The somatic cells of the human body contain 20,000 to

25,000 genes. Yet in any of these cells, only a relatively

small percentage of all genes are active.

In the more than 200 different cell types present in the

body, different cell-specific gene sets are transcribed,

while the rest of the genome is transcriptionally
inactive.

In addition, programs of gene expression become more

and more restricted during development and

differentiation as embryonic cells gradually become
specialized adult cells with distinct phenotypes.

 The prevailing view has been that regulation of gene

expression is coordinated by promoter, promoter
proximal, enhancer, and other cis-regulatory
elements as well as DNA-binding proteins and
transcription factors.
The newly emerging field of epigenetics is providing
us with a basis for understanding how heritable

changes other than those in DNA sequence can

influence phenotypic variation.

These advances greatly extend our understanding of

the molecular basis of gene regulation and apply to

wide-ranging areas including genetic disorders,
cancer, and behavior.

An epigenetic trait : is a stable, mitotically and

meiotically heritable phenotype that results from
changes in gene expression without alterations in the
DNA sequence.
Epigenetics: is the study of the ways in which these
changes alter cell- and tissue-specific patterns of gene
expression.
The epigenome: refers to the epigenetic state of a cell.
During its life span, an organism has one genome, but
this genome can be modified in diverse cell types at
different times to produce many epigenomes.

Current research efforts are focused on several

aspects of epigenetics:

how an epigenome arises in developing and

differentiated cells.

what mechanisms maintain these states, and how they are

transmitted via mitosis and meiosis, making them

heritable traits.
In addition, because epigenetically controlled alterations

to the genome are associated with cancer and some

common diseases such as diabetes and asthma, efforts are
also being directed at the development of drugs that can
modify or reverse disease-associated epigenetic changes
in cells.

Epigenetic pathway
Several systems and pathways that result in the
establishment, maintenance, and inheritance of the
epigenetic state are recognized. These pathways are
organized into three categories.

1-Epigenators
The first category includes environmental signals
called epigenators that are received by the cell
and that stimulate a response via an intracellular
pathway.

2- epigenetic initiators
Responses to epigenator signals are called
epigenetic initiators. Components of this
second category produce epigenetic changes.
These initiators include proteinprotein signal
transduction pathways, DNA binding proteins,
and noncoding RNAs. The actions of initiators
define the location at which epigenetic changes in
chromatin will take place. DNA sequence
recognition is a necessary part of this response.

3- epigenetic maintainers
Once the epigenetic modifications have occurred, they are

maintained by molecular elements called epigenetic

maintainers.
Components of this third category conserve and sustain
the epigenetic changes in the present and future
generations.
Epigenetic maintainers are not sequence-specific, they
operate anywhere in the genome, and they depend on
initiators to specify the loci at which chromatin
modifications will take place.
Maintainers include DNA methylation and histone
modifications.

Epigenetic Alterations To The Genome

Unlike the genome, which is identical in all cell types

of an organism, the epigenome is cell-type specific

and heritable.
Like the genome, the epigenome can be transmitted to
daughter cells by mitosis and to future generations by
meiosis.
There are three major epigenetic mechanisms:
(1) Reversible modification of DNA by the
addition or removal of methyl groups
(2) modification of histones by the addition or
removal of chemical groups
(3) regulation of gene expression by small,
noncoding RNA molecules.

1-Methylation
In mammals, methylation of DNA takes place after replication

and involves the addition of a methyl group (CH3) to

cytosine, a reaction catalyzed by methyltransferase enzymes.

DNA methylation also occurs during the differentiation of

adult cells.

In both instances, methylation takes place almost exclusively

on cytosine bases adjacent to a guanine, a combination called

a CpG dinucleotide.

Many of these dinucleotides are clustered in regions, called

CpG islands, located in and near promoter sequences adjacent

to genes.

 Islands adjacent to essential genes (housekeeping

genes) and cell-specific genes are unmethylated,
making these genes available for transcription.

Other genes with adjacent methylated CpG islands

are transcriptionally silenced.
The methyl groups in CpG islands occupy the major
groove of DNA, and block the binding of
transcription factors necessary to form transcription
complexes.

The bulk of methylated CpG dinucleotides are found

in repetitive DNA sequences located in
heterochromatic regions of the genome, including the
centromere.

