The science of change Molecular biology

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EPIGENETICS: THE SCIENCE OF CHANGE
MOLECULAR BIOLOGY
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INTRODUCTION
The word “epigenetic” literally means “in addition to changes in genetic sequence.” The term
has evolved to include any process that alters gene activity without changing the DNA sequence,
and leads to modifications that can be transmitted to daughter cells (although experiments show
that some epigenetic changes can be reversed). There likely will continue to be debate over
exactly what the term means and what it covers.
MULTIPLE MECHANISMS
Many types of epigenetic processes have been identified, they include:
 Methylation
 Acetylation
 Phosphorylation
 Ubiquitylation
 Sumolyation.
Other epigenetic mechanisms and considerations are likely to surface as work proceeds.
Epigenetic processes are natural and essential to many organism functions, but if they occur
improperly, there can be major adverse health and behavioral effects.
Perhaps the best known epigenetic process, in part because it has been easiest to study with
existing technology, is DNA methylation. This is the addition or removal of a methyl group
(CH3), predominantly where cytosine bases occur consecutively. DNA methylation was first
confirmed to occur in human cancer in 1983, and has since been observed in many other
illnesses and health conditions.
Another significant epigenetic process is chromatin modification. Chromatin is the complex of
proteins (histones) and DNA that is tightly bundled to fit into the nucleus. The complex can be
modified by substances such as acetyl groups (the process called acetylation), enzymes, and
some forms of RNA such as microRNAs and small interfering RNAs. This modification alters
chromatin structure to influence gene expression. In general, tightly folded chromatin tends to be
shut down, or not expressed, while more open chromatin is functional, or expressed.
One effect of such processes is imprinting. In genetics, imprinting describes the condition where
one of the two alleles of a typical gene pair is silenced by an epigenetic process such as
methylation or acetylation. This becomes a problem if the expressed allele is damaged or
contains a variant that increases the organism’s vulnerability to microbes, toxic agents, or other
harmful substances. Imprinting was first identified in 1910 in corn, and first confirmed in
mammals in 1991.
Researchers have identified about 80 human genes that can be imprinted, although that number is
subject to debate since the strength of the evidence varies. That approximate number isn’t likely
to rise much in years to come.
SUBSTANTIAL CHANGES

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Most epigenetic modification, by whatever mechanism, is believed to be erased with each new
generation, during gameto-genesis and after fertilization. However, one of the more startling
reports published in 2005 challenges this belief and suggests that epigenetic changes may endure
in at least four subsequent generations of organisms. Other studies have found that epigenetic
effects occur not just in the womb, but over the full course of a human life span. Younger twin
pairs and those who shared similar lifestyles and spent more years together had very similar
DNA methylation and histone acetylation patterns. But older twins, especially those who had
different lifestyles and had spent fewer years of their lives together, had much different patterns
in many different tissues, such as lymphocytes, epithelial mouth cells, intra-abdominal fat, and
selected muscles.
OTHER DRIVERS OF CHANGE
Substances aren’t the only sources of epigenetic changes. The licking, grooming, and nursing
methods that mother rats use with their pups can affect the long-term behavior of their offspring,
and those results can be tied to changes in DNA methylation and histone acetylation at a
glucocorticoid receptor gene promoter in the pup’s hippocampus. Along with behavior, mental
health may be affected by epigenetic changes. The past decade has also been productive in
developing strong links between aberrant DNA methylation and aging. Some of the strongest,
decade-old evidence shows progressive increases in DNA methylation in aging colon tissues,
and more recent evidence links hypermethylation with atherosclerosis. Altered, age-related
methylation has also been found in tissues in the stomach, esophagus, liver, kidney, and bladder,
as well as the tissue types.
LINKS TO DISEASE
Among all the epigenetics research conducted so far, the most extensively studied disease is
cancer, and the evidence linking epigenetic processes with cancer is becoming “extremely
compelling,”.
