Epigenetics of the male gamete
Douglas T. Carrell, Ph.D., H.C.L.D.
Departments of Surgery (Urology), Obstetrics and Gynecology, and Physiology, University of Utah School of Medicine, Salt Lake City, Utah
Objective: To review and summarize the current understanding of the epigenetic status of human sperm in regards to protamination, specific
localization and modifications of retained histones, and DNA methylation.
Design: Review of the relevant literature.
Setting: University-based clinical and research laboratories.
Patient(s): Fertile and infertile men.
Intervention(s): None.
Main Outcome Measure(s): Critical review of the literature.
Result(s): Sperm from normospermic, fertile men have epigenetic modifications consistent with gene ‘‘poising’’ at the promoters of genes involved in
development, including the localization of retained histones with bivalent histone modifications and hypomethylation of DNA. These epigenetic marks
are altered in some patients with abnormal spermatogenesis, and in some men who exhibit unexplained, altered embryogenesis during IVF therapy.
Conclusion(s): The sperm epigenome implies a poising of the paternal genome for embryogenesis and a possible role in the establishment of totipotency
of the embryo and may help in understanding some causes of reduced fertility and transmission of disease risk. (Fertil SterilÒ 2012;97:267–74. Ó2012 by
American Society for Reproductive Medicine.)
Key Words: Epigenetics, methylation, histones, protamines, transcription, embryogenesis
I
t is interesting, and perhaps of special note to reproductive biologists,
that the field of epigenetics is related
to one of the great historical debates of
embryology. This debate can be distilled
to a disagreement between whether
development of an organism was the
result of growth and expansion of
‘‘pre-formed’’ elements (the school of
‘‘preformation’’) or the result of chemical
reactions within cells that are sequentially executed to result in a mature
organism (the school of ‘‘epigenesis’’)
(1, 2). The discovery during the 20th
century that chromosomes contained
heritable components, followed by the
elucidation of the structure of DNA and
the deciphering of the genetic code,
progressively lent strong credence to
and helped refine the concept of
epigenesis, but how the identical
genetic sequences in the cells of an
embryo ultimately resulted in a fully
differentiated individual remained, and
continues to be, the central question of
developmental biology (3).
Epigenetics is defined as ‘‘the study
of mitotically and/or meiotically heritable changes in gene function that
cannot be explained by changes in
DNA sequence’’ (4). These ‘‘epigenetic’’
components not only have a profound
impact on developmental processes
but also have relevance in many diverse
areas of biology and medicine, including cancer biology, the study of environmental effects, and the study of
aging (5, 6).
Sperm cells are unique in their
morphology and function. Historically,
sperm have been viewed as specialized
delivery capsules containing, and perhaps protecting during transport, a
high-value cargo of 23 chromosomes
to the oocyte. Although the oocyte similarly contributes 23 chromosomes to
the embryo, the disparate sizes of the
two gametes, as well as the obvious unbalanced contribution of the oocyte to
the embryo’s complement of cellular
organelles, RNAs, and cellular machinery, have in some cases minimized the
Received December 13, 2011; revised December 19, 2011; accepted December 20, 2011.
D.T.C. has nothing to disclose.
Reprint requests: Douglas T. Carrell, Ph.D., H.C.L.D., Andrology and IVF Laboratories, University of
Utah School of Medicine, 675 S. Arapeen Dr. #205, Salt Lake City, UT 84108 (E-mail: douglas.
[email protected]).
Fertility and Sterility® Vol. 97, No. 2, February 2012 0015-0282/$36.00
Copyright ©2012 American Society for Reproductive Medicine, Published by Elsevier Inc.
doi:10.1016/j.fertnstert.2011.12.036
VOL. 97 NO. 2 / FEBRUARY 2012
potential contribution of the sperm to
successful embryogenesis (7). Nevertheless, some studies, and numerous anecdotal observations, have hinted at
a potential role of sperm contribution
to embryogenesis (8–10). Therefore,
understanding the epigenetics of sperm
may contribute to understanding
paternal effects on embryogenesis, as
well as a better understanding of germ
cell biology and the pluripotency of
embryonic stem cells.
