The distinctive patterns of gene expression that are asso-
ciated with specialized embryonic and adult cell types,
as well as the modulation of specific gene programmes
in response to physiological and pathological signal-
ling, require multiple levels of transcriptional control.
This control occurs through transcriptional regula-
tors that bind specific DNA sequences, leading to the
modification of chromatin structure, which in turn
controls the accessibility of DNA to regulatory factors.
The main factor influencing chromatin structure is
the state of amino-acid residues within histone tails,
which serve as targets for a variety of reversible post-
translational modifications that modulate nucleosome
structure and gene transcription, both positively and
negatively1,2. Acetylation, one of the most widespread
modifications of histones, serves as a key modulator
of chromatin structure and gene transcription, and
provides a mechanism for coupling extracellular sig-
nals with the genome by regulated acetylation and
Histone acetylation modulates transcription in mul-
tiple ways. Acetylation of ?-amino groups of lysine resi-
dues within histone tails neutralizes their positive charge,
thereby relaxing chromatin structure. This interferes
with the generation of higher-order chromatin struc-
tures, and increasing the accessibility of transcription
factors to their target genes4. Acetylated histones also
serve as binding sites for bromodomain proteins, which
often act as transcriptional activators. Conversely, his-
tone deacetylation favours transcriptional repression by
allowing for chromatin compaction5. Direct acetylation
and deacetylation of transcription factors has also been
shown to have positive and negative consequences on
gene transcription, respectively6.
Histone acetylation is a dynamic process control-
led by the antagonistic actions of two large families of
enzymes — the histone acetyltransferases (HATs) and the
histone deacetylases (HDACs). The balance between
the actions of these enzymes serves as a key regulatory
mechanism for gene expression and governs numerous
developmental processes and disease states.
The vast majority of studies of HDAC functions have
involved biochemical analyses in vitro, studies in cul-
tured cells with HDAC inhibitors, HDAC knockdown
by small interfering RNA (siRNA), or overexpression
of HDACs. Although these experiments demonstrate
the biochemical functions of HDACs as transcriptional
repressors, they are non-predictive of their role in vivo.
An important question regarding the functions and
mechanisms of action of HDACs in vivo is whether
HDACs act primarily to control global changes in the
state of chromatin or whether they also have more spe-
cific functions in the regulation of key downstream
genes and transcriptional programmes. The existence
of many HDAC isoforms in eukaryotic cells also raises
questions about possible specificity or redundancy of
functions. Ongoing human clinical trials that are inves-
tigating the use of HDAC inhibitors as a treatment for a
variety of disorders mean that it is vital these questions
are answered. The recent creation of knockout mice lack-
ing HDAC genes has revealed highly specific functions
for individual HDAC isoforms during development
Department of Molecular
Biology, University of Texas
Southwestern Medical Center,
6000 Harry Hines Boulevard,
Dallas, Texas 75390-9148,
Correspondence to E.N.O.
9 December 2008
The many roles of histone deacetylases
in development and physiology:
implications for disease and therapy
Michael Haberland, Rusty L. Montgomery and Eric N. Olson
Abstract | Histone deacetylases (HDACs) are part of a vast family of enzymes that have
crucial roles in numerous biological processes, largely through their repressive influence
on transcription. The expression of many HDAC isoforms in eukaryotic cells raises
questions about their possible specificity or redundancy, and whether they control
global or specific programmes of gene expression. Recent analyses of HDAC knockout
mice have revealed highly specific functions of individual HDACs in development and
disease. Mutant mice lacking individual HDACs are a powerful tool for defining the
functions of HDACs in vivo and the molecular targets of HDAC inhibitors in disease.
32 | JANUARY 2009 | VOLUME 10
and adulthood. These mutant mice are a powerful tool
for defining the functions of HDACs in vivo and for
identifying the molecular targets of HDAC inhibitors
in disease. In this Review, we discuss the developmental
and physiological functions of HDACs that are revealed
by gene deletions in mice, and how these studies can
inform future efforts to exploit HDACs in the settings
of human disease.
Control of gene expression by HDACs
HDACs lack intrinsic DNA-binding activity and are
recruited to target genes via their direct association
with transcriptional activators and repressors, as well
as their incorporation into large multiprotein transcrip-
tional complexes2,4. Thus, the specificity of HDACs for
regulation of distinct gene programmes depends on cell
identity and the spectrum of available partner proteins
in a cell, in addition to the signalling milieu of the cell.
Although diminished histone acetylation at promoter
regions generally correlates with gene silencing, con-
sistent with the well-established functions of HDACs
as transcriptional repressors, there is also evidence that
HDACs can activate some genes. In yeast, for example,
the HDAC Hos2 is required for gene activation, and
deletion of the HDAC1 and 2 homologue, Rpd3, leads to
repression of transcription at telomeric loci7–9. HDACs
have also been linked to transcriptional activation of a
subset of genes in higher eukaryotes10 but, in settings
in which HDAC inhibition leads to downregulation of
specific genes, it is difficult to rule out possible second-
ary effects that result in transcriptional repression. It
should be noted, however, that deletion or inhibition
of HDACs often results in the upregulation or down-
regulation of approximately equivalent percentages of
It has also become clear in recent years that HDACs
can act on numerous cellular substrates in addition to
histones, and that acetylation might rival phosphoryla-
tion in its importance6. In this regard, class IIa deacety-
lases possess only minimal HDAC activity against
acetylated histones, despite extensive evolutionary con-
servation of their deacetylase domain, pointing to the
possible importance of other types of cellular substrates
for their actions13. How these many facets of acetylation
and deacetylation are controlled and integrated, and how
they influence the expression of specific genes as well as
global changes in gene expression, are important issues
for the field.
