In this essay, I propose that DNA‐binding anti‐cancer drugs work more via chromatin disruption than DNA damage. Success of long‐awaited drugs targeting cancer‐specific drivers is limited by the heterogeneity of tumors. Therefore, chemotherapy acting via universal targets (e.g., DNA) is still the mainstream treatment for cancer. Nevertheless, the problem with targeting DNA is insufficient efficacy due to high toxicity. I propose that this problem stems from the presumption that DNA damage is critical for the anti‐cancer activity of these drugs. (...) DNA in cells exists as chromatin, and many DNA‐targeting drugs alter chromatin structure by destabilizing nucleosomes and inducing histone eviction from chromatin. This effect has been largely ignored because DNA damage is seen as the major reason for anti‐cancer activity. I discuss how DNA‐binding molecules destabilize chromatin, why this effect is more toxic to tumoral than normal cells, and why cells die as a result of chromatin destabilization. (shrink)

Chromatin composition differs across the genome, with distinct compositions characterizing regions associated with different properties and functions. Whereas many histone modifications show local enrichment over genes or regulatory elements, marking can also span large genomic intervals defining broad chromatin domains. Here we highlight structural and functional features of chromatin domains marked by histone modifications, with a particular emphasis on the potential roles of H3K27 methylation domains in the organization and regulation of genome activity in metazoans. Chromatin (...) domains are extended genomic regions defined by the continuous enrichment of a given histone modification. Here, we review recent work highlighting the presence of chromatin domains in multiple species, their structural aspects in the context of chromatin architecture, and their potential role on the regulation of genome activity. (shrink)

The organization of the genome into topologically associated domains (TADs) appears to be a fundamental process occurring across a wide range of eukaryote organisms, and it likely plays an important role in providing an architectural foundation for gene regulation. Initial studies emphasized the remarkable parallels between TAD organization in organisms as diverse as Drosophila and mammals. However, whereas CCCTC‐binding factor (CTCF)/cohesin loop extrusion is emerging as a key mechanism for the formation of mammalian topological domains, the genome organization in Drosophila (...) appears to depend primarily on the partitioning of chromatin state domains. Recent work suggesting a fundamental conserved role of chromatin state in building domain architecture is discussed and insights into genome organization from recent studies in Drosophila are considered. (shrink)

Disruptions in chromatin regulator genes are frequently the cause of neurodevelopmental and neuropsychiatric disorders. Chromatin regulators are widely expressed in the brain, yet symptoms suggest that specific circuits can be preferentially altered when they are mutated. Using Drosophila allows targeted manipulation of chromatin regulators in defined neuronal classes, lineages, or circuits, revealing their roles in neuronal precursor self‐renewal, dendrite and axon targeting, neuron diversification, and the tuning of developmental signaling pathways. Phenotypes arising from chromatin regulator disruption (...) are context dependent – defined by interaction networks between the regulators, transcription factors, and chromatin remodeling complex partners. Future challenges are to determine the complexity of partner interactions, and to ascertain the degree to which cognitive deficits are due to loss of chromatin regulator activity in building a circuit or in maintaining homeostasis and activity within it. (shrink)

The process of chromatin diminution in Parascaris and Ascaris is a developmentally controlled genome rearrangement, which results in quantitative and qualitative differences in DNA content between germ line and somatic cells. Chromatin diminution involves chromosomal breakage, new telomere formation and DNA degradation. The programmed elimination of chromatin in presomatic cells might serve as an alternative way of gene regulation. We put forward a new hypothesis of how an ancient partial genome duplication and chromatin diminution may have (...) served to maintain the genetic balance in somatic cells and simultaneously endowed the germ line cells with a selective advantage. (shrink)

Eukaryotic genomes are packaged into a nucleoprotein complex known as chromatin. The term was introduced in 1879 by German cytologist Walther Flemming. While observing the processes of mitosis in a light microscope, Flemming coined the term to describe the easily stainable threads in the nucleus. He predicted that it would not have a long life: “The word chromatin may serve until its chemical nature is known, and meanwhile stands for that substance in the cell nucleus which is readily (...) stained”. However, Flemming’s prediction did not come true. Although the chemical nature of chromatin—a complex of nucleic acid and proteins—was already elucidated at the end of the 19th century, the.. (shrink)

