Int. J. Mol. Sci. 2011, 12, 6544-6565; doi:10.3390/ijms12106544
International Journal of
ATP-Dependent Chromatin Remodeling Factors and Their
Roles in Affecting Nucleosome Fiber Composition
Paolo Piatti, Anette Zeilner and Alexandra Lusser*
Division of Molecular Biology, Innsbruck Medical University, Biocenter, Fritz-Pregl Strasse 3,
6020 Innsbruck, Austria; E-Mails: firstname.lastname@example.org (P.P.); email@example.com (A.Z.)
* Author to whom correspondence should be addressed; E-Mail: firstname.lastname@example.org;
Tel.: +43-512-9003-70210; Fax: +43-512-9003-73100.
Received: 22 July 2011; in revised form: 20 September 2011 / Accepted: 28 September 2011 /
Published: 6 October 2011
Abstract: ATP-dependent chromatin remodeling factors of the SNF2 family are key
components of the cellular machineries that shape and regulate chromatin structure and
function. Members of this group of proteins have broad and heterogeneous functions
ranging from controlling gene activity, facilitating DNA damage repair, promoting
homologous recombination to maintaining genomic stability. Several chromatin
remodeling factors are critical components of nucleosome assembly processes, and recent
reports have identified specific functions of distinct chromatin remodeling factors in the
assembly of variant histones into chromatin. In this review we will discuss the specific
roles of ATP-dependent chromatin remodeling factors in determining nucleosome
composition and, thus, chromatin fiber properties.
Keywords: chromatin; histone variant; chromatin remodeling factor; centromere; linker
histone; chromatin assembly
Chromatin is an extremely complex structure that serves to compact eukaryotic DNA in order to
comply with the size restrictions of the nucleus. In addition, the way in which chromatin is organized
and in which its arrangement is modulated endows it with an extraordinary regulatory potential. At its
most basic level of organization, chromatin consists of repeating spherical particles termed
Int. J. Mol. Sci. 2011, 12
nucleosomes. Nucleosomes are formed by the wrapping of 147 bp of DNA in 1.7 left-handed
superhelical turns around a core of small, evolutionary conserved, highly basic histone proteins .
Two molecules each of the histones H3 and H4 interact via the so-called “histone-fold” domains to
generate a protein tetramer, which associates with two heterodimers of the histones H2A and H2B to
form the nucleosome core . Nucleosomes are connected by short stretches of linker DNA resulting
in a fiber with a diameter of ~10 nm that has a beads-on-a-string-like appearance [2,3]. Although this
structure may seem uniform from a superficial perspective, a tremendous amount of research during
the past decades has provided ample evidence that nucleosomes can differ from each other with
respect to their structure, the type of histones that they contain as well as the nature and extent of
chemical modifications on both the DNA and histones. In addition, the positioning of the nucleosomes
along the DNA can show striking variation, including regular arrangements with constant spacing
(e.g., in constitutive heterochromatin), irregular arrays of nucleosomes (typically in active genes) or
regions that are devoid or depleted of nucleosomes (e.g., at enhancers and promoters) [4,5].
Importantly, chromatin structure is not static. On the contrary, the organization and composition of
chromatin is constantly changing thereby facilitating or preventing access for DNA-utilizing proteins
to their substrate. In this review we will discuss some of the mechanisms that contribute to the shaping
of chromatin structure not only at the level of the 10 nm fiber but also in higher-order levels of
chromatin organization. We will give special attention to the ATP-dependent chromatin remodeling
machines and their diverse roles in modulating the composition of nucleosomes and chromatin fibers.
2. Chromatin Remodeling Machines and Their Impact on Nucleosome Structure
Chromatin organization is regulated on various levels and by a multitude of diverse proteins and
non-coding RNAs. On one hand, enzyme complexes that use DNA for transcription, replication,
recombination or repair actively contribute to changing chromatin structure. For instance, RNA and
DNA polymerases travel along the DNA double helix and by doing so introduce torsional stress that
can promote the loss of histones ahead of them and facilitate the reassembly of nucleosomes in their
wake . Although most of this stress is constantly released by the action of topoisomerases, it is
likely that DNA-utilizing processes exert distinct effects on local as well as regional chromatin
structure. Other mechanisms that profoundly affect chromatin structure are posttranslational
modifications of nucleosomal histones, the incorporation of so-called variant histone proteins and of
other non-histone architectural proteins, such as high mobility group (HMG) proteins, as well as the
energy-consuming remodeling of nucleosomes by ATP-dependent remodeling machines [7–10].
ATP-dependent chromatin remodeling factors typically are large protein complexes that contain an
ATPase subunit, which belongs to the sucrose non-fermenting 2 (SNF2) family of ATPases/helicases
[11,12]. SNF2-like ATPases can be grouped into 23 subclasses according to sequence differences in
their ATPase domains and the presence of additional protein motifs . The best-studied chromatin
remodeling factors belong to the SWI/SNF (switch/sucrose non-fermenting), the ISWI (imitation
switch), the CHD (chromo helicase DNA binding) and the INO80 (inositol auxotroph 80)
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2.1. The Role of ATP-Dependent Chromatin Remodeling Factors in Nucleosome Positioning
Several recent studies that mapped the positions of nucleosomes at a genome-wide level in different
organisms and cell types have reported the existence of rather well conserved patterns of nucleosome
occupancy in particular at the 5' and 3' ends of genes (e.g., [18–22]). Using micrococcal digestion
combined with deep-sequencing technology, it was shown for yeast, Drosophila and humans that
promoters are commonly marked by a nucleosome-free or depleted region (NDR) upstream of the
transcriptional start site (TSS). Furthermore, the first nucleosome downstream of the TSS
(+1 nucleosome) usually occupies a distinct position, which is ~50 bp downstream of the TSS in yeast
and at ~ +135 bp in Drosophila and humans [4,5]. Another NDR appears to be distinctive of 3'-ends of
genes. Upstream of this NDR a positioned nucleosome is usually detected although the latter appears
not to be universally conserved [4,5,23]. Although DNA sequence is likely to influence some of the
nucleosome positions, in particular the NDRs, it was postulated that ATP-dependent chromatin
remodeling machines play an important role in determining nucleosome positions in vivo [4,5,24]. This
is especially likely for nucleosomes that occupy energetically unfavorable positions.
Chromatin remodeling enzymes are well equipped to carry out this task. In many elegant in vitro
studies, it has been demonstrated that by using the energy derived from hydrolyzing ATP, these
enzymes can break and/or establish histone-DNA contacts. The results of these actions are manifold
and dependent on the type of remodeler as well as on the functional context [10,25–27]. Numerous
studies exploring the effects of deletion or knock-down of chromatin remodelers have found
wide-spread gene regulation defects . These effects can at least in part be attributed to a role of
these factors in positioning and remodeling of nucleosomes. Two SNF2 subfamilies in particular, the
ISWI and the CHD families, have been shown to be able to move nucleosomes to different
translational positions along the DNA (“sliding”) [29–34]. Consistent with this function, ISWI and
CHD type enzymes have been shown to be associated with active genes [35–38]. They have roles in
remodeling nucleosomes in the vicinity of the TSS [37–40], but they seem also involved in regulating
nucleosome positioning at the 3'-end of genes. In yeast it was observed that loss of Isw2 resulted in
increased production of non-coding transcripts. These transcripts originated from mis-oriented
transcription as a result of aberrant nucleosome positioning at the 3'-end of Isw2 target genes .
Likewise, yeast Chd1 was shown to be involved in organizing the nucleosomal fiber at the 3'-end of
genes, since deletion of CHD1 resulted in transcription termination defects and aberrant nucleosomal
arrangements at the 3'-ends of the CYC1 and ASC1 genes . Very recently, the Mi-2/CHD3-related
ATPase Mit1 (Mi2-like protein interacting with Clr three 1), which is part of the SHREC (Snf2/Hdac-
containing Repressor Complex) complex in Schizosaccharomyces pombewas shown to profoundly
affect nucleosome positioning globally and at specific heterochromatic sites [23,42].
Chromatin remodeling complexes of the SWI/SNF family have also been extensively characterized
in vitro and in vivo. One salient feature of this type of remodeler is its ability to disrupt nucleosome
structure more profoundly than ISWI and CHD enzymes (e.g., [43–46]). SWI/SNF enzymes can eject
histones from nucleosomes, they can transfer dimers and tetramers to other DNA molecules
(e.g., [43,47–49]) and they can catalyze nucleosome sliding reactions [10,50]. Thus, in vivo SWI/SNF
ATPases have been identified as crucial regulators of gene activation, and they have been shown to be
able to generate NDRs .
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Almost all SNF2-type motors are part of (large) protein complexes. The accessory subunits can
gravely impact on the biochemical properties of a remodeler complex. For instance, association of the
ISWI motor protein with the ATP-dependent chromatin assembly factor 1 (Acf1) subunit, strongly
stimulates the efficiency by which it can assemble and remodel nucleosomes . In a similar manner
the chromatin remodeling activity of the SWI/SNF ATPases BRG1 (brahma related gene 1) and
hBRM (human brahma) are significantly enhanced by the INI1 (integrase interactor 1) and the
brahma-associated factors BAF155 and BAF170 complex subunits . Nevertheless, a recent study
demonstrated that the ATPases themselves exhibit strikingly different characteristics with respect to
their nucleosome sliding properties. When Drosophila ISWI and CHD1 as well as human Snf2H, Brg1
and Mi-2 (dermatomyositis specific autoantigen Mi-2) were tested side by side in an in vitro sliding
assay, each remodeler moved the nucleosome to different positions although the underlying DNA
sequence was the same in all cases . Hence, it is conceivable that in vivo different chromatin
remodeling factors may establish specific local nucleosome positions in addition to histone
displacement. The action of these enzymes, therefore, will not only facilitate but also impede the
access of factors to their binding sites on the DNA.