The HDAC superfamily
The HDAC superfamily is vast and ancient, dat-
ing back to prokaryotes. Here we focus on the mam-
malian HDACs and the lessons learned from genetic
Mammalian genomes encode 11 proteins with a
highly conserved deacetylase domain (FIG. 1). These
proteins can be classified into four families (class I, IIa,
IIb and IV), which differ in structure, enzymatic func-
tion, subcellular localization and expression patterns. In
addition to these classical HDACs, mammalian genomes
encode another group of deacetylases, the sirtuins, which
are sometimes referred to as class III HDACs. There have
been several recent reviews on the sirtuins14–17, so they
will not be covered here.
Nature Reviews | Genetics
hypertrophy after stress
MEF2 14-3-3 14-3-3
PhenotypeTime of lethality Protein domains
defect in growth plate
hypertrophy after stress
912SSS S E11HDAC7
M.H. and E.O.,
Figure 1 | The histone deacetylase (HDAC) superfamily, showing protein domains, loss-of-function phenotypes in
mice and time point of lethality of the knockouts. Green rectangles indicate the conserved HDAC domain; numbers
following the HDAC domain indicate the number of amino acids. Myocyte enhancer factor 2 (MEF2)-binding sites are
marked by a blue square, and binding sites for the 14-3-3 chaperone protein are also shown. E, embryonic day;
ND, not determined; P, days postnatal; S, serine phosphorylation sites; ZnF, zinc finger.
NATURE REVIEWS | GENETICS
VOLUME 10 | JANUARY 2009 | 33
A particular state
corresponding to the highest
energy in a chemical reaction.
Class I HDACs. The class I HDAC family consists of
HDAC1, 2, 3 and 8, which share homology with Rpd3 — a
founding member from budding yeast18–20. These HDACs
are expressed ubiquitously, localized predominantly to
the nucleus and display high enzymatic activity toward
histone substrates. They possess relatively simple struc-
tures, consisting of the conserved deacetylase domain
with short amino- and carboxy-terminal extensions.
HDAC1 and HDAC2 are nearly identical and are gen-
erally found together in repressive complexes such as the
Sin3, NuRD, CoREST and PRC2 complexes21. HDAC3 is
found in distinct complexes such as the N-CoR–SMRT
complex, whereas no complex has been described for
HDAC8 (REF. 19).
Class IIa HDACs. HDAC4, 5, 7 and 9 belong to the class
IIa HDAC family. These HDACs have large N-terminal
extensions with conserved binding sites for the tran-
scription factor myocyte enhancer factor 2 (MEF2) and
the chaperone protein 14-3-3, which render HDACs sig-
nal responsive. Following phosphorylation by kinases,
such as calcium/calmodulin-dependent protein kinase
(CaMK) and protein kinase D (PKD), these HDACs bind
14-3-3 and shuttle from the nucleus to the cytoplasm22–25.
The dissociation of class II HDACs from MEF2 allows
the HAT p300 to associate with MEF2 via the HDAC
docking site, thereby converting MEF2 from a transcrip-
tional repressor to a transcriptional activator26–30. The
regulated phosphorylation of class IIa HDACs provides
a mechanism for linking extracellular signals with tran-
scription and has key roles in numerous tissues during
development and disease.
In contrast to other HDACs, class IIa HDACs show
relatively restricted expression patterns. HDAC5 and
HDAC9 are highly enriched in muscles, the heart
and brain31,32. HDAC4 is highly expressed in the brain and
growth plates of the skeleton33, and HDAC7 is enriched
in endothelial cells and thymocytes34 (T-cell precursors
derived from the thymus).
The precise mechanism whereby class IIa HDACs
repress transcription has not been fully elucidated.
Highly purified recombinant class IIa HDACs possess
only minimal catalytic activity, and the activity of class
IIa HDACs purified from mammalian cells has been
shown to be due to contaminating class I HDACs13,35,36.
Moreover, MEF2-interacting transcription repressor
(MITR), which is a splice variant of HDAC9 that lacks
the HDAC domain, is as effective in repression of MEF2-
target genes as the full-length HDAC9 protein, indicat-
ing that the intrinsic catalytic activity of class IIa HDACs
is not required for repression37–39.
The class IIa HDACs have been shown to recruit
class I HDACs through their C-terminal HDAC
domain, which probably accounts for a portion of their
repressive activity35. In addition, the regulatory domains
of class IIa HDACs interact with other transcrip-
tional repressors, such as heterochromatin protein 1
(HP1) and C-terminal-binding protein (CTBP)37,38,40.
Thus, they function as adaptors to nucleate multiple
types of transcriptional regulators and to confer signal-
responsiveness to downstream target genes.
Recently, the biochemical basis for the different
activities of class I and class IIa HDACs has been eluci-
dated. In the catalytic pocket of most HDACs, an ultra-
conserved tyrosine acts as a transition-state stabilizer in
the deacetylation reaction41. This tyrosine is changed to
a histidine in vertebrate class IIa HDACs, and this con-
servative amino-acid change reduces the catalytic activ-
ity of the vertebrate enzymes more than 1,000-fold36,42.
Although this activity is still measurable in vitro, it is
unclear if it is of any biological relevance in vivo, espe-
cially given the observation that class IIa HDACs do
not need their catalytic domain in order to be potent
Class IIb HDACs. HDAC6 and HDAC10 form the class
IIb family. HDAC6 is the main cytoplasmic deacetylase
in mammalian cells43, whereas little is known about
the functions of HDAC10 (REFS 44,45). Among the tar-
gets directly deacetylated by HDAC6 are cytoskeletal
proteins such as ?-tubulin and cortactin, transmem-
brane proteins such as the interferon receptor IFN?R,
and chaperones46–50. HDAC6 is distinct from all other
HDACs, as it harbours two deacetylase domains and a
C-terminal zinc finger.