 Methylation of these sequences contributes to silencing

the transcription and replication of transposable

elements such as LINE and SINE sequences which
constitute a major fraction of the human genome.

Heterochromatic methylation also maintains

chromosome stability by preventing translocation and

other chromosomal abnormalities.

As part of dosage compensation, X chromosomes in

mammalian females are inactivated by converting them

to heterochromatin. These inactivated chromosomes
have altered patterns of DNA methylation. As mentioned
above, CpG methylation in euchromatic regions causes a
parentspecific pattern of gene transcription.

2- Histone Modification
histone modification is an important epigenetic

mechanism of gene regulation.

Recall that chromatin is composed of DNA wound
around an octamer core of histone proteins to form
nucleosomes.
Amino acids in the N-terminal region of these histones
can be covalently modified in several ways, including
acetylation, methylation, and phosphorylation.
These modifications occur at conserved amino acid
sequences in the N-terminal histone tails.
Chemical modification of histones alters the structure of
chromatin, making genes accessible or inaccessible for
transcription.

Features of Histone
Modifications
Amino acids in the Nterminal region of these
histones can be covalently
modified in several ways,
including acetylation,
methylation, and
phosphorylation.

Methyl

Acetyl

Phospho

Normally, when histones are modified by acetylation, a reaction catalyzed

by the enzyme histone acetyltransferase (HAT), chromatin structure
becomes open, making genes on these modified nucleosomes available
for transcription
This modification is reversible, and acetyl groups can be removed by
another enzyme, histone deacetylase (HDAC), changing the chromatin to
a closed configuration, and silencing genes by making them unavailable
for transcription

Covalently attached groups (usually

to histone tails)

Methyl

Acetyl

Phospho

3- RNA Interference
After transcription, small interfering RNA (siRNA)
molecules associate with protein complexes to form
RNA-Induced Silencing Complexes (RISCs).
RISCs bind to mRNA molecules that carry
sequences complementary to siRNA in the RISC.

If the siRNA is not perfectly complementary to the

mRNA, the binding interferes with translation,
resulting in downregulation of gene expression.
If, however, the siRNA in the RISC is perfectly
complementary to sequences in the mRNA, the
mRNA is cleaved and destroyed, effectively

silencing the gene.

Recently, it has been discovered that siRNAs can

silence genes by directly interfering with transcription
initiation. This does not involve any changes in
existing epigenetic promoter modifications, nor does it
require new modifications.
Instead, siRNAs complementary to promoter regions
bind to a promoter. Binding blocks the assembly of the
preinitiation complex by preventing binding of
transcription factor TFIIB and RNA polymerase.

 In sum, epigenetic modifications alter

chromatin structure by several

mechanisms including DNA methylation,
histone acetylation, and RNA interference,
without changing the sequence of DNA.

These epigenetic changes create an

epigenome that in turn, can regulate

normal development or generate responses
to environmental signals.

Epigenetics and Imprinting

Mammals inherit a maternal and a paternal copy of each

autosomal gene, and either or both copies of these genes

can be expressed in the offspring.

Imprinted genes show expression of only the maternal

allele or the paternal allele.

This parent-specific pattern of allele expression is laid

down during gamete formation.

Differential methylation of CpG rich regions produce

allele-specific imprinting and subsequent gene silencing.

 In mice, fewer than 100 genes were thought to be

imprinted, but recent work has identified more
than 1300 imprinted genes.
In humans, more than 150 candidate genes are
thought to be imprinted, but the findings in mice
suggest that many more imprinted genes remain to
be identified in humans.
Once a gene has been methylated and imprinted, it
remains transcriptionally silent during
embryogenesis and development.

Mutations and imprinted genes

At the level of individual genes, having only one

functional allele makes these genes highly susceptible

to the deleterious effects of mutations.

Because imprinted genes are clustered, mutation in

one gene can have an impact on the function of
adjacent imprinted genes, amplifying its impact on the
phenotype.

Mutations in imprinted genes can arise by changes in

the DNA sequence or by epigenetic changes, called
epimutations, both of which are heritable changes in
the activity of a gene.