This brief review will describe the
current understanding of sperm epigenetics by focusing on three main areas:
the extensive modifications to sperm
chromatin as a result of the removal
of histones during spermiogenesis and
their replacement with protamines, the
chemical modifications seen in histones retained in sperm chromatin,
and the methylation to the sperm
DNA itself. Each of these three types
of modifications that are present in
human sperm are potent modulators
of gene transcription, and recent data
suggest they may be factors that affect
transcriptional regulation during embryogenesis. The role of noncoding
RNAs, which are also found in sperm,
is reviewed by Hamatani et al. (11) in
a companion article. A brief discussion
of future avenues of study will be
highlighted.
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VIEWS AND REVIEWS
CHROMATIN MODIFICATIONS DURING
SPERMIOGENESIS
Normal human fertilization is a demanding event during
which sperm are required to traverse a potentially hostile
and barrier-laden female reproductive tract, penetrate the
cumulus oophorus, bind to and penetrate the zona pellucida,
fuse to the oolemma and penetrate into the oocyte, then complete multiple post-penetration events (12). To accomplish
these functions, the sperm cell has an extremely specialized
architecture, including dramatic changes to the chromatin
structure by replacement of most (90%–95%) of the histones
with protamines (13). Protamination of the sperm chromatin
facilitates the nuclear compaction necessary for sperm motility and helps to protect the genome from oxidizing and from
other harmful molecules within the female reproductive tract
(13). In addition, the higher order of packaging of the DNA
after protamination precludes transcriptional activity; therefore, protamination is a nontraditional form of epigenetic
regulation that is unique to sperm cells.
The transition of sperm nuclear proteins from canonical
histones to protamines is a multistep process that is still
poorly understood; however, certain events have been described. Early transition events include the replacement of
select histones with histones variants that are expressed during spermatogenesis, including testis-specific histone 2B, the
most abundant histone variant found in mature sperm (14).
Testis-specific histone 2B has been of particular interest
because of its unique expression pattern of not undergoing
30 -transcript polyadenylation and because it has been shown
through immunohistologic studies to locate in the telomeres
of sperm chromosomes (15).
Concomitant to replacement of some histones with variants, there is an increase of acetylation of other histones. This
hyperacetylation appears to be the trigger to the subsequent
cascade of events that ultimately results in protamine replacement (16). Acetylation is regulated by the interplay of both
acetylases and deacetylases, and one regulator appears to be
pygopus 2, which is a plant homeodomain protein that
acts as a cofactor to Wnt signaling effector complexes (17).
Hyperacetylation of histones results in a ‘‘relaxed’’ chromatin
structure that may be important in facilitating the role
of topoisomerase-induced strand breaks and removal of
histones and their replacement with transition proteins
(18, 19). Testis-specific bromodomain-containing protein
(BRDT) has been shown to interact with acetylated histone
and induce chromatin compaction and differential retention
of the hyperacetylated regions, thus appearing to be a key
factor in the reorganization of sperm chromatin (20, 21).
Transition proteins 1 and 2 (TP1 and TP2) are proteins of
intermediate basicity that are found in stages 12 and 13 of
spermatogenesis, but are removed by stage 14 of spermatogenesis. The proteins bind to the DNA, facilitate removal of
histones, and appear to be critical for subsequent events in
protamine compaction (22). Knock-out of one transition protein results in infertility of the affected mice; however, there
does appear to be some incomplete compensation by the remaining transition protein (23). Interestingly, sperm from
TP knock-out mice become less fertile as they progress
268
through the epididymis, hinting that a possible consequence
of abnormal TP expression may be progressively increased
DNA damage during epididymal transport (24).
Next, transition proteins are completely replaced by
protamines. Humans express two protamines, protamine 1
and protamine 2 (P1 and P2), which are expressed in roughly
equal quantities (25). There is considerable overlap in the
expression of the genes for transition proteins and protamines, and the proteins also show some overlap, but both
the transcripts and the proteins undergo unique translational
regulation that facilitates proper temporal regulation that
is necessary for proper protein function (26). Key posttranslational modifications include phosphorylation, which
is necessary for proper chromatin condensation (27).
Improper processing of the protamine transcripts can
result in increased retention of immature P2 precursors,
a condition associated with subfertility (28, 29). Subfertility
is also associated with an altered P1/P2 ratio (13, 30).
Although conflicting data have been reported, altered
protamine expression has been reported to be related to
a broad range of phenotypes, including diminished sperm
counts, decreased sperm function, and diminished embryo
quality during IVF (26, 31–33). It is of note that most studies
evaluating abnormal protamine expression have evaluated
the mean P1/P2 level for an ejaculate; however, protamine
replacement is clearly variable between sperm in a given
ejaculate (34, 35). It is of note that in infertile men with
abnormal protamination, density-gradient centrifugation selects, generally, for sperm with better protamination, also
highlighting the heterogeneity of protamination and the possible role of improved sperm selection techniques in improving
fertility therapies (36).