Class IV HDAC. HDAC11 is the sole class IV HDAC.
Expression of HDAC11 is enriched in the brain, heart,
muscle, kidney and testis, but little is known about its
function51,52. It is composed of a deacetylase domain that
shows homology to class I and II HDAC domains, with
small N- and C-terminal extensions.
Roles of class I HDACs in development
The ubiquitous expression, high deacetylase activ-
ity towards common substrates and high homology
between class I HDACs suggests functional redundancy
among these HDACs in vivo. However, deletion of each
member of the class I HDAC family in mice leads to
lethality in all cases, demonstrating the unique roles of
each HDAC in the control of specific gene expression
HDAC1. HDAC1-null mice die before embryonic day
10.5 (E10.5) and display severe proliferation defects and
general growth retardation12,53. Proliferation defects can
also be observed in HDAC1-null embryonic stem (ES)
cells and are associated with increased expression of the
cyclin-dependent kinase inhibitors p21 and p27 (REF. 54).
These cells show a significant reduction in total HDAC
activity and modest hyperacetylation of histones H3 and
H4, indicating that HDAC1 is a major deacetylase in ES
cells. Surprisingly, 3% of genes are downregulated and
~5% of genes are upregulated in HDAC1-null ES cells54,
suggesting that HDAC1 does not function simply as a
global repressor of transcription, but instead regulates
specific gene programmes by repressing or activating
Deletion of hdac1 in zebrafish causes a variety of
lethal defects in skeletal and neuronal elements. The spe-
cific target genes responsible for these phenotypes have
not been defined, but they seem to be downstream of
34 | JANUARY 2009 | VOLUME 10
A stable antisense
oligonucleotide that is
commonly used in zebrafish
and Xenopus laevis to inhibit
either the translation or
splicing of mRNAs.
A mutation strategy that uses
insertion vectors to trap or
isolate transcripts from flanking
The process in animal embryos
in which the endoderm and
mesoderm move from the
outer surface of the embryo to
the inside, where they give rise
to the internal organs.
canonical and non-canonical Wnt signalling55–59. None
of the other HDACs has been genetically analysed in
detail in zebrafish, although a recent report described
disrupted liver development following morpholino-
mediated knockdown of hdac3 mRNA (REF. 60).
Surprisingly, conditional deletion of HDAC1 in tissues
such as the heart, brain, skeletal muscle and smooth mus-
cle is well tolerated in mice12, although this is probably
due to redundancy with HDAC2 in later development
and postnatal life (see below)12.
HDAC2. There is disagreement regarding the function of
HDAC2 in vivo. One study found that HDAC2-null mice
die within the first 24 hours after birth with severe car-
diac malformations, including obliteration of the lumen
of the right ventricle owing to excessive proliferation of
cardiomyocytes, as well as bradycardia12 (FIG. 2). By con-
trast, other studies have reported that mice harbouring a
lacZ insertion in Hdac2, which is purported to create
a null mutation, are viable61. The basis for these conflict-
ing results is unclear. It is possible that different genetic
backgrounds account for this difference. Alternatively, the
lacZ insertion allele might be ‘leaky’ and allow adequate
expression of HDAC2 for viability, a phenomenon that
has previously been described for gene trap approaches62.
The transcriptional targets of HDAC2 in the heart
remain to be fully defined. However, the homeodomain-
only protein (HOP), which functions as a positive and
negative regulator of cardiomyocyte proliferation, has
been shown to interact with HDAC2 (FIG. 2). Deletion
of HOP also results in hyperproliferation of develop-
ing cardiomyocytes, suggesting that HDAC2 and HOP
reside in a transcriptionally repressive complex to reg-
ulate cardiac proliferation and differentiation during
Redundant roles of HDAC1 and 2 in cardiac growth
and development. Conditional null alleles for class I
HDACs have permitted an analysis of their functions in
specific tissues, bypassing the early lethality associated
with global gene deletion. Given the lethal phenotypes
resulting from global deletion of HDAC1 and HDAC2,
it was surprising to find that deletion of either HDAC1
or HDAC2 in a variety of tissues, including the heart,
brain, endothelial cells, smooth muscle, and neural crest
cells did not yield obvious phenotypes. By contrast,
deletion of both genes together results in severe phe-
notypes in all tissues examined, pointing to redundant
functions of these HDACs during later development
Conditional deletion of HDAC1 and 2 together in
the cardiac lineage has shown that a single wild-type
allele of either gene is sufficient to support normal
development, whereas deletion of all HDAC1 and 2
alleles results in neonatal lethality, accompanied by
cardiac arrhythmias, dilated cardiomyopathy, and
upregulation of genes encoding skeletal muscle-specific
contractile proteins and calcium channels in the
heart12 (FIG. 2). The earlier that HDAC1 and HDAC2
are deleted, the more dramatic the phenotype. In the
heart, deletion of HDAC1 and HDAC2 at E8.5 causes
lethality 2 days later, whereas mice with a cardiac
deletion at E10.5 survived for more than 3 weeks.