2.2. Chromatin Remodeling Factorsin Replication-Coupled Nucleosome Assembly
During S-phase, when the DNA is replicated, chromatin is completely disassembled and
nucleosomes are reformed at the nascent daughter strands. Thereby, newly synthesized histones must
be incorporated to complement the “old” histones that are reused in the newly established
nucleosomes [54,55]. ATP-dependent factors are likely to adopt a critical position within the DNA
replication process. They are known to not only slide and restructure existing nucleosomes but also to
mediate the formation of new nucleosomes or change the histone composition of nucleosomes [8,56].
ISWI-containing remodeling complexes, such as ACF (ATP-dependent chromatin assembly and
remodeling factor)and RSF (remodeling and spacing factor), and CHD1 have been demonstrated to be
able to generate nucleosome arrays in vitro from purified histones and DNA. ACF and CHD1 perform
this reaction in conjunction with the histone chaperone NAP-1 (nucleosome assembly protein 1), while
RSF does not require a chaperone [52,57–60].
Despite the well-characterized biochemical activities of chromatin remodeling factors it is rather
surprising that information about their involvement in replication-coupled chromatin assembly in vivo
is still limited. To date, only ISWI-type enzymes have been linked to nucleosome formation during
S-phase. In Drosophila the inactivation of the ACF complex by deletion of its Acf1 subunit resulted in
an acceleration of S-phase caused by a shortening of heterochromatin replication timing .
Similarly, the human ISWI homolog SNF2h was proposed to play a role in replication-coupled
heterochromatin assembly [62–64]. In this case, two different SNF2h-containing complexes appear to
be important, since knock-down of the ACF1 subunit of the human ACF complex inhibited
progression through S-phase , while a complex containing Snf2h and the Williams syndrome
transcription factor (WSTF) targeted SNF2h to heterochromatin by interaction with proliferating cell
nuclear antigen (PCNA), which is a processivity factor of DNA polymerase [62,64]. Thus, ISWI
enzymes appear to be involved in replication-coupled heterochromatin assembly. However, in light of
more recent studies implicating ISWI in the incorporation of the linker histone H1 (see below), the
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above-mentioned observations might not fully support this conclusion. A recent report identified the
mammalian SNF2-type ATPase SMARCAD1 as an important regulator of global DNA replication-
associated histone deacetylation. As a consequence of SMARCAD1 knock-down, heterochromatin
establishment, in particular histone H3 lysine 9 trimethylation and HP1 binding was perturbed .
Thus, while SMARCAD1 appears to play a crucial role in thedeacetylation of newly incorporated
histones, which are acetylated, it seems not to be directly involved in histone deposition. Therefore, to
date no chromatin remodeler has been unequivocally demonstrated to mediate the reassembly of either
heterochromatin or euchromatin in the course of DNA replication in vivo.
2.3. Incorporation of Linker Histone H1
The linker histone H1 associates with DNA at nucleosome entry/exit sites and thereby affects the
folding of the 10 nm nucleosomal fiber into higher-order structures with a diameter of about
30 nm [66,67]. It is assumed that the 30 nm fiber makes chromatin less accessible to DNA binding
factors and is thus largely refractory for processes such as transcription. Although several recent
studies have made considerable progress in elucidating the structure of in vitro reconstituted 30 nm
fibers [68–71], their in vivo organization appears to be heterogeneous and is still poorly
understood [72,73]. This may be due in part to the highly dynamic behavior of H1 in vivo. While the
core histones H3 and H4 typically remain bound to the chromatin over several cell generations, H1
turn-over occurs within seconds [74–77].
Several lines of evidence point to a critical role for ATP-dependent chromatin remodelers in H1
assembly. First, it was shown that in vitro ACF and ISWI but not the CHD-type factor CHD1 can
generate periodic H1-containing nucleosome arrays [58,78,79]. Second, in Drosophila, deletion of
ISWI resulted in global decondensation of the transcriptionally hyperactive single X chromosome in
salivary glands of male larvae, and overexpression of a dominant negative allele of ISWI led to
striking alterations in the appearance of autosomes as well as sex chromosomes. These changes were
accompanied by a decrease in chromosomal H1 levels [80,81]. These findings suggest that ISWI is
required for the incorporation of H1 into chromatin in vivo. ISWI is part of multiple
chromatin remodeling complexes, and in a study of the largest subunit of the ISWI-containing
NURF (nucleosome remodeling factor) complex, Nurf301, it was shown that Nurf301 mutant alleles
resulted in a decondensation phenotype of the male X chromosome similar to that of an Iswi
mutation [82,83] suggesting that NURF might be involved in H1 incorporation. However, Nurf301
mutation also causes the upregulation of roX RNA, which is a central component of the male specific
lethal (MSL) complex, which is required for dosis compensation in Drosophila males. Mutations of
roX suppressed the puffing phenotype of the Nurf301 mutants , and it is not clear at this point
whether the derepression of roX in Nurf301 mutants and/or H1 incorporation defects are responsible
for the distortions in chromatin structure observed in the absence of functional Nurf301. There is also
evidence that another ISWI-containing complex, ACF, might contribute to H1 incorporation. In
mutants for the signature subunit of ACF, Acf1, a global shortening of nucleosomal repeat length was
observed . Such changes also occur when H1 levels are strongly reduced [77,84,85] and therefore
might argue for an involvement of ACF in H1 assembly. Given that Acf1 mutants do not exhibit
structural defects on the male X chromosome, it is possible that H1 loading is achieved by the
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combined actions of different ISWI complexes. Regardless of the type of complex, these studies
provide an example that ATP-dependent chromatin remodeling not only affects the structure of the
basic nucleosome fiber but also has important functions in modulating higher-order chromatin folding.
2.4. Incorporation of Variant Histones
A major manifestation of chromatin dynamics is the constant turn-over of chromosomal histones.
Even in post-mitotic cells histones are continually exchanged. During replication-coupled assembly the
so-called “canonical” histones are incorporated. These histones are encoded by multiple gene copies in
higher eukaryotes, and their expression is tightly controlled to reach its maximum in S-phase .
Canonical histones are not incorporated by replication-independent mechanisms. To this end, variant
histones are used . Consequently, in post-mitotic cells canonical histones are gradually replaced
with histone variants. For instance, measurements in long-lived neurons have shown that ~80% of all
H3 histones are of the H3.3 variant type . An important process that requires the
replication-independent assembly of histones is transcription. It has been shown that fast and profound
histone loss occurs at highly transcribed genes, such as the heat shock protein 70 (Hsp70) genes in
Drosophila . Moreover, measurements of the incorporation of GFP-tagged histone H3.3 have
revealed that H3.3 accumulates at transcriptionally active sites [89,90]. Some histone variants are
highly similar in sequence to their replication-coupled counterparts. For instance, H3.3 differs from
H3.2 with only four amino acids. On the other hand, there are histone isotypes, such as macroH2A or
H2A.Bbd (H2A Barr body deficient), whose sequence deviates considerably from the canonical
histone type (Figure 1) [91–93].
Multiple studies have shown that assembly and exchange of histones require the concerted action of
histone chaperones and ATP-dependent chromatin remodeling factors in the context of both
replication-coupled as well as replication-independent processes [56,94]. It has also become apparent
in recent years that in vivo the incorporation of individual histone variants requires distinct types of
ATP-dependent factors together with specialized histone chaperones. Although the mechanisms of
incorporation of a number of variants still await discovery, considerable progress has been made in
elucidating the critical factors involved in the incorporation of variant histones, such as H3.3, the
centromere-specific H3 variant CenH3 and the H2A variant H2A.Z [8,92,95,96].
Regardless of the mechanisms of incorporation, histone variant composition of chromatin correlates
with its functional properties and affects chromatin dynamics locally or in a global manner . For
instance, H3.3 and the H2A variant H2A.Bbd colocalize predominantly with transcriptionally active
chromatin, CenH3 is only found at centromeres, where it generates a chromatin structure suitable to
the formation of the kinetochore, and the variant histone macroH2A is distinctive of the
transcriptionally silent X-chromosome of female mammals . Different histone isotypes may affect
nucleosome and chromatin structure and dynamics in various ways. They may be subject to distinct
posttranslational modifications , alter interactions with components of the DNA-utilizing
machineries, or they may affect the structural properties of variant-containing nucleosomes in a way
that makes the underlying DNA sequence more or less permissible for a certain functional state.
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Figure 1. Schematic representation of mammalian histone isotypes (left panel) and of the
respective remodeling enzymes that have been linked to their incorporation (right panel).
Numbers in parenthesis represent the amino acid sequence lengths of the histone proteins.
Identical colors indicate identical amino acid sequences. Replication-coupled histone
incorporation is denoted by light grey shading, replication-independent assembly is
indicated by dark grey shading [91–93].
