Download Nutritional Genomics

Nutritional Genomics
Evaluating the impact of maternal nutrition on
the epigenome of the offspring
using multi-omics data
Francisco Peñagaricano
Department of Animal Sciences
University of Florida
Nutritional Genomics: effects of maternal nutrition on the epigenome of the offspring
Presentation outline
 Fetal developmental programming
 Nutritional epigenomics
 Maternal methionine supplementation
 Programming of muscle and fat tissue development
 Conclusions and future perspectives
Nutritional Genomics: effects of maternal nutrition on the epigenome of the offspring
Fetal developmental programming
 Fetal programming describes the process whereby a maternal stimulus during a
critical period of development has lasting or lifelong consequences
 Maternal nutrition is a major intrauterine environmental factor in fetal development
Epidemiological studies in humans and experimental studies using animal models:
 Maternal nutrition during different stages of pregnancy can induce permanent
changes in the structure, physiology, and metabolism of the offspring
Nutritional Genomics: effects of maternal nutrition on the epigenome of the offspring
Epidemiological studies: Dutch Famine - “Hunger Winter” in 1944-1945
Exposure to poor nutrition during early gestation
 Lipid profile
 Obesity
 Blood coagulation
Exposure to poor nutrition during mid gestation
 Coronary heart disease
 Lung disease
 Renal function
Exposure to poor nutrition during later gestation
 Glucose metabolism
Maternal undernutrition during gestation has important effects on health in later life
 the timing of the nutritional insult determines which organ system is affected
Nutritional Genomics: effects of maternal nutrition on the epigenome of the offspring
Experimental studies using rodent models
Restriction of maternal protein intake throughout gestation
 decreased birth weight
 perturbations to renin-angiotensin system
 kidney disorders
 adult hypertension
Maternal high-fat over-feeding during pregnancy
abnormal glucose homeostasis


abnormal serum lipid profiles

increased adiposity
Maternal nutrition during pregnancy can induce remarkable effects on fetal development
 predispose the offspring to metabolic, endocrine, and cardiovascular disorders in postnatal life
Nutritional Genomics: effects of maternal nutrition on the epigenome of the offspring
Theory of fetal origins of adult disease – Barker’s hypothesis
The period of gestation has significant impacts on the
subsequent health and wellbeing outcomes for an
individual ranging from infancy to adulthood
David Barker
Nutritional Genomics: effects of maternal nutrition on the epigenome of the offspring
Nutritional epigenomics
Maternal nutrition can induce epigenetic alterations in the fetal genome
 such as DNA methylation, which in turn leads to gene expression changes
Nutritional Genomics: effects of maternal nutrition on the epigenome of the offspring
Nutritional epigenomics
Maternal nutrition can induce epigenetic alterations in the fetal genome
 such as DNA methylation, which in turn leads to gene expression changes
DNA Methylation: major epigenetic mark and a well-known regulator of gene expression
depends on the availability of methyl donors:
 methionine
 choline
 betaine
 vitamin B12
 folic acid
Nutritional Genomics: effects of maternal nutrition on the epigenome of the offspring
Nutritional epigenomics
DNA Methylation: major epigenetic mark and a well-known regulator of gene expression
depends on the availability of methyl donors:
 methionine
 choline
 betaine
 vitamin B12
 folic acid
Nutritional Genomics: effects of maternal nutrition on the epigenome of the offspring
Nutritional epigenomics
Maternal nutrition can induce epigenetic alterations in the fetal genome
 such as DNA methylation, which in turn leads to gene expression changes
DNA Methylation: major epigenetic mark and a well-known regulator of gene expression
depends on the availability of methyl donors:
the link between maternal diet and subsequent modification of gene
expression in the fetal genome is one of the most important molecular
mechanisms proposed to explain the phenomenon of fetal programming
Nutritional Genomics: effects of maternal nutrition on the epigenome of the offspring
Nutritional epigenomics: metastable epialleles in mice
One of the most remarkable examples of epigenetically sensitive genes via maternal diet
agouti viable yellow
axin fused
alleles that are variably expressed in genetically identical individuals due to epigenetic modifications
 metastable epialleles exhibits epigenetic plasticity to maternal diet
Nutritional Genomics: effects of maternal nutrition on the epigenome of the offspring
Nutritional epigenomics: farm animals
Objective: evaluate the effects of restricting the supply of vitamin B12, folate, and methionine
from the diet of ewes from 8 weeks before conception until 6 days after conception
Nutritional Genomics: effects of maternal nutrition on the epigenome of the offspring
Nutritional epigenomics: farm animals
subtle diet manipulation of components of the one-carbon cycle during the periconceptional
period can impact the DNA methylation of the fetal genome, and alter adult health-related traits
 altered immune response, insulin-resistance, and elevated blood pressure
Nutritional Genomics: effects of maternal nutrition on the epigenome of the offspring
Nutritional epigenomics: farm animals
 Methionine supplementation before and around conception seems to be a promising
tool for improving reproductive performance in lactating dairy cattle
 effects of a maternal methyl supplemented diet on the epigenome of the embryo?