Transcriptional analysis in these animals revealed that
only 1.6% of transcripts were upregulated, and
that deletion of HDAC1 and HDAC2 in the heart dere-
pressed specific gene programmes involved in Ca2+ ion
handling and in contractility. Cardiac expression of
multiple fetal calcium channels is transcriptionally
regulated by neuron-restrictive silencer factor (NRSF)
through the recruitment of both class I and class IIa
HDACs66,67. A dominant negative mutant of NRSF that
is unable to bind repressors results in activation of the
fetal gene programme, arrhythmogenesis and sudden
death68. Thus, loss of HDAC1 and HDAC2 allows for
the loss of repression by NRSF and other transcrip-
tion factors, resulting in aberrant transcriptional
activity of genes involved in calcium flux and con-
tractility, leading to cardiac arrhythmia and sudden
HDAC3. HDAC3 mutant mice die before E9.5 owing
to defects in gastrulation69–71. The target genes respon-
sible for this early phenotype are unknown, although
loss of Hdac3 seems to be associated with defective
DNA double-stranded break repair70. Conditional
Nature Reviews | Genetics
Nature Reviews | Genetics
a Wild type
c Wild type
HDAC2 KO (P1)
HDAC1;2 KO (P11)
Transcription of calcium
channels and skeletal
Figure 2 | Control of heart development by histone deacetylase 1 (HDAC1) and
HDAC2. a | Histological sections of hearts from wild type and HDAC2 knockout (KO)
mice at postnatal day 1 (P1). Note the excessive number of cardiomyocytes in the mutant
heart, which fill the chambers of the left ventricle (lv) and right ventricle (rv). b |
Schematic of the role of HDAC2 in the repression of cardiomyocyte proliferation
through inhibition of homeodomain-only protein (HOP). c | Histological sections of
hearts from wild-type mice and mice with a cardiac deletion of HDAC1 and 2 at P11.
Note the dilatation of the right ventricle in the mutant, which is indicative of heart
failure. d | Schematic of the redundant roles of HDAC1 and 2 in regulation of calcium
channel and skeletal muscle genes in cardiomyocytes via repression of neuron-restrictive
silencer factor (NRSF) and other transcription factors. Parts a and c are reproduced, with
permission, from REF. 12 ? (2007) Cold Spring Harbor Laboratory Press.
NATURE REVIEWS | GENETICS
VOLUME 10 | JANUARY 2009 | 35
Proliferation of fibroblasts
resulting in increased collagen
production and consequent
The type of cell that produces
and maintains the cartilaginous
The type of cell that is
responsible for bone formation
by secreting and mineralizing
the bone matrix.
The process of bone formation
in which soft connective tissue
is converted into mineralized
The parts of the skeleton which
form by endochondral
ossification, a process in which
cartilage is replaced by bone.
deletions of HDAC3 have so far been described for
the liver and heart. Loss of HDAC3 in the liver dis-
rupts lipid and cholesterol homeostasis, leading to an
accumulation of lipids and a decrease in glycogen stor-
age71. These changes are caused by derepression of a
gene programme that usually is under the control of
nuclear hormone receptors such as the thyroid hor-
mone receptor and peroxisome proliferator-activated
receptor gamma (PPAR?), which control key steps in
lipid and cholesterol biosynthesis early in the postnatal
liver. Only minor increases could be observed in bulk
histone acetylation and on the promoters of dysregu-
lated genes, indicating that other class I HDACs are
also likely to play a part in liver homeostasis.
Deletion of HDAC3 in cardiomyocytes also led to a
dramatic upregulation of ligand-induced lipid storage
in the heart69. These mice survive until 3–4 months of
age, at which point they show massive cardiac hyper-
trophy and derepression of genes that control fatty-acid
uptake and metabolism. In the heart, these gene pro-
grammes are under the control of the nuclear receptor
PPAR?, and derepression by loss of HDAC3 leads to
abnormalities that mimic the metabolic derangements
observed in diabetic cardiomyopathies. Furthermore,
loss of HDAC3 in the heart results in robust interstitial
fibrosis, which is phenotypically independent of ram-
pant PPAR? activity. However, it is currently unknown
whether the transcription factors that regulate the
fibrotic gene programme are directly repressed by
HDAC3. Overexpresion of HDAC3 in the heart leads
to increased thickness of the myocardium, which is
due to increased cardiomyocyte hyperplasia without
Class IIa HDACs in development and physiology
Each of the four class IIa HDACs have been deleted in
mice and, although each gene seems to be dedicated to
specific programmes of tissue-specific gene expression,
commonalities between the different loss-of-function
phenotypes point to similar mechanisms of action.
Many of these modes of action reflect the repressive
influence of these HDACs on the expression and func-
tion of the MEF2 transcription factor, as well as their
signal responsiveness. Importantly, there is a high
degree of redundancy between the class IIa HDACs. It
is thus possible that each tissue has a hard-wired thresh-
old for class IIa HDAC repression, and that the observed
phenotypes reflect the cell types or gene programmes
that are most sensitive to MEF2 and HDAC activity.
Regulation of skeletogenesis by HDAC4. HDAC4 has
a central role in the formation of the skeleton33. Most
of the bones in the vertebrate skeleton are formed
from a cartilaginous template in which chondrocytes
undergo hypertrophy, which is followed by apoptosis.
Thereafter, osteoblasts, blood vessels and other cell
types invade and produce the mature bone matrix73.
HDAC4 is expressed in prehypertrophic chondrocytes
in vivo, and mice with a global deletion of HDAC4 die
during the first week of life owing to ectopic ossification
of endochondral cartilage, which prevents expansion of
the rib cage and leads to an inability to breathe (FIG. 3).