Many other health issues have drawn attention. Epigenetic immune system effects occur, and can
be reversed, according to research published in the November–December 2005 issue of the
Journal of Proteome Research by Nilamadhab Mishra, an assistant professor of rheumatology at
the Wake Forest University School of Medicine, and his colleagues. The team says it’s the first
to establish a specific link between aberrant histone modification and mechanisms underlying
lupus-like symptoms in mice, and they confirmed that a drug in the research stage, trichostatin A,
could reverse the modifications. The drug appears to reset the aberrant histone modification by
correcting hypo-acetylation at two histone sites.
In studies published in the May–August 2004 issue of International Reviews of Immunology and
the October 2003 issue of Clinical Immunology, it was noted that pharmaceuticals such as the
heart drug pro-cinamide and the anti-hypertensive agent hydralazine cause lupus in some people,
and demonstrated that lupus-like disease in mice exposed to these drugs is linked with DNA
methylation alterations and interruption of signaling pathways similar to those in people.
EPIGENETICS IN CANCER: TARGETING CHROMATIN MODIFICATIONS
Post-translational modifications to histones affect chromatin structure and function resulting in
altered gene expression and changes in cell behavior. Aberrant gene expression and altered

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epigenomic patterns are major features of cancer. Epigenetic changes including histone
acetylation, histone methylation, and DNA methylation are now thought to play important roles
in the onset and progression of cancer in numerous tumor types. Indeed deregulated epigenetic
modifications, especially in early neoplastic development, may be just as significant as genetic
mutations in driving cancer development and growth. The reversal of aberrant epigenetic
changes has therefore emerged as a potential strategy for the treatment of cancer. A number of
com-pounds targeting enzymes that regulate histone acetylation, histone methylation, and DNA
methylation have been developed as epigenetic therapies, with some demonstrating efficacy in
hematological malignancies and solid tumors. This review highlights the roles of epigenetic
modifications to histones and DNA in tumorigenesis and emerging epigenetic therapies being
developed for the treatment of cancer.
Cancer can evolve from a combination of epigenetic and genetic abnormalities resulting in
deregulated gene expression and function. The most common epigenetic modifications observed
are increased methylation of CpG islands within gene promoter regions and deacetylation and or
methylation of histone proteins (1–3). This review will focus on the role altered epigenetic
regulation plays in mediating tumor onset and progression and the development of compounds
that tar-get enzymes that regulate the epigenome as anticancer agents.
Chromatin is a highly ordered structure consisting of repeats of nucleosomes connected by linker
DNA. Chroma-tin consists of DNA, histones, and nonhistone proteins condensed into
nucleoprotein complexes and it functions as the physiological template of all eukaryotic genetic
information.
Chromatin is divided into two distinct conformation states:
(1) Heterochromatin, which is densely compacted and transcriptionally inert
(2) Eu-chromatin, which is de-condensed and transcriptionally active.
Histones are small basic proteins containing a globular domain and a flexible charged NH2
terminus known has the histone tail, which protrudes from the nucleosome. Regulation of gene
expression occurs through posttranslational modifications of the histone tails provided by
covalent modifications including acetylation, methylation, phosphorylation, ubi-quitination,
sumoylation, proline isomerization, and ADP-ribosylation).
Post-translational modifications to histone tails govern the structural status of chromatin and the
resulting transcriptional status of genes within a particular locus. In addition, hyper-methylation
of CpG dinucleo-tides within promoter regions also plays an important role in controlling gene
expression. It is the complex inter-play of posttranslational modification of specific residues on
histone tails coupled with the DNA methylation status at a particular locus that determines if a
particular gene(s) is transcriptionally active or repressed. Chromatin remodeling and DNA
methylation is a highly regulated process con-trolled by enzymes that often exist in large
macromolecular complexes. Posttranslational modifications such as histone acetylation and
methylation and DNA methylation do not necessarily occur as mutually exclusive processes
controlled by the sequential recruitment of different enzymes to a specific genomic region, but
are more likely to be a dynamic process directed by large protein complexes containing different
epigenetic enzymes. DNA methylation, histone modification, and subsequent modulation of

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nucleosome positioning are interlinked processes that act in a coordinated manner to determine
the transcriptional status of a particular gene.