2.5. Chromatin Remodelers and H3.3
A recent crystal structure analysis revealed that incorporation of H3.3 into nucleosomes instead of
the replication-coupled H3.1 and H3.2 forms does not lead to obvious structural effects on the
nucleosome [98,99]. Nevertheless, nucleosomes purified from chicken erythroid cells were found to be
less stable when containing H3.3 . Therefore, it was proposed that a destabilization of
nucleosomes by H3.3 may promote the accessibility of active genes and regulatory regions .
Consistent with this idea is the observation that increased levels of H3.3 are detected over active genes
and at transcription factor binding sites [100–102]. Despite its broad distribution in nuclear chromatin,
H3.3 is not essential for viability of Drosophila [103,104]. Yet it is required for germ cell
development, and male and female flies with mutated H3.3 are sterile [103,104].
H3.3 is predominantly incorporated co-transcriptionally with the exception of an early instance in
development. At fertilization the chromatin of the paternal pronucleus requires drastic reorganization.
In this situation, global reassembly of nucleosomes must occur in order to replace sperm-specific DNA
packaging proteins, the protamines, that are responsible for organizing sperm chromatin. It has been
shown in Drosophila and mouse embryos that during paternal pronuclear rearrangement the histone
variant H3.3 but not H3.1 is loaded onto the DNA [105,106]. In Drosophila, the protamine/histone
Int. J. Mol. Sci. 2011, 12
exchange takes place prior to the onset of DNA replication and transcription and thus, H3.3 deposition
must be independent of a transcription-linked process. The histone chaperone HIRA (histone cell cycle
regulation defective homolog A) was identified as a crucial factor for the loading of H3.3 in this
process, since mutation of HIRA abrogated the incorporation of H3.3 into the paternal
chromatin [105,107]. Similar but not identical defects were observed when the ATP-dependent
chromatin remodeler CHD1 was deleted in the fly. The absence of CHD1 resulted in the accumulation
of H3.3 at the nuclear periphery of paternal pronuclei indicating that CHD1 is required for correct
deposition of H3.3 . Thus, CHD1 and HIRA appear to work together in the
transcription-independent incorporation of H3.3 at this specific developmental instance, a notion that is
corroborated by the observation that both factors physically interact in early Drosophila embryos 
CHD1 may also have a role in the transcription-dependent incorporation of H3.3. This idea is
supported by the finding that in Chd1-defective Drosophila embryos, aberrant H3.3 localization is
detected in transcriptionally active syncytial nuclei . In addition, knock-down of CHD1 in mouse
embryonic stem (ES) cells resulted in compromised pluripotency probably due to an observed decrease
of euchromatin and a concomitant spreading of heterochromatin . Although H3.3 incorporation
was not tested in this study, the fact that H3.3 normally is enriched in euchromatin may point to a
defect in generating proper H3.3-containing nucleosomes. Interestingly, whole-genome H3.3 mapping
experiments have revealed that mammalian HIRA is necessary for H3.3 enrichment at active and
repressed genes [100,110,111] (Figure 2b).
A number of recent reports have implicated another SNF2-type chromatin remodeling factor, the
α-thalassemia/mental retardation syndrome X-linked (ATRX) protein, in H3.3 incorporation into
chromatin. ATRX was shown to be required for loading of H3.3 into chromatin at telomeres in mouse
ES cells [100,110,111] and at pericentric heterochromatin in mouse embryonic fibroblasts 
(Figure 2b). Thorough biochemical analyses revealed that ATRX cooperates with a novel
H3.3-specific histone chaperone termed DAXX (death domain associated protein) [100,111,112].
Thus, to date two different ATP-dependent chromatin remodeling factors have been implicated in H3.3
incorporation. Both factors function together with distinct chaperones (CHD1 with HIRA, ATRX with
DAXX) to generate H3.3-containing nucleosomes in specific nuclear neighborhoods or developmental
occasions reflecting a highly complex assembly machinery that enables the formation of functionally
distinct chromatin areas (Figure 2b).
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Figure 2. Replication-independent assembly of histone H3 variants. (a) The chromatin
remodeler CHD1 cooperates with the H3.3-specific histone chaperone HIRA to
incorporate H3.3 into the paternal pronucleus at fertilization in Drosophila embryos. The
maternal pronucleus does not require chromatin reorganization and contains predominantly
H3.1. (b) Different chromatin remodeling complexes in conjunction with specific histone
chaperones incorporate H3.3 and CENP-A at distinct chromosomal sites. Dark blue
shading indicates H3.3 incorporation into telomeric and pericentric heterochromatin,
respectively. Lighter blue shading indicates H3.3 assembly at genic locations. Green
shading denotes CENP-A incorporation into chromatin at the centromere.
2.6. Chromatin Remodelers and the Assembly of Centromeric Chromatin
The histone H3 variant CenH3 (also known as CENP-A, CID, Cnp1, Cse4) is incorporated into
chromatin at the centromeres in a transcription-independent fashion [113,114]. Its presence at the
centromere is thought to identify the region for kinetochore assembly, since centromeric DNA
sequences are not conserved between organisms and therefore not likely to contribute to this
task [113,115]. The assembly of centromeric chromatin appears to involve a great number of proteins.
Despite considerable research efforts over the past years it is still not entirely clear as to which factors
are directly involved in CenH3 assembly and which ones act in an indirect manner [113–116]. For
example, a recent study showed that the yeast SWI/SNF complex acts to remove the yeast centromeric
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histone Cse4 from nucleosomes outside of the centromere. Thus, it acts to confine Cse4 to the single
centromeric nucleosome that defines centromeres in Saccharomyces but is not involved in the loading
of Cse4 . A large step towards elucidating centromeric chromatin assembly has been made with
the discovery of a CenH3-specific histone chaperone, termed HJURP (Holliday junction recognition
protein) [118,119]. HJURP has been demonstrated to directly interact with soluble CenH3. Moreover,
knock-down of HJURP led to the loss of CenH3 signals at the centromere [118,119]. Interestingly,
HJURP is distantly related to the Saccharomyces cerevisiae Scm3 protein, which also has been shown
to act as a CenH3 chaperone [120,121], but does not have any apparent homologs in Drosophila .
Among the ATP-dependent factors the ISWI-containing complex RSF was reported to interact with
CenH3-containing mononucleosomes in human cells and to play a role in the incorporation of human
CenH3 . However, as the effects of RSF knock-down are relatively mild, it is likely that there are
additional proteins involved . Indeed, in chicken DT40 cells as well as in fission yeast, CHD1
and its homolog Hrp1, respectively, have been linked to a role in the assembly of CenH3-containing
nucleosomes [124,125]. In contrast, no such role could be demonstrated for CHD1 in
Drosophila . Similarly, no centromere defects have been reported for Drosophila Rsf1
mutants ). Thus, flies appear to neither possess a bona fide HJURP homolog, nor are the roles of
CHD1 and RSF in CenH3 incorporation conserved. Instead, a Drosophila-specific factor, termed
CAL1 (chromosome alignment defect 1), was demonstrated to interact with CenH3 and to be required
for its loading to chromatin . To date, no ATP-dependent factor was found to participate in this
process. These results indicate that different organisms might use different mechanisms and factors to
ensure CenH3 assembly at centromeres.
2.7. Chromatin Remodelers and H2A.Z Exchange
The replacement of H2A/H2B dimers in nucleosomes with dimers containing the variant histone
H2A.Z/H2B is a common event in all eukaryotes. Its importance is emphasized by the fact that H2A.Z
is essential for viability in Drosophila, Tetrahymena and mouse [129–131]. Incorporation of H2A.Z
into nucleosomes does not result in large structural alterations, but nevertheless causes some intriguing
changes. On one hand, H2A.Z-containing nucleosomes possess a larger acidic patch at the surface of
the octamer that was proposed to serve in the interaction with the H4 N-terminal tail of a neighboring
nucleosome . Indeed, in vitro H2A.Z-containing nucleosome arrays were shown to be more
tightly compacted than H2A-containing nucleosomes . These observations are consistent with the
results from genome-wide analyses of H2A.Z distribution that found that H2A.Z is present in
heterochromatic areas of the genome [134,135].
On the other hand, however, the interface between H2A.Z/H2B and H3/H4 dimers in the crystal
structure was found to be slightly less stable than in H2A-nucleosomes and thus may render these
nucleosomes more prone to disruption . Aside from heterochromatic sites, H2A.Z is particularly
enriched in nucleosomes at the transcription start sites of genes [99,136]. In line with the predictions of
the crystal structure analysis, H2A.Z-containing nucleosomes isolated from chicken erythroid cells
displayed reduced stability [99,136]. However, H2A.Z-containing nucleosomes were only less stable
when they simultaneously contained H3.3 but not when they contained H3 [99,136]. Hence, it appears
that the combination of H2A.Z with either H3.3 or H3.1 confers quite distinct properties to these
Int. J. Mol. Sci. 2011, 12
nucleosomes. This may in part explain the seemingly contradictory presence of H2A.Z in
heterochromatin and euchromatin.