Proper protamine replacement results in a more efficient
chromatin packaging that results in a high level of compaction of the sperm nucleus. Protamine compaction occurs
through the formation of disulfide bonds between the protamines and by the formation of toroidal chromatin structures
(see Fig. 1) (37). Approximately 50 kb of DNA are packaged
in each toroidal subunit (38). However, 5%–10% of DNA in
fertile men, and more in some infertile men, remains bound
to histones (39, 40). Ward has recently proposed a unique
hypothesis on the structural arrangement of histone- and
protamine-bound regions of DNA in the mature sperm cell
in which each toroid is equivalent to one loop domain of
DNA with linker regions associated to matrix attachment regions (Fig. 1) (41, 42). This model emphasizes the protection of
protamine-bound DNA from damage by the toroidal compaction and subsequent stacking of toroids, and the vulnerability
of linker regions and histone-bound regions to DNA degradation by endonucleases (43). This structural packaging may
help explain the correlation between increased histone
retention and elevated DNA damage reported in several
studies (31, 44, 45).
The data indicate that protamination is critical for normal
sperm chromatin condensation and protection from damage
during transport, as well as an epigenetic role in silencing
the regions of the genome bound to protamines (46). Abnormal protamination is a relatively common event in infertile
men with abnormal semen analyses but relatively rare in
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Fertility and Sterility®
FIGURE 1
Chromatin remodeling and epigenetic modifications in human sperm. DNA methylation is the first line of epigenetic modification of chromatin
through methylation of position of cytosines found in CpG dinucleotides. An intermediate step in demethylation is the formation of
5-hydroxymethylcytosine residues, which are also observed in mature sperm. DNA is bound to histone octamers with unique modifications that
present a second level of regulation of gene transcription. Most histones are removed from the elongating spermatid and replaced with
protamines that result in a higher order of DNA packaging and silence gene expression. Retained histones are interspersed between protamine
toroids and may be bound to matrix attachment regions, which facilitates replication of loop domains in the embryo.
Carrell. Epigenetics of the male gamete. Fertil Steril 2012.
men with known fertility and normal semen quality (29, 47).
Protamination can be assessed using various techniques,
varying from histone-specific stains to purification and
quantification of the nuclear proteins, with a common reporting measure being the ratio of protamine 1 to protamine 2
(P1/P2 ratio) (48, 49). Generally, most reports have shown
that abnormal protamine replacement of histones is
associated with decreased semen quality, and in some
reports, diminished IVF embryo quality and pregnancy rates
(30, 33, 50, 51).
HISTONE VARIANTS AND MODIFICATIONS
IN HUMAN SPERM
Until recently, the utility of retaining histones in the sperm
nucleus was not understood and was largely considered the
result of inefficient or leaky protamination. Although protamination is a sperm-specific epigenetic modification, the
more classical epigenetic factors include histone modifications, DNA methylation variability, and the action of microVOL. 97 NO. 2 / FEBRUARY 2012
RNAs. Recent studies indicate that retained histones are
central to the epigenetic status of sperm, are not randomly
distributed throughout the genome, and that unique histone
modifications that help regulate genome activation and
silencing also appear to be programmatically distributed in
the sperm genome (39, 52, 53). These data imply that sperm
chromatin may transmit epigenetic mechanisms that affect
postfertilization transcriptional regulation (53).
Nucleosomes are octamers composed of two dimers of
H3-H4 histones and two dimers of H2A-H2B, with each
nucleosome linked by histone H1 (54). Histones are unique
proteins in that they are able to undergo relatively simple
chemical modifications that drastically alter their binding
to DNA and the accessibility of other regulatory factors to
the DNA, thereby altering gene activation (55). These chemical modifications are primarily accomplished by modifications of lysine and serine residues on the histone tail, and
through phosphorylation, methylation, acetylation, and ubiquitination (56, 57). In addition to chemical modifications,
tissue-specific histone variants are present, including
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VIEWS AND REVIEWS
testes-specific histone H2B, which is retained abundantly in
mature sperm (14). The modification of histones is dynamic
and dependent on specific enzymes, the regulation of which
is an active field of study (55). Histones conferring a generally
‘‘active’’ transcription state include acetylation of H3 and H4,
which generally facilitates an open chromatin structure and
binding of transcription factors (58). Deacetylation, conversely, is associated with an ‘‘inactive’’ transcriptional status,
and generally correlates with DNA methylation (59). Histone
lysine methylation is another key regulator of activation, with
H3K9 and H3K27 histones generally associated with ‘‘inactive’’ status (Table 1) (60).