EPIGENETIC DEREGULATION IN CANCER DEVELOPMENT
Epigenetic mechanisms controlling transcription of genes involved in cell differentiation,
proliferation, and survival are often targets for deregulation in malignant development. In
addition to affecting transcription of protein-encoding mRNAs, noncoding microRNAs (miRs)
that can regulate the expression of a myriad of cellular proteins by affecting mRNA stability
and/or translation are similarly modulated by epigenetic means altered patterns of epigenetic
modifications are common in many human diseases including cancer and there is evidence that
epigenetic dysregulation can be a preliminary transforming event. For example epigenetic
changes such as global DNA hypomethylation and promoter-specific hypermethy-lation are
commonly observed in benign neoplasias as well as early-stage tumors. This suggests that
epigenetic alterations are early events in the loss of cellular homeostasis, and may in some
instances precede genetic mutations and genomic instability.
The deregulation of epigenetic modifiers has been characterized in many malignancies and the
disruption of a number of histone modifying proteins, by mutations, deletions, or overexpression,
is supportive of the critical role of epigenetic effectors in oncogenesis. Indeed individual genes,
entire sets of genes, and miRs may be epigenetically deregulated to promote the “hallmarks of
cancer” such as self-sufficiency in growth signals, insensitivity to growth inhibitory signals,
evasion of apoptosis, increased proliferation potential, sustained angiogenesis, and capability of
metastasis and invasion (8, 10). As detailed below, genetic abnormalities often result in the
deregulated localization of enzymes that actively control DNA methylation or post-translational
modifications of histone tails to specific loci. In these instances, loss of epigenetic regulation
occurs down-stream of oncogenic disruptions to the genome. In instances involving mutation of
a single allele, epigenetic silencing of the other allele can provide a second “hit,” resulting in loss
of heterozygosity (LOH) and inactivation of tumor suppressor genes. Moreover, silencing of
genes can cooperate with oncogenic mutations to promote tumor development and growth.
Interestingly, it is becoming apparent that the expression of epigenetic-regulatory enzymes such
as DNA methyltransferases (DNMT), histone deacetylases (HAT), and histone methyl-
transferases (HMT) can be controlled by miRs.
Deregulated epigenetic mechanisms may initiate genetic instability, resulting in the acquisition
of genetic mutations in tumor-suppressor genes and activating genetic mutations in oncogenes.
Moreover, epigenetic disruptions in tumors are generally of a clonal nature, indicating
occurrence in early generations of cells. There is a strong causative link between the silencing of
genes involved in DNA re-pair and cell transformation. In addition to negative effects on DNA
repair, pathways such as the Wnt/β-catenin pathway that regulates cell proliferation and
epithelial-to-mesenchymal transition (EMT) can be epigenetically controlled. There is clearly
growing awareness of the importance of epigenetic deregulation in early cancer predisposition
and development as evidenced by the growing list of genes with tumor sup-pressor activity that
are often epigenetically silenced but rarely genetically mutated in the pre-invasive stages of
many cancers.
DNA METHYLATION AND CANCER

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The maintenance of appropriate DNA methylation within CpG dinucleotide islands plays a
significant role in regulation of a wide variety of molecular processes including stability of
chromosomal structure and control of gene expression. In general, DNA methylation in gene
promoter regions results in gene silencing likely because of steric inhibition of transcription
complexes binding to regulatory DNA. DNA methylation can also result in the recruitment of
proteins that bind methylated CpG sequences (methyl-CpG-binding domain [MBD] proteins)
complexed with histone deacetylases (HDACs) and HMTs prompting coordinated epigenetic
modifications of the surrounding chromatin.