Activation of gene transcription in vivo is accompanied by an alteration of chromatin structure. The specific binding of transcriptional activators disrupts nucleosomal arrays, suggesting that the primary steps leading to transcriptional initiation involve interactions between activators and chromatin. The affinity of transcription factors for nucleosomal DNA is determined by the location of recognition sequences within nucleosomes, and by the cooperative interactions of multiple proteins targeting binding sites contained within the same nucleosomes. In addition, two distinct types of enzymatic (...) complexes facilitate binding of transcription factors to nucleosomal DNA. These include type A histone acetyltransferases (e.g. GCN5/ADA transcriptional adaptor complex) and ATP‐driven molecular machines that disrupt histone‐DNA interactions (e.g. SWI/SNF and NURF complexes). These observations raise the important question of what happens to the histones during chromatin remodeling. We discuss evidence supporting the retention of histones at transcription factorbound sequences as well as two alternative pathways of histone loss from gene control elements upon transcription factor binding: histone octamer sliding and histone dissociation. (shrink)

Eukaryotic genome DNA is wrapped around core histones and forms a nucleosome structure. Together with associated proteins and RNAs, these nucleosomes are organized three‐dimensionally in the cell as chromatin. Emerging evidence demonstrates that chromatin consists of rather irregular and variable nucleosome arrangements without the regular fiber structure and that its dynamic behavior plays a critical role in regulating various genome functions. Single‐nucleosome imaging is a promising method to investigate chromatin behavior in living cells. It reveals local (...) class='Hi'>chromatin motion, which reflects chromatin organization not observed in chemically fixed cells. The motion data is like a gold mine. Data analyses from many aspects bring us more and more information that contributes to better understanding of genome functions. In this review article, we describe imaging of single‐nucleosomes and their tracked behavior through oblique illumination microscopy. We also discuss applications of this technique, especially in elucidating nucleolar organization in living cells. (shrink)

It is increasingly clear that chromatin is not just a device for packing DNA within the nucleus but also a dynamic material that changes as cellular environments alter. The precise control of chromatin modification in response to developmental and environmental cues determines the correct spatial and temporal expression of genes. Here, we review exciting discoveries that reveal chromatin participation in many facets of plant development. These include: chromatin modification from embryonic and meristematic development to flowering and (...) seed formation, the involvement of DNA methylation and chromatin in controlling invasive DNA and in maintenance of epigenetic states, and the function of chromatin modifying and remodeling complexes such as SWI/SNF and histone acetylases and deacetylases in gene control. Given the role chromatin structure plays in every facet of plant development, chromatin research will undoubtedly be integral in both basic and applied plant biology. BioEssays 24:234–243, 2002. © 2002 Wiley Periodicals, Inc.; DOI 10.1002/bies.10055. (shrink)

The assembly of nucleosomes and higher‐order chromatin structures has been extensively studied in vitro. Provided that non‐specific charge interactions are controlled, all the information for correct assembly is found to be inherent in the macromolecular components. Cellular extracts which can assemble chromatin in vitro with nucleosomes correctly spaced on the DNA have been studied in detail and also used to investigate the role of chromatin structure in transcription. However, the mechanisms of chromatin assembly in vivo are (...) still controversial. (shrink)

We present a molecular model of eukaryotic gene transcription. For the β‐globin locus, we hypothesise that a transcription machine composed of multiple RNA polymerase II (PolII) assembles using the locus control region as a foundation. Transcription and locus remodelling can be achieved by pulling DNA through this multi‐PolII ‘reading head’. Once a transcription complex is formed, it may engage an active gene in several rounds of transcription. Observed intergenic sense and antisense transcripts may be the result of PolII pulling the (...) DNA through the reading head whilst searching for the promoter of a gene. Support for this hypothesis is provided using various data from the literature. In the model, DNA is packed in a 30‐nm chromatin fibre, thus gene regulatory regions separated by kilobases are close in space. This, and the need to store transcription‐induced supercoiling, may explain why functionally interacting regions are often separated by many kilobases. (shrink)

One facet of the control of gene expression is long‐range promoter regulation by distant enhancers. It is an important component of the regulation of genes that control metazoan development and has been appreciated for some time but the molecular mechanisms underlying this regulation have remained poorly understood. A recent study by Cleard and colleagues1 reports the first in vivo evidence of chromatin looping and boundary element promoter interaction. Specifically, they studied the function of a boundary element within the cis‐regulatory (...) region of the Abdominal‐B (Abd‐B) gene of Drosophila melanogaster. BioEssays 29: 7–10, 2007. © 2006 Wiley Periodicals, Inc. (shrink)