This lethal phenotype is accompanied by precocious
and ectopic hypertrophy of chondrocytes, resulting
in the conversion of cartilaginous skeletal elements
to ossified bone. Runt related transcription factor 2
(RUNX2) and the MEF2C transcription factor, which
interact with HDAC4, have vital roles in the control
of chondrocyte hypertrophy and bone formation74.
In the absence of HDAC4, transcriptional activation
of these factors is unrestrained, leading to excessive
bone formation33. Consistent with this mechanism,
forced expression of RUNX2 or a constitutively active
form of MEF2 in developing chondrocytes mimics
the HDAC4 loss-of-function phenotype75. Conversely,
forced expression of a signal-resistant mutant form of
HDAC4 in chondrocytes in vivo inhibits chondrocyte
hypertrophy and differentiation (FIG. 3).
Thus, by repressing the activity of MEF2C and
RUNX2 in developing chondrocytes, HDAC4 is able
to delay chondrocyte hypertrophy and thereby control
the timing and extent of ossification of endochondral
bones (FIG. 3). MEF2 directly regulates the expression
of extracellular matrix protein genes, such as collagen
type X alpha 1, and of vascular endothelial growth fac-
tor (VEGF), which is required for angiogenesis in the
Nature Reviews | Genetics
Normal bone and cartilage
Absence of bone
Figure 3 | Control of chondrocyte hypertrophy by histone deacetylase 4 (HDAC4). a | Ribs from neonatal mice
stained for bone (red) and cartilage (blue). Deletion of HDAC4 results in ossification of cartilage (designated
by the arrowhead), whereas overexpression of HDAC4 in the cartilage of transgenic mice prevents ossification.
b | Schematic of the repressive influence of HDAC4 on myocyte enhancer factor 2 (MEF2) and runt related
transcription factor 2 (RUNX2) in the pathway for chondrocyte proliferation and hypertrophy. IHH, Indian
hedgehog; KO, knockout; PTHrP, parathyroid hormone-related peptide. Part a is reproduced, with permission, from
REF. 33 ? (2007) Cell Press.
36 | JANUARY 2009 | VOLUME 10
late stages of chondrocyte development75. In addition,
RUNX2 activates the expression of the secreted growth
factor Indian hedgehog (IHH), which has a number
of functions in endochondral bone development. These
functions are mediated by enhancing chondrocyte pro-
liferation and stimulating the synthesis of parathyroid
hormone-related peptide (PTHrP), which in turn
inhibits differentiation of prehypertrophic to hyper-
trophic chondrocytes76. RUNX2 expression is also con-
trolled by MEF2, and RUNX2 is a target for regulation
by HDAC4 (REF. 75).
Control of cardiovascular growth and function by
HDAC5 and 9. Mice lacking either HDAC5 or HDAC9
are viable, whereas compound mutant mice lacking
both HDAC5 and 9 show a propensity for lethal ven-
tricular septal defects and thin-walled myocardium,
which typically arise from abnormalities in growth
and maturation of cardiomyocytes32. Given the inter-
action between class IIa HDACs and MEF2, and the
central role of MEF2 in the control of cardiomyocyte
differentiation, the developmental cardiac defects in
these double mutant mice probably result from super-
activation of MEF2. This would be expected to lead to
precocious differentiation and cell-cycle withdrawal of
cardiomyocytes, causing hypocellularity of the myo-
cardium. In addition, class IIa HDACs participate in
multiprotein complexes and modulate the activities
of numerous other transcription factors involved in
myocardial growth, such as the serum response fac-
tor, myocardin and calmodulin binding transcription
activator 2 (CAMTA2)77. Thus, the absence of HDAC5
and 9 probably perturbs the precisely coordinated gene
expression programmes required for myocyte differen-
tiation, proliferation and morphogenesis that underlie
The adult heart typically responds to stress by a
pathological growth response that ultimately leads to
loss of cardiac function78–80. MEF2 is sufficient and nec-
essary to drive the pathological cardiac hypertrophy
and heart failure that takes place in response to injury81.
HDAC5 and 9 have redundant roles in the suppres-
sion of cardiac growth in response to stress signalling.
Mice lacking either HDAC5 or 9 are hypersensitive to
cardiac stress resulting from excess workload or neuro-
humoral signalling (FIG. 4). These stimuli typically acti-
vate the calcineurin and CaMK–PKD pathways, which
in turn lead to phosphorylation of class IIa HDACs,
promoting their nuclear export82. Deletion of class IIa
HDACs eliminates the counter-regulatory mechanism
that restrains cardiac growth and sensitizes MEF2 and
perhaps other transcription factors so that they become
activated by stress-dependent intracellular signals.
Functions of class IIa HDACs in skeletal muscle.
Numerous functions for class IIa HDACs have been
described in skeletal muscle. Skeletal muscle fibres dif-
fer in their contractile and metabolic properties, which
reflect different patterns of gene expression83. Slow-
twitch, or type I, myofibres exhibit an oxidative metabo-
lism, are rich in mitochondria, are heavily vascularized
and are resistant to fatigue. By contrast, fast-twitch, or
type II, myofibres exhibit glycolytic metabolism, are
involved in rapid bursts of contraction and fatigue
rapidly. The calcium-dependent protein kinases CaMK
and PKD have been implicated in the transduction of
calcium signals that upregulate the expression of oxi-
dative, slow fibre-specific genes in skeletal muscle84.