Abnormal patterns in DNA methylation were the first examples of epigenetic deregulation to be
characterized in human cancers, either as a result of DNMT overexpression or aberrant
recruitment. Tumor cell-specific promoter hypermethylation in genes that play important roles in
regulating cell cycle, apoptosis, DNA repair, differentiation, and cell adhesion is often a
hallmark of diseases. In addition, hypomethylation of repetitive sequences may result in
chromosomal and genetic instability, leading to further oncogenic events. Transcriptional
silencing via DNA hypermethylation can often be associated with poor clinical outcome in
several malignancies. For example silencing of CDKN2A and CDKN1A has been associated
with poor clinical outcome in acute leukemias . The development of high-throughput approaches
such as methylated DNA immunoprecipitation (Methyl-DIP) and differential methylation
hybridization (DMH) to survey the epigenome of normal and cancer cells for alterations in
methylated regions of the genome should allow for the identification of other cancer-related
silenced genes that will expand our knowledge of the epigenomic changes that occur during
cellular transformation.
HISTONE MODIFICATIONS
Although a number of histone modifications undoubtedly playimportant roles in epigenetic
deregulation, acetylationand methylation are the two histone modifications that have been
clinically associated with pathological epigenetic disruptions in cancer cells. In particular, the
loss of acetylation and methylation of specific residues in core his-tones H3 and H4 have been
identified as a marker of tumor cells.
HISTONE ACETYLATION AND CANCER
The acetylation status of histones H3 and H4 seem to largely dictate the fate of chromatin
assembly, transcription, and gene expression. Histone acetylation is governed by the opposing
activities of HATs and HDACs. Three main families of HATs transfer acetyl groups to lysine
residues ofthe nucleosome core histones: the MOZ/YBF2/SAS2/TIP60 (MYST) family, the
GCN5 N-acetyltransferase (GNAT) family, and the CBP/p300 family. HATs are recruited as co-
activators of transcription by transcription factors, usually in the context of large chromatin
remodeling complexes.
Altered HAT activity has been reported in both hematological and solid cancers, by inactivation
of HAT activity through gene mutation or through deregulation of HAT activity by viral
oncoproteins.
TARGETING CANCER VIA EPIGENETIC THERAPY
In contrast to genetic mutations, most epigenetic modifications may be reversible and
preventable. The resetting of aberrant epigenetic states in neoplastic cells is an expanding

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therapeutic approach to treat or prevent cancer. Pharmacological targeting of DNA methylation
and histone acetylation and methylation is now possible and is a promising therapeutic approach.
INHIBITORS OF DNA METHYLATION
Hydralazine was originally approved for use as an antihypertensive, and recently reported to be
an inhibitor of DNA methylation able to reactivate the expression of tumor-suppressor genes in
cancer. Pharmacologic inhibition of DNA methylation causes the trapping of DNMTs and their
targeted degradation resulting in re-expression of genes that have been aberrantly silenced by
hypermethylation, concomitant with inhibition of clonal expansion and tumor cell growth,
induction of cell differentiation, and cancer cell death. How DNMT inhibitors (DNMTi)
specifically affect tumor cells is not well understood however as these agents function by being
incorporated into newly replicated DNA, only rapidly dividing cells such as tumor cells will be
targeted. Moreover, as discussed above, the epigenetic silencing of tumor suppressor genes can
be an essential oncogenic event and may therefore result in “addiction” of the transformed cell to
the silencing of specific genes. Accordingly, tumor cells may be exquisitely sensitive to the
reversion of this gene silencing phenotype by DNMTi. It is currently not known if the
reactivation of epigenetically silenced genes is the only molecular event that underpins the
biological and therapeutic effects of DNMTi, and if so if specific reactivation of only one or a
number of epigenetically silenced genes is necessary. Extensive clinical studies indicate that
DNMTi induce manageable short-term side effects at doses that show therapeutic efficacy.
However, the long-term effects of chronic epigenetic deregulation and inhibition of DNMT
activity remains to be fully evaluated.