H2A.Z replacement is carried out by a dedicated ATP-dependent remodeler, termed
SWR1 [137–141] (Figure 1). SWR1 belongs to the INO80 subclass of chromatin remodelers and is
characterized by a split ATPase domain . It is part of a multiprotein complex, which also contains
subunits necessary for H2A.Z recognition and for binding to acetylated H3/H4 . A recently
published study demonstrated that the second member of the INO80 subfamily, INO80, also affects
H2A.Z-containing nucleosomes  (Figure 1). Interestingly, INO80 performs the opposite reaction
to SWR1 by catalyzing the exchange of H2A.Z/H2B dimers for H2A/H2B. Deletion of INO80 in yeast
resulted in aberrant localization of H2A.Z in promoter and coding regions. Moreover, replication fork
progression defects of ∆ino80 mutants were alleviated by reduced expression of H2A.Z, suggesting
that the misincorporation of H2A.Z in the absence of INO80 causes the observed defect .
A number of histone chaperones have been implicated in H2A.Z dynamics. Nap1 was shown to
enable H2A.Z/H2B dimer exchange in an in vitro reaction  and was also detected in purified
SWR1 complex fractions . In S. cerevisiae, another H2A.Z-specific chaperone, termed Chz1, was
identified, which together with Nap1 represents the two major H2A.Z/H2B chaperones in this
organism . Interestingly in the absence of both Chz1 and Nap1 additional proteins, such as the
FACT complex, the karyopherin Kap114 and two peptidylprolyl cis-trans isomerases termed Fpr3 and
Fpr4, were shown to interact with H2A.Z/H2B dimers . Further studies in yeast indicated that
NAP-1 is important for chaperoning the soluble pool of H2A.Z, whereas Chz1 does not interact with
H2A.Z in the cytoplasm .
3. Do Chromatin Remodeling Factors Incorporate Non-Histone Chromosomal Proteins?
As discussed above, ATP-dependent remodeling factors are crucial components of the machineries
that deposit histones and generate patterns of nucleosomes with diverse composition (Figure 1). Apart
from histones and their variants, however, there are other abundant non-histone architectural proteins,
such as HMG proteins or heterochromatin protein 1 (HP1), that associate with the chromatin and shape
its structure and dynamics. Do these factors also require motor proteins to bind correctly to the
nucleosome fiber? Although this intriguing question has not been investigated in great detail so far,
some recent studies provide evidence that suggests this may indeed be the case.
The HMG proteins are among those non-histone architectural proteins that have been studied most
extensively [9,147,148]. HMG proteins generally act to decrease the compactness of the chromatin
fiber and therefore render chromatin more accessible to regulatory factors [147,149]. They bind to
chromatin in a highly dynamic and reversible way either by directly contacting the nucleosome and/or
via co-factors. There are three subfamilies of HMG proteins, termed HMGA, HMGB and
HMGN [9,148]. Members of the HMGN group, in particular, have been shown to bind to nucleosomes
at the entry/exit sites of the DNA and therefore compete with the linker histone H1 for nucleosome
binding sites [150,151]. They also exhibit exchange dynamics that are similar to those of H1 . As
detailed above, H1 incorporation into chromatin is strongly dependent on the ISWI chromatin
assembly factor [61,81]. By analogy, HMGN proteins might also require an ATP-dependent factor for
efficient chromatin association. In a recent study addressing the effects of HMGN1 and HMGN2 on
Int. J. Mol. Sci. 2011, 12
chromatin remodeling by the ATP-dependent factors ACF and the SWI/SNF-family protein BRG1, it
was shown that ACF can assemble extended periodic nucleosome arrays containing HMGN proteins in
vitro . Although in vivo studies have not yet been carried out, these experiments provide an
intriguing hint for a possible function of ACF and potentially other chromatin remodeling factors in
the assembly of not only histones but also of non-histone architectural proteins into chromatin.
Another candidate remodeling factor for the incorporation of non-histone chromosomal proteins
may be ATRX. As discussed above, ATRX has recently been characterized to be required for the
incorporation of histone H3.3 into pericentric and telomeric chromatin [100,110–112]. Yet, ATRX has
also been shown to physically interact with HP1, which is an abundant protein localized in
heterochromatin [154–157]. Two recent studies provided evidence for a function of ATRX in the
loading of HP1 to chromatin. In Drosophila, deletion of ATRX resulted in the loss of HP1α from
pericentricβ-heterochromatin . Along the same lines, depletion of ATRX in mouse ES cells led to
a strong decrease of HP1α localization at telomeric chromatin . Although these findings point to a
role of ATRX in the association of HP1α with heterochromatin, biochemical studies will be necessary
to determine, if indeed ATRX uses its catalytic activity to incorporate HP1 or if the observed
phenotypes are the result of recruitment defects. Previous in vitro experiments have demonstrated that
the ACF remodeling factor greatly stimulates the association of HP1 with reconstituted nucleosome
arrays. This stimulation, however, was found to be dependent on the Acf1 subunit of the complex and
did not involve ATP-hydrolysis . Nevertheless, although the evidence is somewhat circumstantial
at the moment, it will be interesting to elucidate whether non-histone architectural proteins are actual
targets of ATP-dependent chromatin remodeling machines.
Nucleosome assembly is not only necessary for preserving chromatin structure and, thus, genome
integrity, it is also a process that strongly impacts on the functional properties of the nucleosomal fiber.
In particular, the incorporation of specific histone variants into nucleosomes on one hand serves to
modulate the biophysical properties of nucleosomes but can also endow nucleosomes with distinct
abilities to interact with regulatory factors or to receive specific posttranslational modification marks.
Moreover, the presence of histone variants at functionally distinct regions in the genome has been
postulated to serve as a means to transmit epigenetic information across cell generations [159,160].
With the discovery of several novel histone incorporation pathways over the past years, it has become
clear that ATP-dependent chromatin remodeling factors in conjunction with specific histone
chaperones act at the center of these processes. It will be interesting to see in the future, whether
dedicated partnerships of ATP-dependent motor proteins with histone chaperones exist for the
assembly of all histone variants and even non-histone architectural proteins and what the biological
consequences of their actions are.
We apologize to all colleagues whose work we could not cite due to space limitations. We are
grateful to Peter Loidl for critical reading of the manuscript. Research in the authors’ lab is supported
by the Austrian Science Fund (FWF):START Y275-B12; F4408-B19.
Int. J. Mol. Sci. 2011, 12
1. Luger, K.; Mader, A.W.; Richmond, R.K.; Sargent, D.F.; Richmond, T.J. Crystal structure of the
nucleosome core particle at 2.8 A resolution. Nature 1997, 389, 251–260.
Olins, A.L.; Olins, D.E. Spheroid chromatin units (v bodies). Science 1974, 183, 330–332.
Woodcock, C.L.; Safer, J.P.; Stanchfield, J.E. Structural repeating units in chromatin. I. Evidence
for their general occurrence. Exp. Cell Res. 1976, 97, 101–110.
Jiang, C.; Pugh, B.F. Nucleosome positioning and gene regulation: advances through genomics.
Nat. Rev. Genet. 2009, 10, 161–172.
Radman-Livaja, M.; Rando, O.J. Nucleosome positioning: how is it established, and why does it
matter? Dev. Biol. 2010, 339, 258–266.
Lavelle, C. Transcription elongation through a chromatin template. Biochimie 2007, 89, 516–527.
Kouzarides, T. Chromatin modifications and their function. Cell 2007, 128, 693–705.
Jin, J.; Cai, Y.; Li, B.; Conaway, R.C.; Workman, J.L.; Conaway, J.W.; Kusch, T. In and out:
histone variant exchange in chromatin. Trends Biochem. Sci. 2005, 30, 680–687.
Hock, R.; Furusawa, T.; Ueda, T.; Bustin, M. HMG chromosomal proteins in development and
disease. Trends Cell Biol. 2007, 17, 72–79.
10. Clapier, C.R.; Cairns, B.R. The biology of chromatin remodeling complexes. Annu. Rev.
Biochem. 2009, 78, 273–304.
11. Flaus, A.; Martin, D.M.; Barton, G.J.; Owen-Hughes, T. Identification of multiple distinct Snf2
subfamilies with conserved structural motifs. Nucleic Acids Res. 2006, 34, 2887–2905.
12. Gorbalenya, A.E. Helicases: amino acid sequence comparisons and structure-function
relationships. Curr. Opin. Struct. Biol. 1993, 3, 419–429.
13. Eberharter, A.; Becker, P.B. ATP-dependent nucleosome remodelling: factors and functions.
J. Cell Sci. 2004, 117, 3707–3711.
14. Lusser, A.; Kadonaga, J.T. Chromatin remodeling by ATP-dependent molecular machines.
Bioessays 2003, 25, 1192–1200.
15. Bao, Y.; Shen, X. INO80 subfamily of chromatin remodeling complexes. Mutat. Res. 2007, 618,
16. Marfella, C.G.; Imbalzano, A.N. The Chd family of chromatin remodelers. Mutat. Res. 2007,
17. Hargreaves, D.C.; Crabtree, G.R. ATP-dependent chromatin remodeling: genetics, genomics and
mechanisms. Cell Res. 2011, 21, 396–420.
18. Yuan, G.C.; Liu, Y.J.; Dion, M.F.; Slack, M.D.; Wu, L.F.; Altschuler, S.J.; Rando, O.J.
Genome-scale identification of nucleosome positions in S. cerevisiae. Science 2005, 309,
19. Mavrich, T.N.; Ioshikhes, I.P.; Venters, B.J.; Jiang, C.; Tomsho, L.P.; Qi, J.; Schuster, S.C.;
Albert, I.; Pugh, B.F. A barrier nucleosome model for statistical positioning of nucleosomes
throughout the yeast genome. Genome Res. 2008, 18, 1073–1083.