MEF2 is a target for calcium signalling in skeletal mus-
cle and is a key regulator of the slow myofibre pheno-
type. This function of MEF2 is mediated through its
regulation by class IIa HDACs: in slow myofibres, class
Nature Reviews | Genetics
HDAC4, 5, 9
HDAC4, 5, 9
Transcription of genes involved in hypertrophy,
fibrosis and pathological remodelling
CaMK, PKDNuclear export
HDAC9 KOWild type
Figure 4 | Control of pathological cardiac hypertrophy by class IIa histone deacetylases (HDACs). a | Histological
sections of hearts from wild-type and HDAC9 knockout (KO) adult mice. Mice were subjected to cardiac stress by
expression of a cardiac-specific transgene encoding activated calcineurin, which drives pathological hypertrophy.
Note that HDAC9 knockout mice have normal hearts in the absence of stress, but display cardiomegaly in response to
stress, owing to loss of the growth-inhibitory function of HDAC9. b | Schematic of the repressive influence of class IIa
HDACs on myocyte enhancer factor 2 (MEF2) and pathological cardiac remodelling. Stress-inducible kinases, such as
calcium/calmodulin-dependent protein kinase (CaMK) and protein kinase D (PKD), induce the phosphorylation of
class IIa HDACs, which creates docking sites for the 14-3-3 chaperone protein, resulting in nuclear export with
consequent activation of MEF2 and its downstream target genes, which are involved in cardiac remodelling. Part a is
reproduced, with permission, from REF. 31 ? (2007) Cell Press.
NATURE REVIEWS | GENETICS
VOLUME 10 | JANUARY 2009 | 37
A module for fine tuning a
IIa HDACs are selectively degraded by the proteasome
and MEF2 exerts a transcriptional activation function.
Among the target genes of MEF2 in slow myofibres
are type IIx myosin heavy chain, myosin light chain 2,
slow troponin I and myoglobin. In support of this
mode of function of class IIa HDACs in skeletal muscle,
genetic deletion of class IIa HDACs in this tissue dere-
presses MEF2 and results in conversion of fast fibres to
slow fibres85 (FIG. 5).
HDAC9 has also been shown to modulate the
response of skeletal muscle to motor innervation.
Electrical activity from motor neurons represses the
expression of many muscle genes, including those
encoding acetylcholine receptor subunits. In response
to denervation, these genes are derepressed, resulting
in hypersensitivity of the muscle fibre to acetylcholine.
Mice lacking HDAC9 are extremely sensitive to den-
ervation-induced changes in gene expression, whereas
mice that overexpress HDAC9 in skeletal muscle are
rendered insensitive to the effects of denervation86.
In addition to being a signal-responsive modulator of
gene transcription, the expression of HDAC9 is tightly
modulated. A highly conserved MEF2-binding site in
the proximal promoter of the HDAC9 gene drives the
expression of HDAC9, thereby establishing a negative
feedback loop in which MEF2 drives the expression of
its own repressor87 (FIG. 5). This feedback loop is thought
to provide robustness and fine-tuning to the gene pro-
grammes controlled by HDAC9 and MEF2, and to
provide a myogenic ‘rheostat’ that modulates muscle
differentiation in response to extracellular cues.
Control of endothelial function by HDAC7. During
embryogenesis, HDAC7 is specifically expressed in
the endothelial cells that form the inner lining of the
cardiovascular system34. Genetic deletion of HDAC7
in mice results in embryonic lethality, owing to a loss
of integrity of endothelial-cell interactions and con-
sequent rupture of blood vessels and haemorrhaging
(FIG. 6). Vascular disruption in HDAC7-null mice is
accompanied by upregulation of matrix metallopro-
teinase 10 (MMP10), an endoprotease that is secreted
by endothelial cells and that degrades the extracellular
matrix, thereby perturbing endothelial-cell and smooth
muscle-cell interactions. The inappropriate expression
of MMP10 can be traced to its regulation by MEF2;
in the absence of HDAC7, MEF2 activity is elevated,
leading to pathological levels of MMP10 (FIG. 6).
Concurrently, tissue inhibitor of metalloproteinase 1
(TIMP1) is downregulated in endothelial cells, presum-
ably as a secondary consequence of vascular demise.
The downregulation of TIMP1 in the face of enhanced
expression of MMP10 would be expected to further
exacerbate vascular destruction34.
The involvement of HDAC7 in the control of
MMP10 expression and vascular integrity has poten-
tially important implications for a variety of human dis-
orders. Vascular leakage causes circulatory collapse and
contributes to the pathogenesis of numerous usually
life-threatening diseases, such as atherosclerosis and
aneurysm. Moreover, the imbalance between MMP
and TIMP activity has been shown to profoundly influ-
ence vascular integrity following myocardial infarction
Nature Reviews | Genetics
Transcription of muscle
Transcription of slow
HDAC4, 5, 9
CaMK, PKDNuclear export
HDAC4, 5, 9
a Wild typeHDAC5;9 KO
Figure 5 | Control of slow myofibre gene expression by class IIa histone deacetylases (HDACs). a | Histological
sections of soleus muscle from wild-type and HDAC5;9 double mutant knockout (KO) mice stained for type I myosin
heavy chain, a marker of type I slow myofibres. Note the increase in slow myofibres after deletion of class IIa HDACs.
b | Schematic of the repressive influence of class IIa HDACs on myocyte enhancer factor 2 (MEF2), which acts together
with PGC-1? (peroxisome proliferator-activated receptor gamma, coactivator 1 alpha) and NFAT (nuclear factor of
activated T-cells) to promote the formation of slow myofibres. Signalling by calcium/calmodulin-dependent protein
kinase (CaMK) and protein kinase D (PKD) induces the phosphorylation of class IIa HDACs, which creates docking sites
for the 14-3-3 chaperone protein, resulting in nuclear export with consequent activation of slow myofibre genes.
c | A MEF2-dependent negative feedback loop for the control of HDAC9 expression during muscle differentiation.