EPIGENETIC THERAPIES IN COMBINATION
Cooperation between different epigenetic modifications in driving oncogenic gene expression
supports the rationale of combining epigenetic therapies. Both HDAC inhibition and DNA de-
methylating agents have shown clinical efficacy as single agents; yet combination of the two
therapies has been shown to have strong synergistic effects on the reactivation of silenced genes
and anti-proliferative and cyto-toxic effects on cancer cells. Although de-methylating agents
have shown clinical efficacy in a subset of hematologic tumors, there is evidence that this
treatment does not fully revert aberrant epigenetic states and may not protect against the
recurrence of aberrant gene silencing directed by chromatin modifications. The dense
methylation of genes precludes the activation of gene expression by HDAC inhibitors as single
agents; however, combination with DNA demethylating agents has shown synergistic effects in
inducing the expression of heavily methylated genes and inhibiting cancer cell proliferation and
survival. Such combinations are currently being investigated in a number of clinical trials.
Treatments targeting epigenetic processes can potentiate the effects of other antineoplastic
treatments, including traditional chemotherapy and radiation. Examples of such combinations
include epigenetic therapies with agents targeting microtubule stability (docetaxel, paclitaxel),
proteosomal degradation (bortezomib), and molecular protein chaperones (geldanomycin) . The
rationale for these combinations is based on the observation that epigenetic therapies, especially
HDAC inhibition, lower the apoptotic threshold of tumor cells, making them more sensitive to
other agents.

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Based on the rationale that epigenetic deregulation is a driver of tumorigenesis, the combination
of epigenetic therapies with targeted anticancer treatments is also under clinical investigation. In
preclinical models, combinations of HDAC inhibitors with targeted therapies such as signal
transduction inhibitors have consistently shown additive or synergistic effects on induction of
apoptosis in cancer cell lines and in vivo tumor models. This includes combination of epigenetic
with targeted therapies such as imatinib in CML, gefitinib in lung cancer, and tras-tuzumab in
breast cancer, among others.
An appreciation of the significant role of epigenetic defects in cancer onset and progression has
increased remarkably in recent years. It is now understood that deregulated epigenetic
mechanisms can cause, as well as compound, the effects of oncogenic mutations to promote
tumor develop-ment and growth. The management of aberrant epigenetic states as a way to
target early tumor development as well as tumor progression is therefore a logical therapeutic
approach. The efficacy of epigenetic therapies in the treatment of myelodysplastic syndromes
and prevention of leukemic transformation reinforces the importance of epigenetic deregulation
prior to cancer onset. As such, epigenetic therapy is a promising approach for the prevention and
treatment of malignancies. One of the most exciting aspects of epigenetic therapy is the ability to
potentiate responses to existing therapies, which effectively multiplies the arsenal against cancer
progression. An understanding of the link between epigenetic deregulation and cancer is
applicable to prognosis as well as treatment. Further definition and refinement of profiles of
histone and DNA modification patterns should be invaluable for the purposes of detection,
diagnosis, and prognosis of cancer as well as the prediction of therapeutic responses.
MUTATED SILENCERS, HEREDITARY DISEASES, AND THEIR EFFECTS
Genetic mutations occur when nucleotide sequences in an organism are altered. These mutations
lead to not only observable phenotypic influences in an individual, but also alterations that are
undetectable phenotypically. The sources for these mutations can be errors during replication,
spontaneous mutations, and chemical and physical mutagens (UV and ionizing radiation,
heat).Silencers, being encoded in the genome, are susceptible to such alterations which, in many
cases, can lead to severe phenotypical and functional abnormalities. In general terms, mutations
in silencer elements or regions could lead to either the inhibition of the silencer’s action or to the
persisting repression of a necessary gene. This can then lead to the expression or suppression of
an undesired phenotype which may affect the normal functionality of certain systems in the
organism. Among the many silencer elements and proteins, REST/NSRF is an important silencer
factor that has a variety of impacts, not only in neural aspects of development. In fact, in many
cases, REST/NSRF acts in conjunction with RE-1/NRSE to repress and influence non-neuronal
cells. Its effects range from frogs (Xenopuslaevis) to humans, with innumerous effects in
phenotype and also in development. In Xenopuslaevis, REST/NRSF malfunction or damage has
been associated to abnormal ectodermal patterning during development and significant
consequences in neural tube, cranial ganglia, and eye development. In humans, a deficiency in
the REST/NSRF silencer element has been correlated to Huntington's disease due to the decrease
in the transcription of BDNF.
