The more we learn about our genome, the more mysteries arise.
For example, how can people with the same disease-causing mutation have different disease progression and symptoms?
And despite the fact that it’s been more than 15 years since the human genome was sequenced, why can’t we explain the significance of the vast majority of genomic variations that occur in noncoding, or “junk,” elements of the genome?
Now, Pier Lorenzo Puri, M.D., a professor in the Development, Aging, and Regeneration Program at Sanford Burnham Prebys, has used a cutting-edge technique called Hi-C, which maps millions of interactions between proteins and tightly coiled DNA, called chromatin, to shed light on this mystery.
The study, published in Molecular Cell, shows that a specific protein called MyoD – a master regulator of muscle development – reshapes chromatin’s architecture to alter gene expression – revealing fundamental insights into how genetic variations may affect our health.
“One of the greatest mysteries of medicine is how people with the same mutation can have different symptoms,” says Puri.
“Our study indicates that some genetic variations may affect our health by altering how DNA coils and interacts in its 3-D shape.
This alteration may be helpful or detrimental – and could even explain why some people seem to be naturally athletic.”
In the study, the scientists used several genomic technologies to map the interactions between MyoD and chromatin as cells turned into skeletal muscle upon MyoD expression.
Among other findings, the scientists determined that MyoD rearranged chromatin’s shape during this process – similar to the retying of a tangled shoelace.
Importantly, the researchers found that MyoD-driven reconfiguration of 3-D chromatin architecture is mediated by interactions between noncoding elements of the genome – where most disease-associated genetic variants occur.
These findings demonstrate that the noncoding genome can act as a structural element that defines the chromatin architecture – key information that will help predict the functional outcomes of these variants.
Puri is already applying this insight to help solve other genomic mysteries.
He plans to review a worldwide database of gene variations with unknown significance – meaning that scientists are unsure if the change is harmless or a risk factor for disease.
Then, he aims to create models that help us better understand the impact of these genetic variations on an individual’s ability to respond to environmental changes and eventually develop disease.
“It’s possible that many genetic variations alter chromatin folding.
Instead of directly causing disease, the changes may increase or decrease our disease risk,” explains Puri.
“I hope that my next studies will shed light on these genomic mysteries and help more people get definitive answers about what lies in their DNA.”
Epigenetics refers to the control of gene expression via mechanisms not directly related to the DNA coding sequence .
As a result, all cells in an organism have very different phenotypes despite having the same genome.
Epigenetics modulates and regulates gene expression through various epigenomic “marks”, the term given to chemical compounds added to DNA or histone proteins and recognized by enzymes that either lay down or remove the specific mark.
These marks change the spatial conformation of chromatin: either compacting it, thereby preventing the binding of transcription factors to the DNA, or opening it, allowing transcription factor binding and usually upregulating cellular processes.
DNA methylation—the addition of methyl groups to the 5-carbons of cytosine residues in CpG islands to give rise to 5-methylcytosines—works together with histone modifications to regulate gene expression.
DNA methylation tends to act at promoters to induce gene silencing, while histone acetylation usually unwinds chromatin.
DNA methylation is catalyzed by enzymes in the DNA methyltransferase (DNMT) family, which recruit functional complexes containing DNA methylation domains, leading to transcriptional inhibition or maintaining a repressive chromatin state.
Acetylation of histone tails is controlled by two enzyme families: histone acetyltransferases (HATs), which transfer an acetyl group, and histone deacetylases (HDACs) [4,5,6], which remove acetyl groups.
Other epigenetic marks are also described and include histone post-translational modifications such as methylation, ubiquitination, sumoylation, phosphorylation, biotinylation, and ADP-ribosylation, which either promote or suppress gene expression.
The pattern of these marks on histone tails is often referred to as the histone code, which dictates the binding of effector proteins that in turn results in specific cellular processes.
Non-coding RNAs (ncRNAs) are another type of specific epigenetic mark that mediate various intracellular processes .
A ncRNA is a functional RNA molecule transcribed from DNA but not translated into protein. The best characterized ncRNAs are microRNAs (miRNAs), which are short, single-stranded, 19–24 nucleotide ncRNAs. miRNAs regulate gene silencing at the transcriptional and/or translational level of protein-coding genes .