It has long been assumed that the architectural proteins of chromatin (the histones, for example) are unrelated to their functional proteins (transcription factors, polymerases, etc). New studies(1,2)drastically change this perspective. It appears that a portion of the general transcription initiation complex TFIID is made up of proteins that not only carry marked sequence and structural resemblances to the core histones of the nucleosome, but also form an octameric complex similar to the histone octamer. This can now be seen as (...) part of a general pattern of continuity among structural and functional chromatin proteins. (shrink)

Graphical AbstractThe loosening of chromatin structures gives rise to unrestricted access to DNA and thus transcription factors (TFs) can bind to their otherwise masked target sequences. Regions bound by the same set of TFs tend to be located in close proximity and this might increase the probability of activating illegitimate genomic rearrangements.

Specificities associated with chromosomal linearity are not restricted to telomeres. Here, recent results obtained on fission and budding yeast are summarized and an attempt is made to define subtelomeres using chromatin features extending beyond the heterochromatin emanating from telomeres. Subtelomeres, the chromosome domains adjacent to telomeres, differ from the rest of the genome by their gene content, rapid evolution, and chromatin features that together contribute to organism adaptation. However, current definitions of subtelomeres are generally based on synteny and (...) are largely gene‐centered. Taking into consideration both the peculiar gene content and dynamics as well as the chromatin properties of those domains, it is discussed how chromatin features can contribute to subtelomeric properties and functions, and play a pivotal role in the emergence of subtelomeres. (shrink)

Eukaryotic genomes are packaged into nucleosomal chromatin, and genomic activity requires the precise localization of transcription factors, histone modifications and nucleosomes. Classic work described the progressive reassembly and maturation of bulk chromatin behind replication forks. More recent proteomics has detailed the molecular machines that accompany the replicative polymerase to promote rapid histone deposition onto the newly replicated DNA. However, localized chromatin features are transiently obliterated by DNA replication every S phase of the cell cycle. Genomic strategies now (...) observe the rebuilding of locus‐specific chromatin features, and reveal surprising delays in transcription factor binding behind replication forks. This implies that transient chromatin disorganization during replication is a central juncture for targeted transcription factor binding within genomes. We propose that transient occlusion of regulatory elements by disorganized nucleosomes during chromatin maturation enforces specificity of factor binding. (shrink)

The process by which the genetically identical cell lineages of a multicellular organism acquire the propensity to express distinct arrays of gene products is among the most significant and fascinating questions in modern biology. Not surprisingly, this complex process requires control at several levels, each level providing a condition that is necessary but not sufficient for transcription to occur. Evidence suggests that one level of control concerns a region of DNA much larger than the transcription unit itself – the (...) class='Hi'>chromatin domain. This domain must be in a specific chromatin conformation in order to permit transcription; other control mechanisms may be required to bring about overt transcription. A hypothesis concerning the nature of chromatin domains and the relationship between early replication and transcription potential is presented. (shrink)

The eukaryotic genome is packaged into a periodic nucleoprotein structure termed chromatin. The repeating unit of chromatin, the nucleosome, consists of DNA that is wound nearly two times around an octamer of histone proteins. To facilitate DNA‐directed processes in chromatin, it is often necessary to rearrange or to mobilize the nucleosomes. This remodeling of the nucleosomes is achieved by the action of chromatin‐remodeling complexes, which are a family of ATP‐dependent molecular machines. Chromatin‐remodeling factors share a (...) related ATPase subunit and participate in transcriptional regulation, DNA repair, homologous recombination and chromatin assembly. In this review, we provide an overview of chromatin‐remodeling enzymes and discuss two possible mechanisms by which these factors might act to reorganize nucleosome structure. BioEssays 25:1192–1200, 2003. © 2003 Wiley Periodicals, Inc. (shrink)

Just as the faithful replication of DNA is an essential process for the cell, chromatin structures of active and inactive genes have to be copied accurately. Under certain circumstances, however, the activity pattern has to be changed in specific ways. Although analysis of specific aspects of these complex processes, by means of model systems, has led to their further elucidation, the mechanisms of chromatin replication in vivo are still controversial and far from being understood completely. Progress has been (...) achieved in understanding: 1. The initiation of chromatin replication, indicating that a nucleosome‐free origin is necessary for the initiation of replication; 2. The segregation of the parental nucleosomes, where convincing data support the model of random distribution of the parental nucleosomes to the daughter strands; and 3. The assembly of histones on the newly synthesized strands, where growing evidence is emerging for a two‐step mechanism of nucleosome assembly, starting with the deposition of H3/H4 tetramers onto the DNA, followed by H2A/H2B dimers. (shrink)