Furthermore, ongoing studies indicate that NRSE is involved in the regulation of the ANP gene,
which when over expressed, can lead to ventricular hypertrophy.Mutations in the Polycomb-
group (PcG) complexes also presented significant modifications in physiological systems of

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organisms. Hence, modification in silencer elements and sequences can result in either
devastating or unnoticeable changes.Specialized cells called the notochord
(a) Induces ectoderm above it to become the primitive nervous system.
(b) Neural tube forms
(c) Gives rise to the brain and spinal cord.
(d) Neural crest cells will migrate to different regions throughout the embryo to initiate
development of glia, pigments, and other neural structures.
Abnormal ectoderm patterning will cause abnormal and no neural folding.
REST/NRSF in Xenopuslaevis
The effects and influences of RE1/NRSE and REST/NRSF are significant in non-neuronal cells
that require the repression or silencing of neuronal genes. These silencer elements also regulate
the expression of genes that do not induce neuron-specific proteins and studies have shown the
extensive impact these factors have in cellular processes. In Xenopuslaevis, RE1/NRSE and
REST/NRSF dysfunction or mutation demonstrated significant impact on neural tube, cranial
ganglia, and eye development. All of these alterations can be traced to an improper patterning of
the ectoderm during Xenopus development. Thus, a mutation or alteration in either the silencing
region RE1/NRSE or silencer REST/NRSF factor can disrupt the proper differentiation and
specification of the neuroepithelial domain and also hinder the formation of skin or
ectoderm.The lack of these factors result in a decreased production of bone morphogenetic
protein (BMP), which translates into a deficient development of the neural crest. Hence, the
effects of NRSE and NRSF are of fundamental importance for neurogenesis of the developing
embryo, and also in the early stages of ectodermal patterning. Ultimately, inadequate functioning
of these factors can result in aberrant neural tube, cranial ganglia, and eye development in
Xenopus.
REST/NSRF AND HUNTINGTON'S DISEASE
Huntington's disease (HD) is an inherited neurodegenerative disorder, with symptoms emerging
during an individual’s mid-adulthood. The most noticeable symptoms of this progressive disease
are cognitive and motor impairments, as well as behavioral alterations.These impairments can
develop into dementia, chorea, and eventually death. At the molecular level, HD results from a
mutation in the huntingtin protein (Htt). More specifically, there is an abnormal repetition of a
CAG sequence towards the 5’-end of the gene, which then leads to the development of a toxic
polyglutamine (polyQ) stretch in the protein. The mutated Htt protein affects an individual’s
proper neural functions by inhibiting the action of REST/NRSF.
REST/NRSF is an important silencer element that binds to regulatory regions to control the
expression of certain proteins involved in neural functions. The mechanistic actions of huntingtin
are still not fully understood, but a correlation between Htt and REST/NRSF exists in HD
development. By attaching to the REST/NRSF, the mutated huntingtin protein inhibits the action
of the silencer element, and retains it in the cytosol. Thus, REST/NRSF cannot enter the nucleus
and bind to the 21 base-pair RE-1/NRSE regulatory element. An adequate repression of specific
target genes are of fundamental importance, as many are involved in the proper development of
neuronal receptors, neurotransmitters, synaptic vesicle proteins, and channel proteins. A
deficiency in the proper development of these proteins can cause the neural dysfunctions seen in
Huntington's disease. In addition to the lack of repression due to the inactive REST/NRSF,

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mutated huntingtin protein can also decrease the transcription of the brain-derived neurotropic
factor (BDNF) gene. BDNF influences the survival and development of neurons in the central
nervous system as well as the peripheral nervous system. This abnormal repression occurs when
the RE1/NRSE region within the BDNF promoter region is activated by the binding of
REST/NRSF, which leads to the lack of transcription of the BDNF gene.Hence, the anomalous
repression of the BDNF protein suggests a significant impact in Huntington's disease.