Long non-coding RNAs (lncRNAs) are another subset of RNAs over 200 nucleotides in length that function as chromatin remodelers, transcriptional regulators, and post-transcriptional regulators.
Many lncRNAs complex with chromatin-modifying proteins to recruit their catalytic activity to specific genomic sites, thereby modifying chromatin states and influencing gene expression. With little or no protein-coding potential, lncRNAs instead participate in various intracellular processes , and recent studies have identified that certain lncRNAs are specifically associated with certain cancers .
With the development of high-resolution sequencing and high-throughput technologies, a large number of biologically functional ncRNAs have now been identified .
In addition, there is crosstalk between DNA methylation and histone modifications ; for example, histone methyltransferases, histone demethylases, and accessory proteins interact and coordinate the chromatin state and DNA methylation and methylation status of histones are tightly associated .
Moreover, various ncRNAs are also closely associated with other epigenetic marks, which form extensive crosstalk throughout the cell, or the “epigenetic network” .
More recently, histone variants, mostly of canonical histones H2A, H2B, and H3, with specific properties have been identified in humans and other higher eukaryotes.
Most of them are H2A variants, among which macroH2A1 generates alternative splice isoforms, i.e., macroH2A1.1 and macroH2A1.2. MacroH2A1 isoforms appear to be critical regulators of chromatin structure and chromatin dynamics during cellular senescence, regeneration, and fasting [14,15].
The function of these histone variant proteins and their molecular mechanisms in health and during an organism’s lifespan are reviewed in .
The discovery of histone variants and their diversity has added further complexity to context-dependent biological systems and their regulation, such as during health, aging, and in pathological conditions including cancer.
This is an area of ongoing research, since chromatin dynamics throughout life are likely to change and the epigenome suffers from a progressive loss in configuration during aging .
The resulting abnormal chromatin state during aging is characterized by different incorporated histone variants, nucleosome remodeling, altered histone modification patterns, and altered DNA methylation patterns, resulting in the recruitment of different chromatin modifiers, abnormal gene expression patterns, and genomic instability. Among the multiple different variants of histone H2A, macroH2A has been implicated in aging; its level increases in an age-dependent manner during replicative senescence in cultured human fibroblast cells and also in several tissues of aged mice and primates .
These active or repressive marks are also dependent on lifestyle and environmental factors. “Environmental epigenetics” refers to how environmental exposures affect epigenetic changes .
Life experiences, habits, and our environment shape what and who we are by virtue of their impact on our epigenome and health; for instance, although identical twins share the same genome and are superficially phenotypically similar, they are unique individuals with definable differences.
These differences result from distinct gene expression influenced by epigenetic factors. Behavior, nutrition, and exposure to toxins and pollutants are among the lifestyle factors known to be associated with epigenetic modifications.
For example, nutrition is a key environmental exposure from gestation to death that impacts our health by influencing epigenetic phenomena.
In another example, recent epidemiological data suggest that the increased incidence of cancer observed in the developed world since the 1960s may partly be due to exposure to endocrine-disrupting chemicals (EDCs), to which humans and wildlife are exposed daily from multiple sources. These are discussed in detail below.
Cancer: Current Status and Future Prospects
Cancer is the second most common cause of death in most countries and will remain so as elderly people are most susceptible to cancer and the population is rapidly aging, at least in the west. While age-standardized cancer mortality rates are projected to decrease in the European Union (EU) [17,18] and the United States (US) [19,20] due to advances in screening, prevention, and treatment, the incidence has increased in Europe and the US for testicular and prostate cancers over the last 50 years [21,22,23].
In the United Kingdom (UK), the combined incidence of all cancers has increased for all age groups since the early 1990s, with the greatest increase seen in young people aged 0–24 years.
Cancer is a genetic disease characterized by inherited or sporadic mutations in genes that maintain tissue homeostasis, control the cell cycle, or regulate apoptosis.
Cancer is also an epigenetic disease characterized by mutations in chromatin-remodeling enzymes and epigenome alterations resulting from aberrant attachment or removal of DNA or histone protein marks. Accumulating evidence suggests that many adult diseases, including cancers, have epigenetic origins.