Two recent studies characterized the distribution of
retained histones throughout the sperm genome. Arpanahi
et al. (61) used endonuclease digestion of human and mouse
sperm chromatin to identify endonuclease-sensitive regions
and found that endonuclease-sensitive regions were more
likely to be histone-bound regions that were largely found
to be associated with regulatory regions of the genome, including gene promoters. Our laboratory used a different approach to study this question, using repeated micrococcal
nuclease digestion and gel purification, which separates
histone-bound chromatin from protamine-bound chromatin.
This chromatin purification was followed by microarray analysis and deep sequencing of the histone-bound and
protamine-bound DNA to identify, at base pair resolution,
the location of histones. From this study, which was performed using sperm from fertile, normospermic controls, the
data clearly show enriched retention of the histones at the
promoters of gene families involved in development, along
with retention in micro-RNAs and imprinted genes. Conversely, protamines were not enriched at any gene family (39).
These two studies indicate a potential epigenetic function
for retained histones in the developing embryo; however, the
biological relevance of protamination and histone function
after fertilization has been questioned because sperm protamines are rapidly replaced by oocyte-derived histones in
the zygote (62). Therefore, we hypothesized that if an epigenetic role was associated with the retained histones, the location of specific histone variants or modifications would be
necessary and would further indicate a consistent, programmatic function. Indeed, subsequent chromatin immunoprecipitation purification of the histone classes, followed by
microarray and sequencing studies, elucidated an interesting
pattern in which key developmental genes were bivalently
marked with H3K4me3 and H3K27me3, similar to bivalently
marked developmental genes of embryonic stem cells
(39, 63). Regions with this bivalent marking are also DNA
demethylated (discussed in detail below), which together
strongly indicated a ‘‘poised’’ state.
Interestingly, regions of the genome enriched for testesspecific histone H2B, which makes up a significant portion
of retained histones, were enriched in the promoters of genes
for ion channels and genes involved in spermatogenesis, but
not developmental genes, indicating a ‘‘historical’’ epigenetic
record of spermatogenesis. Histone variant H2AZ, which has
been implicated in poising in certain cell types, was enriched
in pericentromeric heterochromatin regions, which had
previously been reported in immunostaining studies (8, 64).
270
Regions enriched with H3K4me3, but not bivalent for
H3K4me3 and H3K27Me, also appear to be enriched for
spermatogenesis-related genes, whereas H3K4me2 regions
are enriched for developmental genes. Thus, the histone packaging appears to be both a historical record of spermatogenesis and a future program for developmental processes (8).
It is of interest to note that a recent study characterized
histone modifications genomewide in zebrafish, a species
that does not use protamination to condense sperm chromatin. Wu et al. (65) reported that the zebrafish sperm genome
was similar to the human sperm genome in that there were
both activating and silencing histone modifications that
were similar to the patterns observed in human sperm. Activating modifications were seen at genes involved in spermatogenesis (a historical record), and in combination with
silencing modifications (bivalency or in some cases multivalency) at developmental gene promoters (a possible future
plan). Methylation patterns were also similar to human sperm
methylation. Wu et al. were also able to correlate the level of
activating marks with actual temporal expression of genes in
the embryo. This study is a fascinating validation of the concept of a sperm epigenetic role in a species distant from
humans with a distinct chromatin packaging organization
void of protamines and transition proteins (66).
SPERM DNA METHYLATION
DNA methylation is a potent epigenetic mark that promotes
gene silencing, is essential to allele-specific imprinting of certain genes, and is key in X chromosome inactivation (67, 68).
Methylation occurs at the 5-carbon position of cytosines
found in cytosine-phosphate-guanine dinucleotides (CpGs)
through the action of the DNMT family of proteins, which
are responsible for both the initiation of methylation
and subsequent maintenance of methylation marks (69).
Cytosine-phosphate-guanine dinucleotides found in high
concentrations near the gene promoter are termed ‘‘CpG
islands’’ and are potent inhibitors of transcription when
methylated (70). This inhibition or permissive role of
CpGs dependent on their methylation status is accomplished
largely through recruitment and/or binding of other factors
directly involved in transcription (71, 72). In addition,
genetic elements, including base pair sequence, influence
the methylation status of CpGs (73).
