DNA Methylation Developmental Programming and Cellular Memory: The Molecular Consequences of Folate Depletion In Utero

Summary Poor maternal nutrition can have profound effects on the long-term health and wellbeing of offspring and may cause premature ageing. These adverse effects on health include common diseases such as obesity, type 2 diabetes and cardiovascular disease. The risk of these conditions appears to be exacerbated when the offspring gain weight more quickly than would have been expected. In preliminary studies, we observed that mice born to mothers given diets low in folate (a B vitamin) during pregnancy and lactation became heavier adults than those born to mothers with normal folate supply despite the fact that there were no differences in body weight between the two groups of mice at weaning and that all mice received the same diet from weaning. This suggests that the maternal nutritional insult was 'remembered' by the mouse tissues and expressed as more rapid growth. Change in epigenetic markings is one of the most important mechanisms believed to be responsible for cellular 'memorisation' of early life experiences. Epigenetics describes chemical changes to the genome which regulate when and where genes are expressed (turned on) but which do not alter the primary DNA sequence. The best understood of the epigenetic marks is DNA methylation i.e. the addition of CH3 (methyl) groups to DNA. In our mouse studies we found that there were fewer CH3 groups in the DNA from adult mice born to the folate-depleted mothers. In follow up studies we have found that maternal folate depletion resulted in altered expression of over 600 genes in fetal liver. In this project we will investigate in more detail the effects of low maternal folate supply on growth and body fatness of the offspring and we will examine the effects of feeding high v. low fat diets from weaning. We anticipate that the high fat diet (based on the composition of Western human diets) will exacerbate the effects of the maternal nutritional insult. We will use state-of-the-art magnetic resonance imaging to allow us to measure how much fat is in the live mice and in which parts of the body it is stored. In humans, fat stored in the abdomen is associated with higher risk of several common disease including type 2 diabetes, cardiovascular disease and bowel cancer. We will then explore the mechanisms responsible for these phenotypic changes. We will focus on identification of genes whose expression has been changed through altered epigenetic markings and attempt to elucidate the molecular events leading to the genes being switched off, or switched on, inappropriately.

An accumulating body of evidence supports the premise that nutritional deprivation in utero has lifelong consequences for health, including a predisposition to obesity, cardiovascular disease and type 2 diabetes, particularly in association with a plentiful food supply post-weaning. Thus, it is presumed that nutritional status in utero is recorded and remembered in a manner that programmes the foetus to use nutrients in a way that improves survival when the nutrient supply is poor but to be adapted inappropriately to a plentiful or excessive intake. Epigenetic processes, including DNA methylation, provide a compelling mechanism for such effects, manifest through influences on gene expression. Our preliminary data reveal that folate depletion in utero in the mouse did not affect weight at weaning but increased the weight of offspring at 100 d and induced global DNA hypomethylation in liver and affected expression of multiple genes in foetal liver. These observations provide the impetus to examine in detail the influence of this nutritional stress on adiposity and on the methylation and expression of specific genes. Our hypotheses are that i) folate depletion in utero predisposes mice to increased weight and abdominal adiposity in adulthood, exacerbated by a high-fat post-weaning diet and ii) the effect is mediated through the methylation of specific genes, which affects expression through transcription factor binding. These hypotheses will be investigated through measuring the effect of folate depletion in utero in mice, followed by a control or high-fat post-weaning diet, on weight gain and adiposity and by examining effects at late gestation on DNA methylation in liver, using a microarray-based approach, and on gene expression and DNA methylation at 180 d. Effects of promoter methylation on gene expression and transcription factor binding will be investigated for selected differentially methylated and expressed genes using in vitro molecular approaches.