Myogenic basic helix-loop-helix (bHLH) transcription factors activate the expression of MEF2, which then amplifies and
sustains the expression of myogenic bHLH genes. Myogenic bHLH factors and MEF2 also cooperate to activate skeletal
muscle differentiation genes. In addition, MEF2 activates the expression of HDAC9, which in turn represses MEF2
activity. Signals that influence myogenesis activate HDAC kinases and thereby repress HDAC9 activity, providing a
‘rheostat’ mechanism for the control of myogenesis. Part a is reproduced from REF. 85.
38 | JANUARY 2009 | VOLUME 10
Inhibition concentration 50%.
The concentration of inhibitor
that is required to inhibit 50%
of the activity of an enzyme
compared with an uninhibited
and during tumour angiogenesis88,89. Hence, strategies
to maintain the repressive influence of HDAC7 on
MEF2 — for example, through inhibition of the kinase
cascades that lead to the phosphorylation of HDAC7
and its dissociation from MEF2 — would be expected
to be beneficial with respect to maintaining vascular
Control of cytoskeletal dynamics by HDAC6. Mice with
a deletion of HDAC6 are the only HDAC mutant ani-
mals published so far that do not have an obvious phe-
notype. However, they do display a dramatic increase in
acetylated tubulin, in line with the notion that HDAC6
is the main tubulin deacetylase43. Given the large body
of experimental in vitro evidence that clearly shows that
HDAC6 has important functions in modulating the
misfolded protein response and cytoskeletal dynamics,
the lack of phenotype in the HDAC6 mutants is some-
what surprising. It is possible that redundancy with
HDAC10 can explain the lack of in vivo phenotype.
Therapeutic actions of HDAC inhibitors
The involvement of histone acetylation and deacetyla-
tion in so many aspects of development and tissue
homeostasis might suggest that systemic inhibition of
HDACs with pharmacologic inhibitors would result in
nonspecific and catastrophic effects as a consequence
of global derepression of gene expression. Thus, it is
striking that systemic HDAC inhibition with com-
pounds that broadly inhibit most or all HDACs is
well tolerated in vivo and blocks numerous disease-
associated gene expression programmes in a seemingly
Given the dramatic phenotypes that result from
HDAC gene deletions, why are HDAC inhibitors so
well tolerated in vivo? We propose three explanations,
which are not mutually exclusive. First, a genetic dele-
tion of an HDAC results in the complete absence of
the enzyme, whereas inhibitors do not result in com-
plete inhibition of activity. Second, a genetic deletion
of an HDAC eliminates the gene product permanently,
whereas the actions of an inhibitor are transient. Third,
and perhaps most importantly, HDACs participate in
multiprotein transcriptional complexes. Genetic dele-
tion of an HDAC perturbs the complexes in which it
would normally be associated, whereas inhibitors are
believed to block enzymatic activity without necessarily
disrupting the repressive complex.
Classical HDAC inhibitors such as trichostatin A
(TSA) or suberoylanilide hydroxamic acid (SAHA) are
mostly ‘pan–HDAC’ inhibitors; that is, they block, with
similar affinities, the activity of all isoforms except class
IIa HDACs. For example, the IC50 values for SAHA are:
HDAC1 = 37.1 nM; HDAC3 = 44.6 nM; and HDAC6 =
40.9 nM90. Given that different HDAC isoforms govern
dramatically different gene expression programmes in
development and disease, it seems plausible that iso-
form-selective inhibitors should lead to improved effi-
cacy and drug safety. The recent mechanistic insights
into the biochemistry of class IIa HDACs should also
be taken into account36,41,91. Many screening studies
used class IIa HDACs purified from mammalian cells
for the development of class IIa isoform-specific inhibi-
tors and, surprisingly, compounds were identified that
blocked class IIa but not class I activity. These com-
pounds probably function as small-molecule inhibi-
tors of protein–protein interactions and not as bona
fide HDAC inhibitors92. As these molecules are enter-
ing clinical trials, it is important to realize that they
might show biological properties distinct from classical
HDAC inhibitors from multiple chemical classes
have entered clinical trials, and SAHA (marketed as
Vorinostat, brand name Zolinza) has been approved
for treatment of cutaneous manifestations of advanced,
refractory T-cell lymphoma in a select group of
patients93. The exact mechanism for the effect of HDAC
inhibitors on tumour cells is currently unknown, and
numerous explanations, such as changes in gene tran-
scription, direct induction of apoptosis, production
of reactive oxygen species and induction of cell-cycle
arrest, have been proposed92,94–96. The specific HDAC
isoforms that mediate this antiproliferative effect also
remain to be clearly identified. Genetic deletion of
HDAC3 leads to cell-cycle dependent DNA damage
coupled with defective double-stranded break repair70.
HDAC3-null cells are thus sensitized to ionizing radia-
tion, a phenomenon that has also been observed with
HDAC inhibitors97. Therefore, some of the effects
observed with HDAC inhibition might be mediated via
HDAC3, although the involvement of other isoforms
can not be ruled out98. The existence of conditional
alleles for all class I HDACs might allow the creation
of transformed cancer cell lines with conditional alle-
les for all the different class I HDAC isoforms (and
their combinations), which would make a systematic
analysis of HDAC requirement in cancer cells possible.
a Wild type
Transcription of genes
involved in endothelial
Figure 6 | Control of endothelial integrity by histone deacetylase 7 (HDAC7).
a | Wild-type and HDAC7 knockout (KO) embryos at embryonic day 10.5 (E10.5). The
absence of HDAC7 results in vascular rupture, pericardial oedema and haemorrhaging
throughout the mutant embryos. b | Schematic of the role of HDAC7 in maintenance of
vascular integrity. HDAC7 is expressed specifically in endothelial cells, where it represses
the activity of myocyte enhancer factor 2 (MEF2). In the absence of HDAC7, MEF2
activity is elevated, resulting in upregulation of matrix metalloproteinase 10 (MMP10)
and degradation of cell–cell interactions required for vascular integrity. Deletion of
HDAC7 also leads to downregulation of tissue inhibitor of metalloproteinase 1 (TIMP1),
presumably through indirect mechanisms, which further enhances MMP10 activity.