Nutritional Epigenetics in Health and Disease
Nutrition is one of the most studied and better understood environmental epigenetic factors. Associations have been observed between adverse prenatal nutritional conditions, postnatal health, and increased risk of disease.
For instance, at its extreme, the Dutch Famine Birth Cohort resulting from the Dutch Famine of 1944–1945 has been used to study the effects of starvation during pregnancy and subsequent health and developmental outcomes including, but not limited to, increased risk of type II diabetes mellitus, cardiovascular disease, metabolic disorders, and decreased cognitive function in later life [24,25,26].
The first months of pregnancy seem to have the greatest effect on disease risk; children conceived during the Dutch Famine tended to have smaller-than-usual offspring, suggesting that effects may persist and impact our children and even beyond. It seems likely that the fetus epigenetically adapts in response to a limited supply of nutrients.
In humans, persistent epigenetic differences associated with prenatal exposure to famine have been ascribed to a lower degree of methylation of a gene implicated in insulin metabolism than their unexposed siblings .
The evidence for transgenerational effects of poor maternal diet on human populations with respect to metabolic outcomes was examined in .
There is evidence from historical records that the grandchildren of women exposed to famine and other dietary alterations during pregnancy are more likely to experience health complications than their control counterparts.
The potential molecular mechanisms of transgenerational inheritance suggest methylation of gametes via the paternal and maternal lineage . Indeed, further transmission via the paternal line is highly likely to occur via epigenetic modulation of the spermatozoan nucleus.
Two examples from historical cohorts illustrate this transgenerational transmission via the paternal lineage. One showed that female grandchildren (F2) from the paternal grandmother (F0) who experienced poor food availability during her own growth were at higher risk of cardiovascular mortality.
The second more recent example showed that adult grand-offspring whose fathers were exposed to famine in utero had higher BMIs than a control population. The evidence that both maternal and paternal diets influence metabolic phenotypes in offspring in mammals through epigenetic information transmission is reviewed in . Over molecular mechanisms with respect to the fetal origins of adult disease have been suggested including mitochondrial dysfunction and oxidative stress as among the earliest events described in offspring exposed to nutrient restriction .
Nutrients can either act directly by inhibiting epigenetic enzymes such as DNMT, HDAC, or HAT or by altering the availability of substrate necessary for those enzymatic reactions. This in turn modifies the expression of critical genes and impacting on our overall health and longevity (see [12,31] for reviews).
A number of studies have reported the epigenetic effects of diet on phenotype and susceptibility to diseases throughout life. Folate metabolism is linked to phenotypic changes through DNA methylation, as folate, a water-soluble B vitamin, is a source of one-carbon for the synthesis of AdoMet, which is necessary for DNA methylation [12,30,31].
Other methyl donor nutrients such as choline can also alter the DNA methylation status and subsequently impact gene expression .
Maternal methyl donor nutrient availability in early pregnancy is essential for proper fetal development, with consequences for health and disease susceptibility or cancer in the children throughout life. In one animal study, a maternal diet restricted in methyl nutrients during periconception affected DNA methylation patterns in offspring and was the cause of altered phenotypes .
Conversely, dietary restriction but without severe nutritional deprivation has been shown in several models to extend lifespan . Calorie restriction has an anti-inflammatory effect through the inhibitory effects of critical genes not limited to NF-κB . From the epigenetic standpoint, there is clear crosstalk between DNA methylation and histone modifications , suggesting that the chromatin structure may also determine DNA methylation .
In this context of epigenetic interactions, sirtuin 1, a NAD+-dependent HDAC whose substrate specificity includes histone proteins, has been suggested to be activated by some dietary components (for example resveratrol, a type of natural phenol present in grape skins).
Sirtuin 1 mediates some of the effects of dietary restriction that delay or reverse some of the physiological changes associated with aging through effects on DNA methylation .
Diets high in these methyl-donating nutrients can rapidly impact gene expression, especially during early development when the epigenome is first established, and can have long-lasting effects in adult life.