Objective: evaluate the effect of methionine supplementation before and around conception on
the transcriptome of preimplantation embryos from lactating dairy cows
Nutritional Genomics: effects of maternal nutrition on the epigenome of the offspring
Maternal Met supplementation and embryo transcriptome
Animals and maternal diets
Holstein dairy cows were randomly assigned to one of two treatments differing in level of
dietary methionine from calving until embryo flushing
(1) methionine-rich; diet formulated to deliver 2,875 g metabolizable protein (MP) with 6.8 lysine
%MP and 2.4 Met %MP; (2) control; same basal diet but formulated to contain only 1.9 Met %MP
Cows were superovulated and embryos were recovered 6 d after synchronized ovulations
 high quality preimplantation embryos from individual cows were pooled and then
analyzed using RNA sequencing technology
Nutritional Genomics: effects of maternal nutrition on the epigenome of the offspring
Maternal Met supplementation and embryo transcriptome
1. Lab work - total RNA extraction, amplification, library preparation,
and sequencing (following Illumina mRNA-Seq protocol)
2. Map the reads to the reference genome
3. Reconstruct transcript models
4. Generate a final assembly merging together each of the assemblies
5. Count the number of reads that map to each gene
6. Identify differentially expressed genes
Statistical Analysis
7. Perform a gene-set enrichment analysis (look at the expression changes of sets of genes)
Nutritional Genomics: effects of maternal nutrition on the epigenome of the offspring
Maternal Met supplementation and embryo transcriptome
Exploration of overall gene expression
Multidimensional Scaling Analysis
The plot shows that dimension 1 clearly
separated control from methionine samples
 biological replicates are consistent
 multiple differentially-expressed genes between treatments can be detected
Nutritional Genomics: effects of maternal nutrition on the epigenome of the offspring
Maternal Met supplementation and embryo transcriptome
Evaluation of overall gene expression
Controlling FDR at 0.10, 276 genes showed at least a 2-fold
expression difference between treatments
The expression of most genes was decreased by maternal
methionine supplementation
Several of the most significant genes are related to embryonic
development and immune response
Nutritional Genomics: effects of maternal nutrition on the epigenome of the offspring
Maternal Met supplementation and embryo transcriptome
Gene-set enrichment analysis
Evaluation of different Gene Ontology terms, InterPro motifs, and KEGG pathways
 Several pathways were enriched with differentially expressed genes
 Many pathways related to embryo development and immune system
GO terms - Tube development, Immune response, Cytokine binding
InterPro motifs - Several terms associated with immunoglobulin motifs
KEGG pathway - cytokine-cytokine receptor interaction
 Remarkable down-regulation of gene expression due to maternal methionine supplementation
Nutritional Genomics: effects of maternal nutrition on the epigenome of the offspring
Maternal Met supplementation and embryo transcriptome
Conclusions
 Although the embryos showed identical morphological appearance (same grading for
developmental stage and quality), they showed significant transcriptomic differences
 Expression of several genes that are critical for embryonic and adult function were decreased
by methionine supplementation, probably due to increased DNA methylation
 Our findings provide evidence that maternal diet rich in methionine can significantly
modulate gene expression in bovine preimplantation embryos
Nutritional Genomics: effects of maternal nutrition on the epigenome of the offspring
Fetal programming and livestock production
 Postnatal effects of maternal nutrition on growth, productivity, and reproduction
of the offspring could have important implications in the livestock industry
 maternal nutrition can alter subsequent growth, skeletal muscle development, body
composition, and energy metabolism in postnatal life of offspring in livestock species
Nutritional Genomics: effects of maternal nutrition on the epigenome of the offspring
Fetal programming and livestock production
Studies by Rick N Funston in University of Nebraska
Maternal protein supplementation during late gestation in beef cattle
 positively associated with postnatal growth and adipose deposition in steer progeny
 postweaning body weight and fertility in heifer progeny
Studies by Amy E Radunz in Ohio State University
Different prepartum energy sources fed during late gestation in both beef cattle and sheep
resulted in changes in postnatal muscle and fat deposition in the progeny
Nutritional Genomics: effects of maternal nutrition on the epigenome of the offspring
Programming of muscle and fat tissue development
Maternal nutrition can impact muscle and adipose tissues development
Objective: evaluate the effect of different maternal diets during mid-to-late gestation on
the transcriptome of fetal muscle and adipose tissues in sheep
Nutritional Genomics: effects of maternal nutrition on the epigenome of the offspring
Programming of muscle and fat tissue development
Animals and maternal diets
Multiparous Polypay ewes were naturally bred to a single sire and from days 67 ± 3 of gestation until
necropsy (days 130 ± 1), they were individually fed one of three isoenergetic diets:
 alfalfa haylage (HY; fiber)
 corn (CN; starch)
 dried corn distiller’s grains (DG; fiber, protein & fat)
