.Our approach to testing this hypothesis was to determine the overlap between genes that bind Sirt1, those regulated at the mRNA level in response to DR and those that show an age-related change in methylation status, predicted on the principle that overlaps greater than would be expected by chance would support our hypothesis.By representing all genes using ENSEMBL identifiers, we identified overlaps between a list of 418 genes identified as interacting physically with the histone deacetylase Sirt1(3) , a list of 2954 genes found to respond to DR, compiled from 11 published studies, and a list of 1040 genes whose DNA methylation status changed in mouse small intestine in association with ageing (2) . All overlaps were found to be greater than would be expected by chance using the principle of hypergeometric distribution, as shown in Table 1. The promoter regions of 80 of the 91 genes that responded to DR and also bound Sirt1 (i.e. 88 %) were found to include CpG islands (defined as having a GC content of greater that 50 % and CpG observed/expected ratio of greater than 0.6 (5) ), representing sites of potential differential methylation in ageing tissues; this proportion is also greater than would be expected by chance (representation factor = 1.47, P = 3.395 · 10 -9 ), based on the assumption that 60 % (4) of mouse genes have associated CpG islands. Table 1. Overlaps between genes responsive to DR, genes that bind Sirt1 and genes that show changes in methylation status with ageing, based on in silico analysis. 'n' indicates the number of genes common to both groups; RF indicates the representation factor (i.e. observed/expected); P indicates the hypergeometric probability Genes binding Sirt1 (418 genes) Genes that show methylation changes with ageing (1040 genes) Genes responsive to DR (2954) n 91; RF = 1.60; P = 2.695 · 10 -6 n 191; RF = 1.27; P = 5.47 · 10Genes binding Sirt1 (418 genes) n 37; RF = 2.05; P = 1.66 · 10Overall, this in silico analysis supports our hypothesis that Sirt1-mediated effects of DR include effects on DNA methylation. Future studies should examine directly the extent to which DR and/or Sirt1 expression influence genome methylation status and should determine if such effects are causal with respect to ageing and/or lifespan extension.

Energy restriction is the only known robust dietary measure effective in increasing lifespan across a wide variety of species. Our current research indicates a role for the best-established dietary mimetic of energy restriction, the red grape polyphenol resveratrol, in altering information carried by DNA at a level superimposed on the information provided by the sequence of bases, known as epigenetic marking. These effects appear to share a common pathway with some of the effects of energy restriction that involve the protein Sirt1. Preliminary observations made in human intestinal cells reveal that isoflavones found in the soyabean may share some of these functional properties and so highlight the potential for a diet rich in these compounds to promote healthy ageing.

We aimed to determine if epigenetic effects could be a component of the mechanism through which the dietary polyphenol resveratrol brings about its reported beneficial effects on healthspan and if such effects are though activation of SIRT1, the histone deacetylase purported to play a role in lifespan extension resulting from dietary restriction (DR). Resveratrol reduced histone expression in human intestinal Caco‐2 cells through an effect on histone gene expression (opposing an aging‐related observed increase in mouse intestine) and increased DNA methylation. The ER antagonist fulvestrant abrogated the effect on histone expression. SIRT1 overexpression alone had no effect on histone expression or DNA methylation but overexpression abrogated both effects of resveratrol. Genes with altered expression resulting from resveratrol treatment or SIRT1 knockdown did not overlap but both sets of genes showed an overlap greater than expected by chance with genes affected by DR. The findings indicate that resveratrol has epigenetic effects that are mediated through the ER and are independent of SIRT1 and that such actions may contribute to effects of resveratrol to increase healthspan.