20. Mavrich, T.N.; Jiang, C.; Ioshikhes, I.P.; Li, X.; Venters, B.J.; Zanton, S.J.; Tomsho, L.P.; Qi, J.;
Glaser, R.L.; Schuster, S.C.; et al. Nucleosome organization in the Drosophila genome. Nature
2008, 453, 358–362.
Int. J. Mol. Sci. 2011, 12
21. Valouev, A.; Ichikawa, J.; Tonthat, T.; Stuart, J.; Ranade, S.; Peckham, H.; Zeng, K.; Malek, J.A.;
Costa, G.; McKernan, K.; et al. A high-resolution, nucleosome position map of C. elegans
reveals a lack of universal sequence-dictated positioning. Genome Res. 2008, 18, 1051–1063.
22. Schones, D.E.; Cui, K.; Cuddapah, S.; Roh, T.Y.; Barski, A.; Wang, Z.; Wei, G.; Zhao, K.
Dynamic regulation of nucleosome positioning in the human genome. Cell 2008, 132, 887–898.
23. Lantermann, A.B.; Straub, T.; Stralfors, A.; Yuan, G.C.; Ekwall, K.; Korber, P.
Schizosaccharomyces pombe genome-wide nucleosome mapping reveals positioning
mechanisms distinct from those of Saccharomyces cerevisiae. Nat. Struct. Mol. Biol. 2010, 17,
24. Zhang, Z.; Wippo, C.J.; Wal, M.; Ward, E.; Korber, P.; Pugh, B.F. A packing mechanism for
nucleosome organization reconstituted across a eukaryotic genome. Science 2011, 332, 977–980.
25. Flaus, A.; Owen-Hughes, T. Mechanisms for ATP-dependent chromatin remodelling: the means
to the end. FEBS J. 2011, 278, 3579–3595.
26. Langst, G.; Becker, P.B. Nucleosome remodeling: one mechanism, many phenomena? Biochim.
Biophys. Acta 2004, 1677, 58–63.
27. Morettini, S.; Podhraski, V.; Lusser, A. ATP-dependent chromatin remodeling enzymes and their
various roles in cell cycle control. Front. Biosci. 2008, 13, 5522–5532.
28. Moshkin, Y.M.; Mohrmann, L.; van Ijcken, W.F.; Verrijzer, C.P. Functional differentiation of
SWI/SNF remodelers in transcription and cell cycle control. Mol. Cell. Biol. 2007, 27, 651–661.
29. Guschin, D.; Wade, P.A.; Kikyo, N.; Wolffe, A.P. ATP-Dependent histone octamer mobilization
and histone deacetylation mediated by the Mi-2 chromatin remodeling complex. Biochemistry
(Mosc.) 2000, 39, 5238–5245.
30. Brehm, A.; Langst, G.; Kehle, J.; Clapier, C.R.; Imhof, A.; Eberharter, A.; Muller, J.; Becker,
P.B. dMi-2 and ISWI chromatin remodelling factors have distinct nucleosome binding and
mobilization properties. EMBO J. 2000, 19, 4332–4341.
31. Stockdale, C.; Flaus, A.; Ferreira, H.; Owen-Hughes, T. Analysis of nucleosome repositioning by
yeast ISWI and Chd1 chromatin remodeling complexes. J. Biol. Chem. 2006, 281, 16279–16288.
32. Langst, G.; Bonte, E.J.; Corona, D.F.; Becker, P.B. Nucleosome movement by CHRAC and
ISWI without disruption or trans- displacement of the histone octamer. Cell 1999, 97, 843–852.
33. Hamiche, A.; Sandaltzopoulos, R.; Gdula, D.A.; Wu, C. ATP-dependent histone octamer sliding
mediated by the chromatin remodeling complex NURF. Cell 1999, 97, 833–842.
34. Rippe, K.; Schrader, A.; Riede, P.; Strohner, R.; Lehmann, E.; Langst, G. DNA sequence- and
conformation-directed positioning of nucleosomes by chromatin-remodeling complexes. Proc.
Natl. Acad. Sci. USA 2007, 104, 15635–15640.
35. Stokes, D.G.; Tartof, K.D.; Perry, R.P. CHD1 is concentrated in interbands and puffed regions of
Drosophila polytene chromosomes. Proc. Natl. Acad. Sci. USA 1996, 93, 7137–7142.
36. Srinivasan, S.; Armstrong, J.A.; Deuring, R.; Dahlsveen, I.K.; McNeill, H.; Tamkun, J.W. The
Drosophila trithorax group protein Kismet facilitates an early step in transcriptional elongation
by RNA Polymerase II. Development 2005, 132, 1623–1635.
37. Whitehouse, I.; Tsukiyama, T. Antagonistic forces that position nucleosomes in vivo. Nat. Struct.
Mol. Biol. 2006, 13, 633–640.
Int. J. Mol. Sci. 2011, 12
38. Sala, A.; Toto, M.; Pinello, L.; Gabriele, A.; Di Benedetto, V.; Ingrassia, A.M.; Lo Bosco, G.;
Di Gesu, V.; Giancarlo, R.; Corona, D.F. Genome-wide characterization of chromatin binding
and nucleosome spacing activity of the nucleosome remodelling ATPase ISWI. EMBO J. 2011,
39. Petesch, S.J.; Lis, J.T. Rapid, transcription-independent loss of nucleosomes over a large
chromatin domain at Hsp70 loci. Cell 2008, 134, 74–84.
40. Morettini, S.; Tribus, M.; Zeilner, A.; Sebald, J.; Campo-Fernandez, B.; Scheran, G.; Worle, H.;
Podhraski, V.; Fyodorov, D.V.; Lusser, A. The chromodomains of CHD1 are critical for
enzymatic activity but less important for chromatin localization. Nucleic Acids Res. 2011, 39,
41. Alen, C.; Kent, N.A.; Jones, H.S.; O’Sullivan, J.; Aranda, A.; Proudfoot, N.J. A role for
chromatin remodeling in transcriptional termination by RNA polymerase II. Mol. Cell 2002, 10,
42. Sugiyama, T.; Cam, H.P.; Sugiyama, R.; Noma, K.; Zofall, M.; Kobayashi, R.; Grewal, S.I.
SHREC, an effector complex for heterochromatic transcriptional silencing. Cell 2007, 128,
43. Kwon, H.; Imbalzano, A.N.; Khavari, P.A.; Kingston, R.E.; Green, M.R. Nucleosome disruption
and enhancement of activator binding by a human SW1/SNF complex. Nature 1994, 370,
44. Cairns, B.R.; Lorch, Y.; Li, Y.; Zhang, M.; Lacomis, L.; Erdjument-Bromage, H.; Tempst, P.;
Du, J.; Laurent, B.; Kornberg, R.D. RSC, an essential, abundant chromatin-remodeling complex.
Cell 1996, 87, 1249–1260.
45. Cote, J.; Quinn, J.; Workman, J.L.; Peterson, C.L. Stimulation of GAL4 derivative binding to
nucleosomal DNA by the yeast SWI/SNF complex. Science 1994, 265, 53–60.
46. Imbalzano, A.N.; Schnitzler, G.R.; Kingston, R.E. Nucleosome disruption by human SWI/SNF is
maintained in the absence of continued ATP hydrolysis. J. Biol. Chem. 1996, 271, 20726–20733.
47. Lorch, Y.; Zhang, M.; Kornberg, R.D. RSC unravels the nucleosome. Mol. Cell 2001, 7, 89–95.
48. Owen-Hughes, T.; Utley, R.T.; Cote, J.; Peterson, C.L.; Workman, J.L. Persistent site-specific
remodeling of a nucleosome array by transient action of the SWI/SNF complex. Science 1996,
49. Dechassa, M.L.; Sabri, A.; Pondugula, S.; Kassabov, S.R.; Chatterjee, N.; Kladde, M.P.;
Bartholomew, B. SWI/SNF has intrinsic nucleosome disassembly activity that is dependent on
adjacent nucleosomes. Mol. Cell 2010, 38, 590–602.
50. Whitehouse, I.; Flaus, A.; Cairns, B.R.; White, M.F.; Workman, J.L.; Owen-Hughes, T.
Nucleosome mobilization catalysed by the yeast SWI/SNF complex. Nature 1999, 400, 784–787.
51. Hartley, P.D.; Madhani, H.D. Mechanisms that specify promoter nucleosome location and
identity. Cell 2009, 137, 445–458.
52. Ito, T.; Levenstein, M.E.; Fyodorov, D.V.; Kutach, A.K.; Kobayashi, R.; Kadonaga, J.T. ACF
consists of two subunits, Acf1 and ISWI, that function cooperatively in the ATP-dependent
catalysis of chromatin assembly. Genes Dev. 1999, 13, 1529–1539.
53. Phelan, M.L.; Sif, S.; Narlikar, G.J.; Kingston, R.E. Reconstitution of a core chromatin
remodeling complex from SWI/SNF subunits. Mol. Cell 1999, 3, 247–253.