CN diet had lower protein than DG and HY diets
DG diet had higher fat than CN and HY diets
Nutritional Genomics: effects of maternal nutrition on the epigenome of the offspring
Programming of muscle and fat tissue development
Animals and maternal diets
Multiparous Polypay ewes were naturally bred to a single sire and from days 67 ± 3 of gestation until
necropsy (days 130 ± 1), they were individually fed one of three isoenergetic diets:
 Tissue sample collection (fetal tissues):
longissimus dorsi muscle, perirenal adipose depot, and subcutaneous adipose depot
 Samples were pooled and then analyzed using RNA sequencing technology
Nutritional Genomics: effects of maternal nutrition on the epigenome of the offspring
Programming of muscle and fat tissue development
Evaluation of gene expression in muscle
Most significant genes are closely related to fetal development, skeletal muscle cell and tissue
differentiation, and muscle myosin complex and sarcomere organization
Nutritional Genomics: effects of maternal nutrition on the epigenome of the offspring
Programming of muscle and fat tissue development
Evaluation of gene expression in fat
Most significant genes are related to fetal development, adipogenesis, cholesterol and fatty
acid biosynthetic process, and metabolism of lipids and lipoproteins
Nutritional Genomics: effects of maternal nutrition on the epigenome of the offspring
Programming of muscle and fat tissue development
Conclusions
 Our findings provide evidence that maternal diet during mid-to-late gestation can modulate
gene expression in fetal tissues in sheep
 Gene set enrichment analysis revealed many functional categories, including terms related to
tissue and organ development, energy metabolism, and chromatin biology
 Maternal starch-based diet mainly altered muscle development while a maternal diet with high
fiber, protein, and fat concentrations mainly impacted adipose tissues
Nutritional Genomics: effects of maternal nutrition on the epigenome of the offspring
Nutritional epigenomics: imprinted genes
 Imprinted genes : genes that are expressed in a parent-of-origin-specific manner
 Epigenetic modifications, such as DNA methylation, regulate expression of imprinted genes
 evaluate the effect of maternal diets on the expression of imprinted genes in fetuses in sheep
Nutritional Genomics: effects of maternal nutrition on the epigenome of the offspring
Nutritional epigenomics: imprinted genes
Gene expression in muscle
insulin like growth factor 2 receptor
Nutritional Genomics: effects of maternal nutrition on the epigenome of the offspring
Nutritional epigenomics: imprinted genes
Gene expression in muscle
DNA Methylation in IGF2R
insulin like growth factor 2 receptor
↑ 𝑷𝒓𝒐𝒕𝒆𝒊𝒏 →→ ↑ 𝑴𝒆𝒕𝒉𝒚𝒍𝒂𝒕𝒊𝒐𝒏
↑ 𝑴𝒆𝒕𝒉𝒚𝒍𝒂𝒕𝒊𝒐𝒏 →→ ↑ 𝑮𝒆𝒏𝒆 𝑬𝒙𝒑𝒓𝒆𝒔𝒔𝒊𝒐𝒏
Nutritional Genomics: effects of maternal nutrition on the epigenome of the offspring
Nutritional epigenomics: future perspectives
At least four key questions:
 if a maternal diet rich in methyl donors increases DNA methylation of the offspring genome
 if these DNA methyl marks are transient or persist across time
 if DNA methylation modulates gene expression
 if there are specific functional sets of genes that underlay fetal programming
Nutritional Genomics: effects of maternal nutrition on the epigenome of the offspring
Nutritional epigenomics: future perspectives
Ongoing Project
maternal nutritional treatments:
control diet and methionine-rich diet
from day -30 until day +90 relative to the start of the breeding season
tissues will be collected across time from the same group of animals
genome-wide DNA methylation and gene expression
Nutritional Genomics: effects of maternal nutrition on the epigenome of the offspring
Nutritional epigenomics: future perspectives
At least four key questions:
 if a maternal diet rich in methyl donors increases DNA methylation of the offspring genome
 if these DNA methyl marks are transient or persist across time
 if DNA methylation modulates gene expression
 if there are specific functional sets of genes that underlay fetal programming
 a deep understanding of these epigenetics mechanisms will benefit livestock production
and may also have a great impact on human health
Nutritional Genomics: effects of maternal nutrition on the epigenome of the offspring
Fetal programming and human health
Methyl supplementation around conception and during early pregnancy
 use of periconceptional and prenatal supplements rich in folic acid
and other methyl donors to prevent potential defects in developing fetuses
 recent studies suggest that maternal diets rich in methyl donors may lead to epimutations
that are likely associated with negative health outcomes, including neurobehavioral disorders
Nutritional Genomics: effects of maternal nutrition on the epigenome of the offspring
Perspectives
 Evidence that maternal diets can indeed modulate gene expression in the offspring
 Functional ramifications of the observed gene expression changes warrant future research
 The epigenetic mechanisms underlying fetal programming are not well understood
 Critical need to understand the effects of maternal diets on offspring epigenome/performance
 Use farm animals as models: potential contributions in food production and also human health
Nutritional Genomics: effects of maternal nutrition on the epigenome of the offspring
Thanks for your attention!
Contact Information:
Phone: (352) 392-1981
E-mail: [email protected]
Website: fpenagaricano-lab.org