Recent investigations have revealed 1) that the isochores of the human genome group into two super‐families characterized by two different long‐range 3D structures, and 2) that these structures, essentially based on the distribution and topology of short sequences, mold primary chromatin domains (and define nucleosome binding). More specifically, GC‐poor, gene‐poor isochores are low‐heterogeneity sequences with oligo‐A spikes that mold the lamina‐associated domains (LADs), whereas GC‐rich, gene‐rich isochores are characterized by single or multiple GC peaks that mold the topologically associating (...) domains (TADs). The formation of these “primary TADs” may be followed by extrusion under the action of cohesin and CTCF. Finally, the genomic code, which is responsible for the pervasive encoding and molding of primary chromatin domains (LADs and primary TADs, namely the “gene spaces”/“spatial compartments”) resolves the longstanding problems of “non‐coding DNA,” “junk DNA,” and “selfish DNA” leading to a new vision of the genome as shaped by DNA sequences. (shrink)

t is now well accepted that defined architectural compartments within the cell nucleus can regulate the transcriptional activity of chromosomal domains within their vicinity. However, it is generally unclear how these compartments are formed. The nuclear periphery has received a great deal of attention as a repressive compartment that is implicated in many cellular functions during development and disease. The inner nuclear membrane, the nuclear lamina, and associated proteins compose the nuclear periphery and together they interact with proximal chromatin (...) creating a repressive environment. A new study by Harr et al. identifies specific protein–DNA interactions and epigenetic states necessary to re‐position chromatin to the nuclear periphery in a cell‐type specific manner. Here, we review concepts in gene positioning within the nucleus and current accepted models of dynamic gene repositioning within the nucleus during differentiation. This study highlights that myriad pathways lead to nuclear organization. (shrink)

The eukaryotic genome is organized into different domains by cis‐acting elements, such as boundaries/insulators and matrix attachment regions, and is packaged with different degrees of condensation. In the M phase, the chromatin becomes further highly condensed into chromosomes. The first step for transcriptional activation of a given gene, at a particular time during development, in any locus, is the opening of its chromatin domain. This locus needs to be kept in this state in each early G1 phase during (...) every cell cycle. Certain distal enhance elements, including locus control regions (LCRs) and enhancers, are believed to perform this target chromatin domain opening process and several models have been proposed to explain distal enhance action. But they did not explain precisely how a given chromatin domain is opened. Based on various studies, we propose a hypothesis for the mechanism of opening chromatin on a large scale. One important mechanism may involved breaking one or two DNA strands and reducing the linking numbers within chromatin domain. The topological changes can overpass some complexes formed on DNA strands and can be transmitted from specific localized points over a broad region, until boundary elements or insulators are reached. These may initiate downstream events such as propagation of histone acetylation and the binding of transcription factors to proximal promoters and may further augment the action mediated by distal enhancer elements. BioEssays 25:507–514, 2003. © 2003 Wiley Periodicals, Inc. (shrink)

How epigenetic mechanisms regulate genome output and response to stimuli is a fundamental question in development and disease. Past decades have made tremendous progress in deciphering the regulatory relationships involved by correlating aggregated (epi)genomics profiles with global perturbations. However, the recent development of epigenetic editing technologies now enables researchers to move beyond inferred conclusions, towards explicit causal reasoning, through 'programing’ precise chromatin perturbations in single cells. Here, we first discuss the major unresolved questions in the epigenetics field that can (...) be addressed by programable epigenome editing, including the context‐dependent function and memory of chromatin states. We then describe the epigenetic editing toolkit focusing on CRISPR‐based technologies, and highlight its achievements, drawbacks and promise. Finally, we consider the potential future application of epigenetic editing to the study and treatment of specific disease conditions. (shrink)

Variegated phenotypes often result from chromosomal rearrangements that place euchromatic genes next to heterochromatin. In such rearrangements, the condensed structure of heterochromatin can spread into euchromatic regions, which then assume the morphology of heterochromatin and become transcriptionally inactive. In position‐effect variegation (PEV) therefore, gene inactivation results from a change in chromatin structure. PEV has been intensively investigated in the fruitfly Drosophila, where the phenomenon allows a genetic dissection of chromatin components. Consequently, many genes have been identified which, when (...) mutated, act as dominant modifiers (suppressors or enhancers) of PEV. Data available already demonstrate that genetic, molecular and developmental analysis of these genes provides an avenue to the identification of regulatory and structural chromatin components, and hence to fundamental aspects of chromosome structure and function. (shrink)