Studies in animals have reported that diets poor in methyl-donating folate or choline before or just after birth cause permanent hypomethylation of parts of the genome. In adults, a methyl-deficient diet decreases DNA methylation, but the changes are reversible when methyl is added back into the diet.
Further, depending on the dietary supplements received by a pregnant mouse, her offspring may have a different phenotype due to differential gene methylation. In an example of epigenome-modifying chemicals, bisphenol A (BPA) is widely used to manufacture numerous plastic products including containers.
The pups of adult mice fed BPA were more likely to have an unhealthy phenotype (yellow/obese, prone to cancer and diabetes) compared to those born from mothers fed BPA with supplemented methyl-rich nutrients like folic acid and vitamin B12 (brown, thin and healthy).
In this case, maternal nutrient supplementation counteracted the negative effects of chemical exposure, underscoring the importance of a good diet rich in fruit and vegetables and other high-quality foods.
Methyl-donating nutrients act as co-substrates for methyl group transfers; the pool of available methyl donors is an important regulator of both DNA and histone methylation capacity  and their production is also dependent on BPA’s epigenetic effects. In this mouse model, altered DNA hypomethylation could be alleviated by folic acid as a dietary methyl donor.
Another striking example of the effects of early diets on epigenetics with consequences on the phenotype can be found in honey bees.
The sterile worker bee differentiates from the fertile queen depending on the larval diet through epigenetic changes in DNA methylation patterns. Larvae designated to become queens are fed exclusively with royal jelly, which contains epigenetically active ingredients that silences a key gene which itself silences a group of queen genes .
Furthermore, DNA methylation changes occur during ageing, and it has become evident that early life nutrition can modulate DNA methylation and influence longevity, in particular by inducing long-term changes in DNA methylation and other marks that affect susceptibility to a range of ageing-associated diseases ; that is, a form of “cellular memory” (see  for a review). In this regard, the lifespan of a queen bee is up to twenty times that of a worker.
Nutritional Epigenetics and Cancer
Folic acid and vitamin B12 are two examples of epigenetically active ingredients that play important roles in DNA metabolism and the maintenance of DNA methylation patterns via chemical reaction of a methyl product. In one in vivo study, dietary folate intake was positively correlated with p16 tumor suppressor gene expression, a critical cancer-associated gene with frequent silencing DNA methylation of its promoter .
Altered p16 gene expression was observed in aged mouse colons, consistent with the known decrease in DNMT expression with aging.
There is a growing body of epidemiological evidence that folate modulates anticarcinogenic properties through epigenetic changes, as folate deficiency reduces the potential for DNA methylation, and abnormal DNA methylation is associated with many types of cancer. Diets rich in fruits and vegetables containing natural anti-oxidants can protect against cancer.
The potential epigenetic effects of several nutritional components in addition to folate, mostly derived from vegetables, have been examined in a number of studies, which have shown reductions in DNA hypermethylation of critical genes resulting in tumor suppression [32,35]. For example, green tea contains polyphenols, which are natural compounds widely distributed in plant foods and with many biological activities including inhibition of DNA methylation.
A variety of dietary factors are potential HDAC and HAT modulators. Some, such as sulforaphane, an isothiocyanate found in broccoli sprouts, or diallyl disulfide, an organosulfur compound in garlic, have been shown to act as HDAC inhibitors [9,35], a class of epigenetic therapeutic described further below. Such epigenetic drugs have been used to treat cancers in clinical trials due to their mode of action in restoring cancer cell differentiation and rendering tumors more sensitive to conventional therapies . Several in vitro studies using these compounds have shown anti-carcinogenic effects associated with HDAC inhibition and histone acetylation [12,37,38]. Table 1 summarizes some dietary components considered to have protective effects against cancer through different epigenetic modifications.