Int. J. Mol. Sci. 2011, 12
54. Groth, A.; Rocha, W.; Verreault, A.; Almouzni, G. Chromatin challenges during DNA
replication and repair. Cell 2007, 128, 721–733.
55. Ransom, M.; Dennehey, B.K.; Tyler, J.K. Chaperoning histones during DNA replication and
repair. Cell 2010, 140, 183–195.
56. Lusser, A.; Kadonaga, J.T. Strategies for the reconstitution of chromatin. Nat. Methods 2004, 1,
57. LeRoy, G.; Loyola, A.; Lane, W.S.; Reinberg, D. Purification and characterization of a human
factor that assembles and remodels chromatin. J. Biol. Chem. 2000, 275, 14787–14790.
58. Lusser, A.; Urwin, D.L.; Kadonaga, J.T. Distinct activities of CHD1 and ACF in ATP-dependent
chromatin assembly. Nat. Struct. Mol. Biol. 2005, 12, 160–166.
59. Torigoe, S.E.; Urwin, D.L.; Ishii, H.; Smith, D.E.; Kadonaga, J.T. Identification of a Rapidly
Formed Nonnucleosomal Histone-DNA Intermediate that Is Converted into Chromatin by ACF.
Mol. Cell 2011, 43, 638–648.
60. Robinson, K.M.; Schultz, M.C. Replication-independent assembly of nucleosome arrays in a
novel yeast chromatin reconstitution system involves antisilencing factor Asf1p and
chromodomain protein Chd1p. Mol. Cell. Biol. 2003, 23, 7937–7946.
61. Fyodorov, D.V.; Blower, M.D.; Karpen, G.H.; Kadonaga, J.T. Acf1 confers unique activities to
ACF/CHRAC and promotes the formation rather than disruption of chromatin in vivo. Genes
Dev. 2004, 18, 170–183.
62. Poot, R.A.; Bozhenok, L.; van den Berg, D.L.; Steffensen, S.; Ferreira, F.; Grimaldi, M.; Gilbert, N.;
Ferreira, J.; Varga-Weisz, P.D. The Williams syndrome transcription factor interacts with PCNA
to target chromatin remodelling by ISWI to replication foci. Nat. Cell Biol. 2004, 6, 1236–1244.
63. Collins, N.; Poot, R.A.; Kukimoto, I.; Garcia-Jimenez, C.; Dellaire, G.; Varga-Weisz, P.D. An
ACF1-ISWI chromatin-remodeling complex is required for DNA replication through
heterochromatin. Nat. Genet. 2002, 32, 627–632.
64. Bozhenok, L.; Wade, P.A.; Varga-Weisz, P. WSTF-ISWI chromatin remodeling complex targets
heterochromatic replication foci. EMBO J. 2002, 21, 2231–2241.
65. Rowbotham, S.P.; Barki, L.; Neves-Costa, A.; Santos, F.; Dean, W.; Hawkes, N.; Choudhary, P.;
Will, W.R.; Webster, J.; Oxley, D.; et al. Maintenance of silent chromatin through replication
requires SWI/SNF-like chromatin remodeler SMARCAD1. Mol. Cell 2011, 42, 285–296.
66. Thoma, F.; Koller, T.; Klug, A. Involvement of histone H1 in the organization of the nucleosome
and of the salt-dependent superstructures of chromatin. J. Cell Biol. 1979, 83, 403–427.
67. Marsden, M.P.; Laemmli, U.K. Metaphase chromosome structure: evidence for a radial loop
model. Cell 1979, 17, 849-858.
68. Routh, A.; Sandin, S.; Rhodes, D. Nucleosome repeat length and linker histone stoichiometry
determine chromatin fiber structure. Proc. Natl. Acad. Sci. USA 2008, 105, 8872–8877.
69. Robinson, P.J.; Rhodes, D. Structure of the “30 nm” chromatin fibre: a key role for the linker
histone. Curr. Opin. Struct. Biol. 2006, 16, 336–343.
70. Robinson, P.J.; Fairall, L.; Huynh, V.A.; Rhodes, D. EM measurements define the dimensions of
the “30-nm” chromatin fiber: evidence for a compact, interdigitated structure. Proc. Natl. Acad.
Sci. USA 2006, 103, 6506–6511.
Int. J. Mol. Sci. 2011, 12
71. Schalch, T.; Duda, S.; Sargent, D.F.; Richmond, T.J. X-ray structure of a tetranucleosome and its
implications for the chromatin fibre. Nature 2005, 436, 138–141.
72. Woodcock, C.L.; Horowitz, R.A. Chromatin organization re-viewed. Trends Cell Biol. 1995, 5,
73. Horowitz, R.A.; Agard, D.A.; Sedat, J.W.; Woodcock, C.L. The three-dimensional architecture
of chromatin in situ: electron tomography reveals fibers composed of a continuously variable
zig-zag nucleosomal ribbon. J. Cell Biol. 1994, 125, 1–10.
74. Kimura, H.; Cook, P.R. Kinetics of core histones in living human cells: little exchange of H3 and
H4 and some rapid exchange of H2B. J. Cell Biol. 2001, 153, 1341–1353.
75. Misteli, T.; Gunjan, A.; Hock, R.; Bustin, M.; Brown, D.T. Dynamic binding of histone H1 to
chromatin in living cells. Nature 2000, 408, 877–881.
76. Lever, M.A.; Th’ng, J.P.; Sun, X.; Hendzel, M.J. Rapid exchange of histone H1.1 on chromatin
in living human cells. Nature 2000, 408, 873–876.
77. Siriaco, G.; Deuring, R.; Chioda, M.; Becker, P.B.; Tamkun, J.W. Drosophila ISWI regulates the
association of histone H1 with interphase chromosomes in vivo. Genetics 2009, 182, 661–669.
78. Fyodorov, D.V.; Kadonaga, J.T. Chromatin assembly in vitro with purified recombinant ACF
and NAP-1. Methods Enzymol. 2003, 371, 499–515.
79. Maier, V.K.; Chioda, M.; Rhodes, D.; Becker, P.B. ACF catalyses chromatosome movements in
chromatin fibres. EMBO J. 2008, 27, 817–826.
80. Deuring, R.; Fanti, L.; Armstrong, J.A.; Sarte, M.; Papoulas, O.; Prestel, M.; Daubresse, G.;
Verardo, M.; Moseley, S.L.; Berloco, M.; et al. The ISWI chromatin-remodeling protein is
required for gene expression and the maintenance of higher order chromatin structure in vivo.
Mol. Cell 2000, 5, 355–365.
81. Corona, D.F.; Siriaco, G.; Armstrong, J.A.; Snarskaya, N.; McClymont, S.A.; Scott, M.P.;
Tamkun, J.W. ISWI regulates higher-order chromatin structure and histone H1 assembly in vivo.
PLoS Biol. 2007, 5, e232.
82. Badenhorst, P.; Voas, M.; Rebay, I.; Wu, C. Biological functions of the ISWI chromatin
remodeling complex NURF. Genes Dev. 2002, 16, 3186–3198.
83. Bai, X.; Larschan, E.; Kwon, S.Y.; Badenhorst, P.; Kuroda, M.I. Regional control of chromatin
organization by noncoding roX RNAs and the NURF remodeling complex in Drosophila
melanogaster. Genetics 2007, 176, 1491–1499.
84. Fan, Y.; Nikitina, T.; Morin-Kensicki, E.M.; Zhao, J.; Magnuson, T.R.; Woodcock, C.L.;
Skoultchi, A.I. H1 linker histones are essential for mouse development and affect nucleosome
spacing in vivo.Mol. Cell. Biol. 2003, 23, 4559–4572.
85. Lu, X.; Wontakal, S.N.; Emelyanov, A.V.; Morcillo, P.; Konev, A.Y.; Fyodorov, D.V.;
Skoultchi, A.I. Linker histone H1 is essential for Drosophila development, the establishment of
pericentric heterochromatin, and a normal polytene chromosome structure. Genes Dev. 2009, 23,
86. Gunjan, A.; Paik, J.; Verreault, A. Regulation of histone synthesis and nucleosome assembly.
Biochimie 2005, 87, 625–635.
87. Henikoff, S.; Ahmad, K. Assembly of variant histones into chromatin. Annu. Rev. Cell Dev. Biol.
2005, 21, 133–153.
Int. J. Mol. Sci. 2011, 12
88. Pina, B.; Suau, P. Changes in histones H2A and H3 variant composition in differentiating and
mature rat brain cortical neurons. Dev. Biol. 1987, 123, 51–58.
89. Ahmad, K.; Henikoff, S. Histone H3 variants specify modes of chromatin assembly. Proc. Natl.
Acad. Sci. USA 2002, 99, 16477–16484.
90. Ahmad, K.; Henikoff, S. The histone variant H3.3 marks active chromatin by
replication-independent nucleosome assembly. Mol. Cell 2002, 9, 1191–1200.
91. Bernstein, E.; Hake, S.B. The nucleosome: a little variation goes a long way. Biochem. Cell Biol.
2006, 84, 505–517.
92. Talbert, P.B.; Henikoff, S. Histone variants–ancient wrap artists of the epigenome. Nat. Rev.
Mol. Cell Biol. 2010, 11, 264–275.