After fertilization, paternal DNA is actively demethylated
and the maternal DNA undergoes passive demethylation (74).
Although the demethylation is extensive, the specific regions
of demethylation are unknown. Subsequently, in the primordial germ cells, the maternal and paternal germ lines are reset,
resulting in remodeling of the parental-specific methylation
patterns of imprinted genes (75). Tissue-specific methylation
is also continued throughout development (Fig. 2) (76). Therefore, the role of sperm methylation in the epigenetic regulation of embryonic events is not well understood, and it will
require evaluation of the methylome of not only the sperm
but also of the embryo.
The genomewide study of sperm histone localization
described above also included an analysis of methylation status at gene promoters. Bivalently and monovalently marked
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Fertility and Sterility®
FIGURE 2
Summary of critical chromatin and DNA methylation modification in sperm, the embryo, and primordial germ cells. Key chromatin changes are
demonstrated by the protamination of sperm during spermiogenesis, followed by replacement of protamines with histones at the pronuclear
stage of embryogenesis. The active demethylation of sperm DNA after fertilization is contrasted with passive demethylation of maternally
derived DNA. Resetting of imprinting occurs in the primordial germ cells and tissue-specific methylation changes occur throughout development.
Carrell. Epigenetics of the male gamete. Fertil Steril 2012.
promoters of developmental genes were shown to be hypomethylated regions of DNA, consistent with the ‘‘poised’’ state of
bivalently marked genes of embryonic stem cells (39). This
finding supports the hypothesis of a potential embryonic
role for the sperm genome, although much is yet to be studied
and determined. Recently, the methylome of human sperm
has been reported (77). This study confirmed the generally hypomethylated state of developmental gene promoters and
highlighted that these hypomethylated regions are broader
and more deeply hypomethylated than hypomethylated promoter regions seen in somatic cells (77).
Although the studies described above are associational
only, they complement a potential epigenetic hypothesis of
epigenetic programming in sperm in which key developmental genes are poised, but not yet actively marked, for rapid activation in development by their hypomethylation of DNA at
regulatory regions along with simultaneous activating and silencing histone modifications, as opposed to the vast majority
of the genome that is silenced by protamine binding and
a small fraction of the genome bound to histone with silencing modifications. These promoter regions of DNA are also
hypomethylated, further poising the gene for activation (78).
INVESTIGATION OF CHROMATIN
MODIFICATIONS IN SPERM OF INFERTILE MEN
Studies have shown a small, but significantly increased, risk of
offspring with imprinting disorders in couples undergoing IVF
VOL. 97 NO. 2 / FEBRUARY 2012
and intracytoplasmic sperm injection (79, 80). Therefore,
the methylation status of imprinted genes in the sperm of
infertile men has been studied extensively and generally
shown altered methylation in the imprinted genes of
oligozoospermic men compared with controls (81–83).
Elevated methylation defects of imprinted genes has also
been observed in other pathologies, including men with
obstructive azoospermia undergoing testicular aspiration of
sperm, men with known defects of protamination, and
patients with idiopathic infertility (84–86). It is important
to consider that the rates of abnormalities seen in the sperm
of infertile men are manyfold higher than the rates of
affected offspring, which highlights the potential corrective
abilities of the embryo and primordial germ cells in
methylation ‘‘resetting.’’ In addition, the selection of the most
morphologically normal sperm during intracytoplasmic
sperm injection from the heterogeneous pool of sperm in
a given ejaculate may decrease the risk of transmission of the
observed methylation defects and be a factor in the disparity
of sperm abnormalities versus transmission of disease.
Given the poising nature of developmental genes, it is
also of interest to study the methylation status of nonimprinted genes, particularly development-related genes
and micro-RNAs. Our laboratory recently reported that no
gross methylation defects were seen, on average, in the promoters of OCT4, SOX2, NANOG, HOCC11, and miR-17 in
men with known abnormalities of protamination, a population previously shown to have elevated risk of defects of
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VIEWS AND REVIEWS
TABLE 1
Summary of activating and silencing histone modifications relevant
to sperm.