Two aspects of the chromatin repeat length (r t) are discussed: (i) Why is r t, longer for slowly dividing cells than in rapidly dividing cells?, and (ii) Why is the temporal evolution of r ta decreasing function of time (t) in mammalian cortical neurons, whereas it is an increasing function of t for granule cells around the time of birth? These questions are discussed in terms of a hypothesis which assumes a correlation between deoxyribonucleic acid (DNA) packaging, transcription, (...) and replication. (shrink)

Chromosomal rearrangements frequently occur at specific places (“hot spots”) in the genome. These recombination hot spots are usually separated by 50–100 kb regions of DNA that are rarely involved in rearrangements. It is quite likely that there is a correlation between the above‐mentioned distances and the average size of DNA loops fixed at the nuclear matrix. Recent studies have demonstrated that DNA loop anchorage regions can be fairly long and can harbor DNA recombination hot spots. We previously proposed that chromosomal (...) DNA loops may constitute the basic units of genome organization in higher eukaryotes. In this review, we consider recombination between DNA loop anchorage regions as a possible source of genome evolution. (shrink)

DNA is packed as chromatin on several levels in the eukaryotic nucleus. Dissection of chromatin with nucleases produces three stable substructures: the nucleosome core particle, the chromatosome and the 30 nm fibre. Whilst the first two allow transcription, the 30 nm fibre is taken to be the first level of transcriptionally dormant chromatin and it has an important functional role in cell differentiation and epigenetic regulation. Its structure has been a subject of continuing discussion since native fibres (...) cannot readily be crystallized. This problem has recently been addressed by reconstitution of fibres on repeats of DNA sequences having nucleosome‐positioning properties and two different structures were reported.1,2The reconstitution results and their interpretations are compared in this review with experimental data from native chromatin and it is shown that the results of Robinson et al.2 conform well with the known structural features of native fibres and are a good first step towards understanding the structure of the fibre. BioEssays 30:1003–1009, 2008. © 2008 Wiley Periodicals, Inc. (shrink)

The use of Drosophila chromosomal rearrangements and transposon constructs involving the white gene reveals the existence of repressive chromatin domains that can spread over considerable genomic distances. One such type of domain is found in heterochromatin and is responsible for classical position‐effect variegation. Another type of repressive domain is established, beginning at specific sequences, by complexes of Polycomb Group proteins. Such complexes, which normally regulate the expression of many genes, including the homeotic loci, are responsible for silencing, white gene (...) variegation, pairing‐dependent effects and insertional targeting. (shrink)

We highlight a recent study exploring the hand‐off of UV damage to several key nucleotide excision repair (NER) proteins in the cascade: UV‐DDB, XPC and TFIIH. The delicate dance of DNA repair proteins is choreographed by the dynamic hand‐off of DNA damage from one recognition complex to another damage verification protein or set of proteins. These DNA transactions on chromatin are strictly chaperoned by post‐translational modifications (PTM). This new study examines the role that ubiquitylation and subsequent DDB2 degradation has (...) during this process. In total, this study suggests an intricate cellular timer mechanism that under normal conditions DDB2 helps recruit and ubiquitylate XPC, stabilizing XPC at damaged sites. If DDB2 persists at damaged sites too long, it is turned over by auto‐ubiquitylation and removed from DNA by the action of VCP/p97 for degradation in the 26S proteosome. (shrink)

Phenotypic plasticity is a crucial feature of aggressive cancer, providing the means for cancer progression. Stochastic changes in tumor cell transcriptional programs increase the chances of survival under any condition. I hypothesize that unstable chromatin permits stochastic transitions between transcriptional programs in aggressive cancers and supports non‐genetic heterogeneity of tumor cells as a basis for their adaptability. I present a mechanistic model for unstable chromatin which includes destabilized nucleosomes, mobile chromatin fibers and random enhancer‐promoter contacts, resulting in (...) stochastic transcription. I suggest potential markers for “unsettled” chromatin in tumors associated with poor prognosis. Although many of the characteristics of unstable chromatin have been described, they were mostly used to explain changes in the transcription of individual genes. I discuss approaches to evaluate the role of unstable chromatin in non‐genetic tumor cell heterogeneity and suggest using the degree of chromatin instability and transcriptional noise in tumor cells to predict cancer aggressiveness. (shrink)