CURRENT RESEARCH ON REST/NRSF AND VENTRICULAR HYPERTROPHY IN
MAMMALS
REST/NRSF in conjunction with RE1/NRSE also acts outside the nervous system as regulators
and repressors. Current research has linked RE1/NRSE activity with the regulation of the
expression of the atrial natriuretic peptide (ANP) gene.An NRSE regulatory region is present in
the 3’ untranslated region of the ANP gene and acts as a mediator for its appropriate expression.
The protein encoded by the ANP gene is important during embryonic development for the
maturation and development of cardiac myocytes. However, during early childhood and
throughout adulthood, ANP expression is suppressed or kept to a minimum in the ventricle.
Thus, an abnormal induction of the ANP gene can lead to ventricular hypertrophy and severe
cardiac consequences. In order to maintain the repression of the gene, NRSF (neuron-restrictive
silencer factor) or REST binds to the NRSE region in the 3’untranslated region of the ANP gene.
Furthermore, the NRSF-NRSE complex recruits a transcriptional corepressor known as
mSin3.This leads to the activity of histone deacetylase in the region and the repression of the
gene. Therefore, studies have revealed the correlation between REST/NRSF and RE1/NRSE in
regulating the ANP gene expression in ventricular myocytes. A mutation in either the NRSF or
NRSE can lead to an undesirable development of ventricular myocytes, due to lack of repression,
which can then cause ventricular hypertrophy. Left ventricular hypertrophy, for example,
increases an individual’s chance of sudden death due to a ventricular arrhythmia resulting from
the increased ventricular mass.[13] In addition to the influence on the ANP gene, the NRSE
sequence regulates other cardiac embryonic genes, such as brain natriuretic peptide BNP,
skeletal α-actin, and Na, K – ATPase α3 subunit. Hence, the regulatory activity of both NRSE
and NRSF in mammals prevents not only neural dysfunctions but also physiological and
phenotypical abnormalities in other non-neuronal regions of the body.
MUTATIONS IN POLYCOMB-GROUP RESPONSE ELEMENTS (PRES)
The Polycomb-group (PcG) regulatory complexes are known for their influence in the epigenetic
regulation of stem cells, especifically in hematopoietic stem cells. The Polycomb Repressive
Complex 1 (PRC 1) is directly involved in the process of hematopoiesis, and functions together
with, for example, the PcG gene “Bmi1”. Studies in mice indicate that organisms with mutated
“Bmi1” demonstrate deficient mitochondrial functioning, and also hindered the ability of
hematopoietic cells to self-renew. Likewise, mutations in PRC2 genes were related to
hematological conditions such as acute lymphoblastic leukemia (ALL), which is a form of
leukemia. Hence, Polycomb-group genes and proteins are involved in the proper maintenance of
hematopoiesis in the body.
CURRENT AND FUTURE QUANDARIES
The accumulated evidence indicates that many genes, diseases, and environmental substances are
part of the epigenetics picture. However, the evidence is still far too thin to form a basis for any

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overarching theories about which substances and which target genes are most likely to mediate
adverse effects of the environment on diseases.