93. Wiedemann, S.M.; Mildner, S.N.; Bonisch, C.; Israel, L.; Maiser, A.; Matheisl, S.; Straub, T.;
Merkl, R.; Leonhardt, H.; Kremmer, E.; et al. Identification and characterization of two novel
primate-specific histone H3 variants, H3.X and H3.Y. J. Cell Biol. 2010, 190, 777–791.
94. Haushalter, K.A.; Kadonaga, J.T. Chromatin assembly by DNA-translocating motors. Nat. Rev.
Mol. Cell Biol. 2003, 4, 613–620.
95. Mellone, B.G.; Zhang, W.; Karpen, G.H. Frodos found: Behold the CENP-a “Ring” bearers. Cell
2009, 137, 409–412.
96. Polo, S.E.; Almouzni, G. Chromatin assembly: a basic recipe with various flavours. Curr. Opin.
Genet. Dev. 2006, 16, 104–111.
97. Loyola, A.; Almouzni, G. Marking histone H3 variants: how, when and why? Trends Biochem.
Sci. 2007, 32, 425–433.
98. Tachiwana, H.; Osakabe, A.; Shiga, T.; Miya, Y.; Kimura, H.; Kagawa, W.; Kurumizaka, H.
Structures of human nucleosomes containing major histone H3 variants. Acta Crystallogr. D
Biol.Crystallogr. 2011, 67, 578–583.
99. Jin, C.; Felsenfeld, G. Nucleosome stability mediated by histone variants H3.3 and H2A.Z.
Genes Dev. 2007, 21, 1519–1529.
100. Goldberg, A.D.; Banaszynski, L.A.; Noh, K.M.; Lewis, P.W.; Elsaesser, S.J.; Stadler, S.;
Dewell, S.; Law, M.; Guo, X.; Li, X.; et al. Distinct factors control histone variant H3.3
localization at specific genomic regions. Cell 2010, 140, 678–691.
101. Jin, C.; Zang, C.; Wei, G.; Cui, K.; Peng, W.; Zhao, K.; Felsenfeld, G. H3.3/H2A.Z double
variant-containing nucleosomes mark “nucleosome-free regions” of active promoters and other
regulatory regions. Nat. Genet. 2009, 41, 941–945.
102. Mito, Y.; Henikoff, J.G.; Henikoff, S. Genome-scale profiling of histone H3.3 replacement
patterns. Nat. Genet. 2005, 37, 1090–1097.
103. Sakai, A.; Schwartz, B.E.; Goldstein, S.; Ahmad, K. Transcriptional and developmental functions
of the H3.3 histone variant in Drosophila. Curr. Biol. 2009, 19, 1816–1820.
104. Hodl, M.; Basler, K. Transcription in the absence of histone H3.3. Curr. Biol. 2009, 19,
105. Loppin, B.; Bonnefoy, E.; Anselme, C.; Laurencon, A.; Karr, T.L.; Couble, P. The histone H3.3
chaperone HIRA is essential for chromatin assembly in the male pronucleus. Nature 2005, 437,
Int. J. Mol. Sci. 2011, 12
106. van der Heijden, G.W.; Dieker, J.W.; Derijck, A.A.; Muller, S.; Berden, J.H.; Braat, D.D.;
van der Vlag, J.; de Boer, P. Asymmetry in histone H3 variants and lysine methylation between
paternal and maternal chromatin of the early mouse zygote. Mech. Dev. 2005, 122, 1008–1022.
107. Bonnefoy, E.; Orsi, G.A.; Couble, P.; Loppin, B. The essential role of Drosophila HIRA for de
novo assembly of paternal chromatin at fertilization. PLoS Genet. 2007, 3, 1991–2006.
108. Konev, A.Y.; Tribus, M.; Park, S.Y.; Podhraski, V.; Lim, C.Y.; Emelyanov, A.V.; Vershilova, E.;
Pirrotta, V.; Kadonaga, J.T.; Lusser, A.; et al. CHD1 motor protein is required for deposition of
histone variant H3.3 into chromatin in vivo. Science 2007, 317, 1087–1090.
109. Gaspar-Maia, A.; Alajem, A.; Polesso, F.; Sridharan, R.; Mason, M.J.; Heidersbach, A.;
Ramalho-Santos, J.; McManus, M.T.; Plath, K.; Meshorer, E.; et al. Chd1 regulates open
chromatin and pluripotency of embryonic stem cells. Nature 2009, 460, 863–868.
110. Wong, L.H.; McGhie, J.D.; Sim, M.; Anderson, M.A.; Ahn, S.; Hannan, R.D.; George, A.J.;
Morgan, K.A.; Mann, J.R.; Choo, K.H. ATRX interacts with H3.3 in maintaining telomere
structural integrity in pluripotent embryonic stem cells. Genome Res. 2010, 20, 351–360.
111. Lewis, P.W.; Elsaesser, S.J.; Noh, K.M.; Stadler, S.C.; Allis, C.D. Daxx is an H3.3-specific
histone chaperone and cooperates with ATRX in replication-independent chromatin assembly at
telomeres. Proc. Natl. Acad. Sci. USA 2010, 107, 14075–14080.
112. Drane, P.; Ouararhni, K.; Depaux, A.; Shuaib, M.; Hamiche, A. The death-associated protein
DAXX is a novel histone chaperone involved in the replication-independent deposition of H3.3.
Genes Dev. 2010, 24, 1253–1265.
113. Torras-Llort, M.; Moreno-Moreno, O.; Azorin, F. Focus on the centre: the role of chromatin on
the regulation of centromere identity and function. EMBO J. 2009, 28, 2337–2348.
114. Dalal, Y.; Furuyama, T.; Vermaak, D.; Henikoff, S. Structure, dynamics, and evolution of
centromeric nucleosomes. Proc. Natl. Acad. Sci. USA 2007, 104, 15974–15981.
115. Sullivan, B.A.; Blower, M.D.; Karpen, G.H. Determining centromere identity: cyclical stories
and forking paths. Nat. Rev. Genet. 2001, 2, 584–596.
116. Allshire, R.C.; Karpen, G.H. Epigenetic regulation of centromeric chromatin: old dogs, new
tricks? Nat. Rev. Genet. 2008, 9, 923–937.
117. Gkikopoulos, T.; Singh, V.; Tsui, K.; Awad, S.; Renshaw, M.J.; Scholfield, P.; Barton, G.J.;
Nislow, C.; Tanaka, T.U.; Owen-Hughes, T. The SWI/SNF complex acts to constrain
distribution of the centromeric histone variant Cse4. EMBO J. 2011, 30, 1919–1927.
118. Dunleavy, E.M.; Roche, D.; Tagami, H.; Lacoste, N.; Ray-Gallet, D.; Nakamura, Y.; Daigo, Y.;
Nakatani, Y.; Almouzni-Pettinotti, G. HJURP is a cell-cycle-dependent maintenance and
deposition factor of CENP-A at centromeres. Cell 2009, 137, 485–497.
119. Foltz, D.R.; Jansen, L.E.; Bailey, A.O.; Yates, J.R., 3rd; Bassett, E.A.; Wood, S.; Black, B.E.;
Cleveland, D.W. Centromere-specific assembly of CENP-a nucleosomes is mediated by HJURP.
Cell 2009, 137, 472–484.
120. Williams, J.S.; Hayashi, T.; Yanagida, M.; Russell, P. Fission yeast Scm3 mediates stable
assembly of Cnp1/CENP-A into centromeric chromatin. Mol. Cell 2009, 33, 287–298.
121. Pidoux, A.L.; Choi, E.S.; Abbott, J.K.; Liu, X.; Kagansky, A.; Castillo, A.G.; Hamilton, G.L.;
Richardson, W.; Rappsilber, J.; He, X.; et al. Fission yeast Scm3: A CENP-A receptor required
for integrity of subkinetochore chromatin. Mol. Cell 2009, 33, 299–311.
Int. J. Mol. Sci. 2011, 12
122. Sanchez-Pulido, L.; Pidoux, A.L.; Ponting, C.P.; Allshire, R.C. Common ancestry of the
CENP-A chaperones Scm3 and HJURP. Cell 2009, 137, 1173–1174.
123. Perpelescu, M.; Nozaki, N.; Obuse, C.; Yang, H.; Yoda, K. Active establishment of centromeric
CENP-A chromatin by RSF complex. J. Cell Biol. 2009, 185, 397–407.
124. Okada, M.; Okawa, K.; Isobe, T.; Fukagawa, T. CENP-H-containing complex facilitates
centromere deposition of CENP-A in cooperation with FACT and CHD1. Mol. Biol. Cell 2009,
125. Walfridsson, J.; Bjerling, P.; Thalen, M.; Yoo, E.J.; Park, S.D.; Ekwall, K. The CHD remodeling
factor Hrp1 stimulates CENP-A loading to centromeres. Nucleic Acids Res. 2005, 33,
126. Podhraski, V.; Campo-Fernandez, B.; Worle, H.; Piatti, P.; Niederegger, H.; Bock, G.;
Fyodorov, D.V.; Lusser, A. CenH3/CID incorporation is not dependent on the chromatin
assembly factor CHD1 in Drosophila. PLoS One 2010, 5, e10120.
127. Hanai, K.; Furuhashi, H.; Yamamoto, T.; Akasaka, K.; Hirose, S. RSF governs silent chromatin
formation via histone H2Av replacement. PLoS Genet. 2008, 4, e1000011.