Modification
Acetylation of H3
Acetylation of H4
Ubiquitination of H2B
H3K4me2
H3K4me3
Ubiquitination of H2A
H3K9me
H3K27me3
H3K4me3 and H3K27me3
Activating
Silencing
Bivalency
X
X
X
X
X
X
X
X
X
Carrell. Epigenetics of the male gamete. Fertil Steril 2012.
methylation of imprinted genes (86, 87). However, defects were
observed in the CREM gene, which may be more of a reflection
and/or cause of abnormal spermatogenesis (87, 88). Recently,
we have evaluated the methylation status of sperm
genomewide in two populations of infertile patients, men
with abnormal protamination and men with unexplained
poor embryogenesis during IVF therapy. Interestingly, gross
abnormalities were observed in 3 of 28 of the men analyzed,
which was confirmed by subsequent bisulfate sequencing
(89). Sperm methylation studies are clearly in their nascent
stage, but highlight the known association of methylation
defects with male infertility. Whether the association is
biologically relevant remains to be shown.
Although numerous studies have shown abnormal protamination in the sperm of some infertile men, only one study
has yet to evaluate localization of histones and characterization of histone modifications genomewide in infertile men
(40). Because of the cost associated with such studies, only
seven patients were evaluated in this study, four men with
known abnormalities of protamination and three men with
unexplained, consistently poor embryogenesis during IVF.
A gross lack of enrichment of histones at developmental
gene promoters was observed in five of the seven patients,
but defects of histone modifications were more subtle, in
that defects were observed at specific genes but not genomewide. Methylation of developmental genes was altered in
some cases, but was also more subtle in most cases (40).
These limited studies of infertile men seem to support the
hypothesis that sperm epigenetic marks may be programmatic, at least in part, and imply a function during embryogenesis. The observation of abnormalities in IVF patients
with unexplained poor embryogenesis is interesting, but it
must be confirmed in much larger studies of highly characterized patients. Such studies have been limited because of the
costs and difficulties in bioinformatics and data analysis of
large genomewide studies. Current efforts are aimed at identifying select candidate alleles that are key factors in embryogenesis, or that are representative of the genome at large, and
are predictive of abnormal epigenetic function.
CONCLUSIONS
The sperm cell has a highly differentiated and specialized
morphology that is essential to facilitate fertilization.
272
Similarly, the epigenome of human sperm is unique, elegant,
and may be essential to embryogenesis. Key elements of the
sperm epigenome include the replacement of most canonical
histones with protamines, the elegant retention of histones at
key regulatory regions of the genome in a poised state that
suggests unique and significant functions in early development, and in complementary hypomethylation of DNA at
such regions. Not discussed in this short review, but likely
an equally important epigenetic modifier in sperm, are noncoding RNAs found in mature sperm. These epigenetic factors
suggest a diverse and critical role of sperm in regulating
embryogenesis.
Advancement of the field of sperm epigenetics will be
dependent on further studies on the basic science of epigenetics, as well as translational studies. The combination of
these studies may result in a much deeper understanding of
the possible causes of abnormal embryogenesis that is so frequently seen in the IVF clinic, as well as possible causes of
recurrent pregnancy loss, developmental anomalies, and
other pathologies. In addition, the understanding of the
epigenetics of sperm and spermatogonia may be key in understanding the mechanisms of pluripotency, which has broad
implications for potential therapies.
The sperm epigenome has been characterized before the
oocyte or early embryo epigenomes because of the relative
ease of obtaining sufficient cells to undertake genomewide
studies. However, techniques are now evolving that will allow
genomewide assessment of embryos and oocytes, in fact the
initial studies of methylation in the oocyte have recently
been reported and further studies are ongoing (74, 90).
Key to understanding sperm epigenetic ramifications will
be a better understanding of the extent and specifics of
remodeling that takes place after fertilization. Future studies
will likely focus on the epigenetics of both gametes as well
as changes observed throughout embryogenesis.
Epigenetic regulation is the presumed link between environmental factors and many observed effects (91). Therefore,
environmental studies will likely play an ever-increasing role
in the possible causes of infertility and in the assessment of
risk factors. Included in such studies will be studies evaluating
the aging process on epigenetics, and on the heritability of
such epigenetic modification. The studies listed above, in
addition to further characterization studies, will soon lead
to a better understanding of the risk of ‘‘subtle epigenetic
abnormalities’’ that are frequently observed and reported in
the literature. Such an understanding will be key in the translation of epigenetics to a useful clinical tool.
Acknowledgments: I thank Bill Kelly, Ben Emery, and Ki
Aston for their assistance in the development of the figures
used in this review.
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