At the FDA, scientists are investigating many drugs that function through epigenetic mechanisms
(although as spokes-woman Christine Parker notes, the agency bases its approvals on results of
clinical trials, not consideration of the mechanism by which a drug works). One such drug,
azacitidine, has been approved for use in the United States to treat myelodysplastic syndrome, a
blood disease that can progress to leukemia. The drug turns on genes that had been shut off by
methylation. The drug’s epigenetic function doesn’t make it a “miracle drug,” however. Trials
indicate it benefits only 15% of those who take it, and a high percentage of people suffer serious
side effects, including nausea (71%), anemia (70%), vomiting (54%), and fever (52%).
Ehrlich points out that azacitidine also has effects at the molecular level—such as inhibiting
DNA replication and apoptosis—that may be part of its therapeutic benefits. The drug’s mixed
results might also be explained in part by a study published in the October 2004 issue of Cancer
Cell by Andrew Feinberg, director of the Johns Hopkins University Center for Epigenetics in
Common Human Disease, and his colleagues. They found that each of two tested drugs,
trichostatin A and 5-aza-2′-deoxycytidine (which is related to azacitidine), can turn on hundreds
of genes while also turning off hundreds of others. If that finding holds in other studies, it
suggests one key reason why it is so difficult to create a drug that doesn’t cause unintended side
effects.
PUBLIC AND PRIVATE
Despite the potentially huge role that epigenetics may play in human disease, investment in this
area of study remains tiny compared to that devoted to traditional genetics work. Several efforts
to change that are under way.
In Europe, the Human Epigenome Project was officially launched in 2003 by the Welcome Trust
Sanger Institute, Epigenomics AG, and the Centre National de Génotypage. The group’s focus is
on DNA methylation research tied to chromosomes 6, 13, 20, and 22. They may be joined soon
by organizations in Germany and India, where scientists plan to work on chromosomes 21 and
X, respectively, says Sanger senior investigator Stephan Beck.
But comprehensively studying all the epigenetic and epigenomic factors related to a multitude of
diseases and health conditions will take much more work. “A comprehensive Human Epigenome
Project is a lot more complicated than a Human Genome Project,” Jones says. “There’s only one
genome,[but an epigenome varies in each and every tissue.” The Human Genome Project was a
worldwide effort that took more than a decade and billions of dollars to complete.
In the United States, the National Cancer Institute and the National Human Genome Research
Institute formally kicked off a major effort 13 December 2005 that will include epigenomic
work. The pilot project of The Cancer Genome Atlas, funded by $50 million each from the two
institutes, is designed to lay the groundwork for comprehensive study of genomic factors related
to human cancer. The initial three-year effort is expected to focus on just two or three of the
more than 200 cancers known to exist, but if it’s successful in developing methods and
technologies, the number of cancers evaluated could then expand. If a high number of cancer

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genes are eventually scrutinized, the effort would be the equivalent of thousands of Human
Genome Projects.
TOOL TIME
If epigenetic work is to continue breaking new ground, many observers say technology will need
to continue advancing. that there must be additional improvements in high-throughput
technologies, analytical techniques, computational capability, mechanistic studies, and
bioinformatic strategies. They also say there is a need for basics such as standardized reagents
and a consistent supply of antibodies for testing.there is also a need to develop a comprehensive
tally of all proteins in the cell and to get better protein modification information. He says
universities are recognizing the demand for the talents needed to solve epigenomics problems,
and are increasing their efforts to cover these topics in various ways, especially at the graduate
school level.
Other groups are doing their part by creating tools to further the field. All the imprinted genes
identified so far are tracked in complementary efforts The European managers of the DNA
Methylation Database have assembled a compendium of known DNA methylations that,
although not comprehensive, still provides a useful tool for researchers investigating the roughly
22,000 human genes.
REFERENCES
 http://www.dnamethsoc.com/main.htm
 http://www.epigenome-noe.net
 http://www.epigenome.org
 http://www.landesbioscience.com/journals/epigenetics/
 http://www.methdb.de/front.html
 http://igc.otago.ac.nz/home.html
 http://www.geneimprint.com/databases/?c=clist
 http://www.mgu.har.mrc.ac.uk/research/imprinting/
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