128. Erhardt, S.; Mellone, B.G.; Betts, C.M.; Zhang, W.; Karpen, G.H.; Straight, A.F. Genome-wide
analysis reveals a cell cycle-dependent mechanism controlling centromere propagation. J. Cell
Biol. 2008, 183, 805–818.
129. Liu, X.; Bowen, J.; Gorovsky, M.A. Either of the major H2A genes but not an evolutionarily
conserved H2A.F/Z variant of Tetrahymena thermophila can function as the sole H2A gene in
the yeast Saccharomyces cerevisiae. Mol. Cell. Biol. 1996, 16, 2878–2887.
130. Faast, R.; Thonglairoam, V.; Schulz, T.C.; Beall, J.; Wells, J.R.; Taylor, H.; Matthaei, K.;
Rathjen, P.D.; Tremethick, D.J.; Lyons, I. Histone variant H2A.Z is required for early
mammalian development. Curr. Biol. 2001, 11, 1183–1187.
131. van Daal, A.; Elgin, S.C. A histone variant, H2AvD, is essential in Drosophila melanogaster.
Mol. Biol. Cell 1992, 3, 593–602.
132. Suto, R.K.; Clarkson, M.J.; Tremethick, D.J.; Luger, K. Crystal structure of a nucleosome core
particle containing the variant histone H2A.Z. Nat. Struct. Biol. 2000, 7, 1121–1124.
133. Fan, J.Y.; Rangasamy, D.; Luger, K.; Tremethick, D.J. H2A.Z alters the nucleosome surface to
promote HP1alpha-mediated chromatin fiber folding. Mol. Cell 2004, 16, 655–661.
134. Zhang, Z.; Pugh, B.F. Genomic Organization of H2Av Containing Nucleosomes in Drosophila
Heterochromatin. PLoS One 2011, 6, e20511.
135. Hardy, S.; Jacques, P.E.; Gevry, N.; Forest, A.; Fortin, M.E.; Laflamme, L.; Gaudreau, L.;
Robert, F. The euchromatic and heterochromatic landscapes are shaped by antagonizing effects
of transcription on H2A.Z deposition. PLoS Genet. 2009, 5, e1000687.
136. Henikoff, S.; Henikoff, J.G.; Sakai, A.; Loeb, G.B.; Ahmad, K. Genome-wide profiling of salt
fractions maps physical properties of chromatin. Genome Res. 2009, 19, 460–469.
137. Mizuguchi, G.; Shen, X.; Landry, J.; Wu, W.H.; Sen, S.; Wu, C. ATP-driven exchange of histone
H2AZ variant catalyzed by SWR1 chromatin remodeling complex. Science 2004, 303, 343–348.
138. Kobor, M.S.; Venkatasubrahmanyam, S.; Meneghini, M.D.; Gin, J.W.; Jennings, J.L.; Link, A.J.;
Madhani, H.D.; Rine, J. A Protein Complex Containing the Conserved Swi2/Snf2-Related
ATPase Swr1p Deposits Histone Variant H2A.Z into Euchromatin. PLoS Biol. 2004, 2, e131.
Int. J. Mol. Sci. 2011, 12
139. Krogan, N.J.; Keogh, M.C.; Datta, N.; Sawa, C.; Ryan, O.W.; Ding, H.; Haw, R.A.; Pootoolal, J.;
Tong, A.; Canadien, V.; et al. A Snf2 family ATPase complex required for recruitment of the
histone H2A variant Htz1. Mol. Cell 2003, 12, 1565–1576.
140. Ruhl, D.D.; Jin, J.; Cai, Y.; Swanson, S.; Florens, L.; Washburn, M.P.; Conaway, R.C.;
Conaway, J.W.; Chrivia, J.C. Purification of a human SRCAP complex that remodels chromatin
by incorporating the histone variant H2A.Z into nucleosomes. Biochemistry (Mosc.) 2006, 45,
141. Luk, E.; Ranjan, A.; Fitzgerald, P.C.; Mizuguchi, G.; Huang, Y.; Wei, D.; Wu, C. Stepwise
histone replacement by SWR1 requires dual activation with histone H2A.Z and canonical
nucleosome. Cell 2010, 143, 725–736.
142. Wu, W.H.; Alami, S.; Luk, E.; Wu, C.H.; Sen, S.; Mizuguchi, G.; Wei, D.; Wu, C. Swc2 is a
widely conserved H2AZ-binding module essential for ATP-dependent histone exchange. Nat.
Struct. Mol. Biol. 2005, 12, 1064–1071.
143. Papamichos-Chronakis, M.; Watanabe, S.; Rando, O.J.; Peterson, C.L. Global regulation of
H2A.Z localization by the INO80 chromatin-remodeling enzyme is essential for genome
integrity. Cell 2011, 144, 200–213.
144. Park, Y.J.; Chodaparambil, J.V.; Bao, Y.; McBryant, S.J.; Luger, K. Nucleosome assembly
protein 1 exchanges histone H2A-H2B dimers and assists nucleosome sliding. J. Biol. Chem.
2005, 280, 1817–1825.
145. Luk, E.; Vu, N.D.; Patteson, K.; Mizuguchi, G.; Wu, W.H.; Ranjan, A.; Backus, J.; Sen, S.;
Lewis, M.; Bai, Y.; et al. Chz1, a nuclear chaperone for histone H2AZ. Mol. Cell 2007, 25,
146. Straube, K.; Blackwell, J.S., Jr.; Pemberton, L.F. Nap1 and Chz1 have separate Htz1 nuclear
import and assembly functions. Traffic 2010, 11, 185–197.
147. Reeves, R. Nuclear functions of the HMG proteins. Biochim. Biophys. Acta 2010, 1799, 3–14.
148. Bianchi, M.E.; Agresti, A. HMG proteins: dynamic players in gene regulation and
differentiation. Curr. Opin. Genet. Dev. 2005, 15, 496–506.
149. Rochman, M.; Malicet, C.; Bustin, M. HMGN5/NSBP1: a new member of the HMGN protein
family that affects chromatin structure and function. Biochim. Biophys. Acta 2010, 1799, 86–92.
150. Catez, F.; Yang, H.; Tracey, K.J.; Reeves, R.; Misteli, T.; Bustin, M. Network of dynamic
interactions between histone H1 and high-mobility-group proteins in chromatin. Mol. Cell. Biol.
2004, 24, 4321–4328.
151. Rochman, M.; Postnikov, Y.; Correll, S.; Malicet, C.; Wincovitch, S.; Karpova, T.S.;
McNally, J.G.; Wu, X.; Bubunenko, N.A.; Grigoryev, S.; et al. The interaction of NSBP1/HMGN5
with nucleosomes in euchromatin counteracts linker histone-mediated chromatin compaction and
modulates transcription. Mol. Cell 2009, 35, 642–656.
152. Phair, R.D.; Scaffidi, P.; Elbi, C.; Vecerova, J.; Dey, A.; Ozato, K.; Brown, D.T.; Hager, G.;
Bustin, M.; Misteli, T. Global nature of dynamic protein-chromatin interactions in vivo:
three-dimensional genome scanning and dynamic interaction networks of chromatin proteins.
Mol. Cell. Biol. 2004, 24, 6393–6402.
153. Rattner, B.P.; Yusufzai, T.; Kadonaga, J.T. HMGN proteins act in opposition to ATP-dependent
chromatin remodeling factors to restrict nucleosome mobility. Mol. Cell 2009, 34, 620–626.
Int. J. Mol. Sci. 2011, 12 Download full-text
154. Berube, N.G.; Smeenk, C.A.; Picketts, D.J. Cell cycle-dependent phosphorylation of the ATRX
protein correlates with changes in nuclear matrix and chromatin association. Hum. Mol. Genet.
2000, 9, 539–547.
155. Lechner, M.S.; Schultz, D.C.; Negorev, D.; Maul, G.G.; Rauscher, F.J., 3rd. The mammalian
heterochromatin protein 1 binds diverse nuclear proteins through a common motif that targets the
chromoshadow domain. Biochem. Biophys. Res. Commun. 2005, 331, 929–937.
156. Kourmouli, N.; Sun, Y.M.; van der Sar, S.; Singh, P.B.; Brown, J.P. Epigenetic regulation of
mammalian pericentric heterochromatin in vivo by HP1. Biochem. Biophys. Res. Commun. 2005,
157. Emelyanov, A.V.; Konev, A.Y.; Vershilova, E.; Fyodorov, D.V. Protein complex of Drosophila
ATRX/XNP and HP1a is required for the formation of pericentric beta-heterochromatin in vivo.
J. Biol. Chem. 2010, 285, 15027–15037.
158. Eskeland, R.; Eberharter, A.; Imhof, A. HP1 binding to chromatin methylated at H3K9 is
enhanced by auxiliary factors. Mol. Cell. Biol. 2007, 27, 453–465.
159. Hake, S.B.; Allis, C.D. Histone H3 variants and their potential role in indexing mammalian
genomes: the “H3 barcode hypothesis”. Proc. Natl. Acad. Sci. USA 2006, 103, 6428–6435.
160. Henikoff, S.; Furuyama, T.; Ahmad, K. Histone variants, nucleosome assembly and epigenetic
inheritance. Trends Genet. 2004, 20, 320–326.
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