A summary of some dietary components considered to have protective effects against cancer.
|Nutrient||Food Origin||Epigenetic Role|
|Methionine||Sesame seeds, brazil nuts, fish, peppers, spinach||SAM synthesis|
|Folic Acid||Leafy vegetables, sunflower seeds, baker’s yeast, liver||Methionine synthesis|
|Vitamin B12||Meat, liver, shellfish, milk||Methionine synthesis|
|Vitamin B6||Meats, whole grain products, vegetables, nuts||Methionine synthesis|
|SAM-e (SAM)||Popular dietary supplement pill; unstable in food||Enzymes transfer methyl groups from SAM directly to the DNA|
|Choline||Egg yolks, liver, soy, cooked beef, chicken, veal and turkey||Methyl donor to SAM|
|Betaine||Wheat, spinach, shellfish, and sugar beets||Break down the toxic byproducts of SAM synthesis|
|Resveratrol||Red wine||Removes acetyl groups from histones, improving health (shown in lab mice)|
|Genistein||Soy, soy products||Increased methylation, cancer prevention, unknown mechanism|
|Sulforaphane||Broccoli||Increased histone acetylation turning on anti-cancer genes|
|Butyrate||A compound produced in the intestine when dietary fiber is fermented||Increased histone acetylation turning on ‘protective’ genes, increased lifespan (shown in the lab in flies)|
|Diallyl sulphide (DADS)||Garlic||Increased histone acetylation turning on anti-cancer genes|
As noted above, miRNAs can regulate DNA methylation and histone modifications, but promoter methylation or histone acetylation can also modulate miRNA expression as part of a complex network with feed-forward and feedback loops. Dysregulated miRNA expression is associated with the development or progression of human cancers through alterations in cell proliferation and apoptosis, but methyl- and folate-deficient diets can also result in aberrant miRNA expression to exert similar, pro-cancer effects . Specifically, miR-222 has been considered a potential biomarker of nutritional status in humans and is implicated in obesity. Certain dietary components may protect against cancer through miRNA regulation, such as curcumin and retinoic acid , the former present in some plants and commonly used as a dietary supplement and food flavoring, and the latter present in any vitamin A-rich food.
Dietary patterns, not only individual nutrients, also influence behavior and phenotype in offspring. For example, Western diets tend to be high in saturated fats, red meats, and empty carbohydrates but low in fresh fruits and vegetables, whole grains, seafood, and poultry. This diet has been linked to many diseases including hypertension, heart disease, diabetes, and obesity and it has generally been linked to an increased risk of cancer [39,40]. In a specific example, nonalcoholic fatty liver disease (NAFLD) is a major public health concern in western societies. Nonalcoholic steatohepatitis (NASH), a form of NAFLD, is characterized by lipid accumulation in hepatocytes, inflammatory cell infiltrates, oxidative stress, and fibrosis and can lead to cirrhosis or hepatocellular carcinoma. The effects of a Western diet  or a high fat diet  on NAFLD development has been investigated in rodents. The risk of NAFLD may increase through an imbalance in fatty acids (FAs) in the Western diet. For instance, substituting linoleic acid with α-linolenic acid or long chain n-3 polyunsaturated fatty acids and decreased the n-6:n-3 FA ratio in high fat, high fructose (HFHF) diet-induced NASH . The data from this study showed that decreasing the n-6:n-3 ratio by introducing healthy fats prevented HFHF-induced NASH by attenuating oxidative stress and inflammation and restoring the antioxidant state.
Epigenetics, such as DNA methylation, may be involved in the impact the western diet has on the human body. The effects of maternal Western diet on offspring physical activity, gene expression, and phenotype were assessed in . Interestingly, differences in F1 female offspring but not in F2 male and female offspring were observed, suggesting that changes in the F1 generation were related to in utero somatic reprogramming. Epigenetic effects of specific FAs have been investigated in a number of studies. Eicosapentaenoic acid (EPA) and arachidonic acid (AA) are products of essential FA metabolism; both FAs are involved in inflammation resolution. EPA has been long regarded as a protective FA, particularly in the light of the favorable cardiometabolic effects of fish oil. One study  demonstrated a strong association between whole peripheral blood DNA methylation and EPA and AA in two distinct human cohorts—lactating infants and adult men—of different ages and developmental stages, thereby linking EPA and AA to DNA hypermethylation. Another study  reported that maternal dyslipidemia caused significant epigenetic changes in placentas and fetal livers and also increased fetal liver triacylglycerol accumulation. It has been shown from animal experiments that cardiovascular and metabolic diseases, particularly in males, may develop from alterations in DNA methylation. A recent study demonstrated how parental diet may affect their offspring’s epigenetic modifications and lead to the development of cardiovascular and metabolic diseases and impact central nervous system plasticity . This study demonstrated that mice exposed to a high-fat, high-sugar diet (HFHSD) prior to and during pregnancy led to DNA modifications of their offspring’s compensatory renin-angiotensin system (RAS), a hormone system that regulates blood pressure, fluid retention, and vascular resistance. In adulthood, offspring from HFHSD-exposed dams exhibited several differences compared to control counterparts including but not limited to a lower level of angiotensin converting enzyme-2 (ACE2) gene expression in the brain stem, kidney, and cecum and higher ACE2 gene activity in the hypothalamus. These data suggest that perinatal exposure to HFHSD resulted in epigenetic modifications of the compensatory brain RAS, potentially affecting plasticity of neuronal networks leading to autonomic dysfunction in the male offspring.