Imprinting patterns are

reprogrammed each generation
For example, females receive a
maternal and a paternal set of
chromosomes. In somatic cells and in
germ cells, the maternal chromosome
set has female imprints, and the
paternal set contains male imprints.
When gamete formation begins in
female germ cells, both chromosome
sets have their imprints erased and are
each reprogrammed by changing the
pattern of methylation to carry a
female imprint pattern that is
transmitted to the next generation
through the egg.

Most human disorders associated with imprinting have their

origins during fetal growth and development.
Imprinting defects cause PraderWilli syndrome, Angelman
syndrome, BeckwithWiedemann syndrome, and several
other diseases (ST Table 3.1).
However, given the number of candidate genes and the
possibility that additional imprinted genes remain to be
discovered, the overall number of imprinting-related genetic
disorders may be much higher.

 In humans, most known imprinted genes encode

growth factors or other growth-regulating

genes.
Generally, maternally expressed alleles of
imprinted genes suppress growth, and paternal
alleles enhance growth.
One autosomal dominant disorder of imprinting,
BeckwithWiedemann syndrome (BWS), offers
insight into how disruptions of epigenetically
imprinted genes lead to an abnormal phenotype.

BeckwithWiedemann syndrome (BWS),

BWS is a prenatal overgrowth disorder with
abdominal wall defects, enlarged organs, large birth

weight, and predisposition to cancer.

BWS is not caused by mutation, nor is it associated

with any chromosomal aberration.

Instead it is a disorder of imprinting and is caused by

abnormal methylation patterns.

BWS
.

 Genes linked to BWS are located in a cluster of

imprinted genes on the short arm of chromosome 11.
This cluster contains more than a dozen imprinted
genes, some of which are paternally expressed, while
others are maternally expressed and all genes in this
cluster regulate growth during prenatal development.
The imprinted region is subdivided into two
separately regulated domains, one of which contains

the closely linked genes IGF2 (insulin growth factor

2) and H19.

 Normally, the paternal allele of IGF2 is expressed, and

the allele on the maternal homolog is imprinted and

silenced. In the case of H19, the situation is usually the
reverse.
The protein encoded by IGF2 is a growth factor, and the
product of the H19 is a long, noncoding RNA that is a
growth repressor.
Expression of these genes is normally controlled by an
imprinting control region (ICR) located within its
chromosomal domain.
Many affected individuals have a loss of imprinting of
the maternal IGF2 allele. This causes both the maternal
and paternal alleles to be transcriptionally active,
resulting in the overgrowth of tissues characteristic of
this disease.

Epigenetics and Cancer

Epidemiological studies investigate the role of

environmental factors in normal phenotypic variation

and as risk factors for disease.

For some complex diseases, there are strong links with

environmental factors such as the association between

smoking and lung cancer.

Following the discovery of cancer-associated genes,

including tumor-suppressor genes and protooncogenes, research into the genetic basis of cancer
focused mainly on mutant alleles of genes involved in
several cellular functions, including the cell cycle,
differentiation, and apoptosis.

 Converging lines of evidence are clarifying the role that

epigenetic changes play in the initiation and

maintenance of malignancy.

The relationship between epigenetics and cancer was

first noted in the 1980s by Feinberg and Vogelstein who

observed that colon cancer cells had much lower levels
of methylation than normal cells derived from the same
tissue.

Subsequent research by many investigators showed that

global hypomethylation is a property of all cancers

examined to date.

In the ensuing years, it has become clear that the

epigenetic states of normal cells are greatly altered
in cancer cells and that other epigenetic changes,
including selective hypermethylation and gene
silencing, are also present in cancer cells.

Cancer is now being viewed as a disease that

involves both epigenetic and genetic changes that
lead to alterations in gene expression (ST Figure 3
6).

 DNA hypomethylation reverses the inactivation of

genes, leading to unrestricted transcription of many

gene sets including oncogenes.

Hypomethylation of repetitive DNA sequences in

heterochromatic regions is associated with an increase

in chromosome rearrangements and changes in
chromosome number, both of which are characteristic
of cancer cells.

In addition, hypomethylation of repetitive sequences

leads to transcriptional activation of transposable DNA

sequences such as LINEs and SINEs, further increasing
genomic instability.