Part a is reproduced, with permission, from REF. 34 ? (2007) Cell Press.
NATURE REVIEWS | GENETICS
VOLUME 10 | JANUARY 2009 | 39
These studies could then be extended by crossing con-
ditional HDAC-null alleles into tumour-prone genetic
One of the most perplexing aspects of HDAC biol-
ogy is that pharmacological inhibition of HDAC activ-
ity provides a therapeutic benefit in such a wide variety
of disease states. TABLE 1 gives an overview of the dis-
ease states in which HDAC inhibition has been shown
to be beneficial as well as the proposed mechanisms
involved. These range from infectious and immuno-
logical diseases to traumatic shock, and from cardiac
hypertrophy to neurodegenerative disease99–105, and
in certain circumstances HDAC inhibitors are even
able to ‘cure’ genetic disease in humans106. Although
a unifying theory explaining how reduced deacetylase
activity is beneficial in such diverse pathophysiological
states is currently unknown, it is tempting to speculate
that most of these diseases have an epigenetic compo-
nent (that is, aberrant histone acetylation), and that
treatment with HDAC inhibitors resets the epigenetic
memory of the cell to a pre-disease state.
Issues for the future
Although the interest in HDAC biology has intensified
with the successful introduction of HDAC inhibitors
in the clinical setting, we are still far from understand-
ing the intricacies of protein acetylation. The number
of identified acetylated non-histone proteins is rapidly
increasing, raising questions regarding whether phe-
notypes resulting from HDAC gene deletion or from
pharmacological inhibition reflect changes in chroma-
tin structure and transcription, or if they reflect hyper-
acetylation of non-histone proteins. In the cases of
non-histone substrates, it will be important to identify
the proteins and understand how acetylation influences
their actions. It also remains to be determined whether
different HDAC isoforms have specific targets in vivo,
as suggested by the specific phenotypes resulting from
genetic deletion of individual HDACs. It is expected
that unbiased profiling of hyperacetylated proteins in
knockout or inhibitor-treated cells using mass spec-
trometry approaches will be a valuable tool in answer-
ing these questions. A recent study in human cancer
cells showed the feasibility of this approach107.
Given the variety of pre-clinical studies in which
HDAC inhibitors have shown a therapeutic benefit,
another major challenge will be to decipher the role of
individual HDACs in specific disease processes and to
develop isoform-specific HDAC inhibitors. Because
HDACs can affect multiple targets, it will be important
to develop inhibitors that selectively block the patho-
logical actions of HDACs. Solving these challenges
will most likely broaden the therapeutic window and
possibly lead to the clinical application of HDAC
inhibitors in a variety of non-oncological disease
Table 1 | Clinical and experimental use of histone deacetlyase (HDAC) inhibitors in diverse disease states
Proposed mechanism Refs
De-silencing of latent virus 99
Cutaneous T-cell lymphoma Upregulation of tumor-suppressor genes, induction of apoptosis 108, 109
GPI deficiency Increased PIGM expression by hyper-acetylation of histones on promoter 106
Inhibition of NF-?B activation
Sickle cell disease
Increase in fetal haemoglobin expression 111
Inhibition of TNF? expression and of inflammation
Asthma Inhibition of cytokine expression and T-cell infiltration 113
Autoimmune encephalitis Upregulation of antioxidant, antiexcitotoxicity and proneuronal factors 114
Colitis Suppression of pro-inflammatory cytokines 115
Cardiac hypertrophy Unknown 102, 116
Dementia Dendrite sprouting, increased synapse number 100
Graft versus host disease Reduction of pro-inflammatory cytokines 101, 117
Inhibition of TNF? and INF?
Muscular dystrophy Induction of follistatin119
Systemic lupus erythematosus Downregulation of pro-inflammatory cytokines 120
Spinal muscular atrophy
Activation of survival motor neuron 2 gene 121
Reduction of TNF? expression
GPI, glycosylphosphatidylinositol; INF?, interferon gamma, NF-?B, nuclear factor kappa B; PIGM, phosphatidylinositol glycan
anchor biosynthesis, class M; TNF?, tumour-necrosis factor alpha.
Inhibiting neuroinflammation 104
40 | JANUARY 2009 | VOLUME 10
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We apologize to the many authors in the field whose work we
were not able to cite because of space constraints. Research
in the Olson laboratory has been supported by grants from
the National Institutes of Health, the D.W. Reynolds Clinical
Cardiovascular Research Center, the Robert A. Welch
Foundation and the Sandler Foundation for Asthma Research.
M.H was supported by a grant from the Deutsche
Forschungsgemeinschaft (DFG, HA 3335/2-1).
Competing interests statement
The authors declare competing financial interests; see web
version for details.
HDAC1 | HDAC2 | HDAC3 | HDAC4 | HDAC5 | HDAC6 |
HDAC7 | HDAC8 | HDAC9 | HDAC10 | HDAC11 | MEF2
Olson laboratory homepage:
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42 | JANUARY 2009 | VOLUME 10