In conclusion, following a western diet even before the chid’s birth may lead to physiological dysfunction via epigenetic changes. Further investigations, such as looking at potential transgenerational transmission in F2 male and female mice by crossing F1 males (offspring from HFHSD-exposed dams) with regular diet-fed females (and the opposite) would be of particular interest.
In contrast to the Western diet, a number of studies have described the health benefits of following the Mediterranean diet, which is associated with a reduced risk of heart disease and cardiovascular mortality as well as overall mortality. The Mediterranean diet traditionally includes fruits, vegetables, pasta and rice, fish and poultry, whole grains, and healthy fats (monounsaturated fats and polyunsaturated fats such as beneficial linolenic acid) and discourages red meat and saturated fats. The favorable effects of a Mediterranean diet as primary prevention of cardiovascular disease were assessed among persons at high cardiovascular risk . The data from this study revealed that the incidence of major cardiovascular events was lower in those assigned to a Mediterranean diet supplemented with extra-virgin olive oil or nuts than among those assigned to a reduced-fat diet . The Mediterranean diet is also associated with a reduced incidence of cancer, and the risk of breast cancer has been reported to be reduced in women who eat a Mediterranean diet supplemented with extra-virgin oil and mixed nuts. In relation to breast cancer, one study demonstrated that dietary patterns affect the mammary gland microbiome, establishing an alternative mechanistic pathway for breast cancer prevention . The impact of maternal Mediterranean diet adherence during pregnancy on offspring behaviors has also been investigated, and maternal adherence to a Mediterranean diet in early pregnancy is associated with positive neurobehavioral outcomes in early childhood and with sex-dependent methylation differences in the control regions of imprinted genes . Figure 1 summarizes how diet affects the epigenome to modify individual and transgenerational phenotypes.
Nutritional Epigenetics: The Future
Nutrients and bioactive food components can therefore reversibly alter the DNA methylation status, histone modifications, and chromatin remodeling, subsequently altering gene expression and having an impact on overall health. Bioactive food components, specific nutrients, and dietary patterns may have beneficial effects and overcome the negative impact of negative life behaviors, such as smoking or exposure to certain chemicals [51,52]. However, nutritional epigenetics is a quite recent subfield of epigenetics, so current knowledge on the precise effects of bioactive food components on epigenetics and their associations with phenotypes are limited. Deciphering the epigenetic signatures triggered by bioactive food components might pave the way for personalized nutritional interventions and aid our understanding of how our bodies respond to specific diets or nutrients . For example, a recent study showed that fruit and juice epigenetic signatures as measured by DNA methylation marks are associated with independent immunoregulatory pathways, suggesting that the health benefits of fruit and juices are distinct. The identification of such differences between foods is the first step toward personalized nutrition .
More information: Alessandra Dall’Agnese et al. Transcription Factor-Directed Re-wiring of Chromatin Architecture for Somatic Cell Nuclear Reprogramming toward trans-Differentiation, Molecular Cell (2019). DOI: 10.1016/j.molcel.2019.07.036
Journal information: Molecular Cell
Provided by Sanford Burnham Prebys Medical Discovery Institute