While widespread hypomethylation is a hallmark of cancer

cells, hypermethylation at CpG islands and inactivation of
certain genes, including tumor-suppressor genes (ST Table
3.2), are also found in many cancers, often in a tumorspecific pattern.
For example BRCA1 is
hypermethylated and
inactivated in breast and
ovarian cancer, and MLH1
is hypermethylated in some
forms of colon cancer.

 Inactivation of tumor-suppressor genes by

hypermethylation is thought to play an important

complementary role to mutational changes that
accompany the transformation of normal cells into
malignant cells.

For example, in a bladder cancer cell line, one allele of

the cell cycle control gene CDKN2A is mutated, and the

other, normal allele is inactivated by hypermethylation
of its CpG island.

The inactivation of both alleles allows these cells to

escape control of the cell cycle and divide continuously.

In many clinical cases, the combination of mutation
and hypermethylation occurs in familial forms of
cancer.

 However, genes other than tumor-suppressor genes are

also hypermethylated in some cancer cells; these include
genes that control or participate in DNA repair,
differentiation, apoptosis, and drug resistance.

In addition to altered patterns of methylation, many cancer

cells also have disrupted histone modification profiles.
In some cases, mutations in the genes encoding members of
the histone-modifying proteins histone acetyltransferase
(HAT) and histone deacetylase (HDAC) are linked to the
development of cancer.

initiating epigenetic changes leading to cancer may occur in

stem cells residing in normal tissue.
Three lines of evidence support this idea.
First, epigenetic mechanisms can replace mutations as a way
of silencing individual tumor-suppressor genes or activating
oncogenes.
Second, global hypomethylation may cause genomic
instability and the large-scale chromosomal changes that
are a characteristic feature of cancer.
Third, because epigenetic modifications can silence multiple
genes, they are more effective than serial mutations of single
genes in transforming normal cells into malignant cells.

A model of cancer based on epigenetic changes

in colon stem cells as initiating events in
carcinogenesis followed by mutational events is
shown in ST Figure 37

Epigenetics and the Environment

Environmental agents including nutrition, chemicals,

and physical factors can alter gene expression by

affecting the epigenetic state of the genome.

In mice, coat color is controlled by the dominant allele Agouti

(A). In homozygous AA mice, the allele is active only during a

specific time during hair development, producing a yellow
band on an otherwise black hair shaft, resulting in the agouti
phenotype.

A nonlethal mutant allele (Avy) causes yellow pigment

formation along the entire hair shaft, producing a yellow fur

color. This allele is the result of the insertion of a transposable
element near the transcription start site of the Agouti gene.

Epigenetics and the Environment

A promoter element within the transposon is responsible for

this change in gene expression.

The degree of methylation in the transposons promoter is

related to the amount of yellow pigment deposited in the hair

shaft and varies from individual to individual.
The result is a wide variation in coat color in genetically
identical mice (ST Figure 38), ranging from yellow
(unmethylated) to pseudoagouti (highly methylated).

In addition to a gradation in coat color, there is also a

gradation in body weight. Yellow mice are more obese than
the brown, pseudoagouti mice.

Epigenetics and the Environment

To evaluate the role of environmental factors in

modifying the epigenome, the diet of pregnant mice

was supplemented with methylation precursors,
including folic acid, vitamin B12, and choline.

In the offspring, variation in coat color was reduced

and shifted toward the pseudoagouti phenotype.

The shift in coat color was accompanied by increased

methylation of the transposons promoter.

Example 1: These two genetically identical mice were

born of genetically identical mothers who were fed
differently in pregnancy and they will have very
different lives

Their identical mothers were fed different amounts of

methylating nutrients or soy genistein during pregnancy

Epigenetics Occurs
LTR Hypomethylated
Transposon sequence

Yellow Mouse
High risk cancer, diabetes,
obesity & reduced lifespan

Maternal
Supplements
With
Genistein
zinc
methionine
betaine
choline,
folate
B12

LTR Hypermethylated
Transposon sequence

Agouti Mouse
Lower risk of cancer,
diabetes, obesity and
prolonged life

Increasing soy supplement genistein alters

gene expression and thus phenotype

Increasing Methylation
Change in coat color
Change to lower lifetime weight
Change to improved lifetime health

PNAS November 14, 2006 Vol. 103 no. 46 17-71-17072