From 1956, when the complex ultrastructure of meiotic chromosomes was discovered, 1 until 1985, when the isolation of meiotic chromosome cores was reported, knowledge of the molecular structure of the meiotic chromosome was at best a dream. The dissection of meiotic chromosome structures has become a realistic challenge through the arrival of isolated symptonemal complexes (SCs), monoclonal and polyclonal antibodies against SCs, the possibility for screening expression libraries for genes that encode SC proteins, the isolation of SC‐associated DNA, and the (...) development of techniques for the in situ recognition of DNA sequences in the context of the meiotic chromosome structure. (shrink)

Here, the emerging data on DNA G‐quadruplexes (G4s) as epigenetic modulators are reviewed and integrated. This concept has appeared and evolved substantially in recent years. First, persistent G4s (e.g., those stabilized by exogenous ligands) were linked to the loss of the histone code. More recently, transient G4s (i.e., those formed upon replication or transcription and unfolded rapidly by helicases) were implicated in CpG island methylation maintenance and de novo CpG methylation control. The most recent data indicate that there are direct (...) interactions between G4s and chromatin remodeling factors. Finally, multiple findings support the indirect participation of G4s in chromatin reshaping via interactions with remodeling‐related transcription factors (TFs) or damage responders. Here, the links between the above processes are analyzed; also, how further elucidation of these processes may stimulate the progress of epigenetic therapy is discussed, and the remaining open questions are highlighted. (shrink)

During early embryonic development in several metazoans, accurate DNA replication is ensured by high number of replication origins. This guarantees rapid genome duplication coordinated with fast cell divisions. In Xenopus laevis embryos this program switches to one with a lower number of origins at a developmental stage known as mid‐blastula transition (MBT) when cell cycle length increases and gene transcription starts. Consistent with this regulation, somatic nuclei replicate poorly when transferred to eggs, suggesting the existence of an epigenetic memory suppressing (...) replication assembly origins at all available sites. Recently, it was shown that histone H1 imposes a non‐permissive chromatin configuration preventing replication origin assembly on somatic nuclei. This somatic state can be erased by SSRP1, a subunit of the FACT complex. Here, we further develop the hypothesis that this novel form of epigenetic memory might impact on different areas of vertebrate biology going from nuclear reprogramming to cancer development. (shrink)

Extensive studies of DNA secondary structure during the past decade have shown that DNA is a dynamic molecule, whose structure depends on the underlying nucleotide sequence and is influenced by the environment and the overall DNA topology. Three major non‐B‐DNA structures have been described (Z‐DNA, triplex DNA and cruciform DNA) which are stabilized by unconstrained negative supercoiling and can be formed under physiological conditions. In this essay we summarize the DNA primary structure features that are pertinent to the formation of (...) these conformers and present data concerning the occurrence of these sequences in the eukaryotic genome. The evidence in favor of the existence of these unusual DNA structures in vivo is discussed. The effect of alternative non‐B‐DNA structures on the way DNA is organized in chromatin is considered, and this is followed by evaluation of the data relating these structures to eukaryotic transcription. Some possible mechanisms by which the effect of non‐B structures on transcription might be exerted are proposed. (shrink)

The role of RNA as a messenger in the expression of the genome has been long appreciated, but its functions in regulating chromatin and chromosome structure are no less interesting. Recent results have shown that small RNAs guide chromatin‐modifying complexes to chromosomal regions in a sequence‐specific manner to elicit transcriptional repression. However, sequence‐specific targeting by means of base pairing seems to be only one mechanism by which RNA is employed for epigenetic regulation. The focus of this review is (...) on large RNAs that act in the dosage‐compensation pathways of flies and mammals. These RNAs associate with chromatin over the length of whole chromosomes and are crucial for spreading epigenetic changes in chromatin structure. They do not appear to act in a sequence‐specific manner but might provide scaffolds for co‐operative binding of chromatin‐associated complexes that enable spreading of chromatin modifications. BioEssays 25:434–442, 2003. © 2003 Wiley Periodicals, Inc. (shrink)

How chromatin bridges are detected by the abscission checkpoint during mammalian cell division is unknown. Here, we discuss recent findings from our lab showing that the DNA topoisomerase IIα (Top2α) enzyme binds to catenated (“knotted”) DNA next to the midbody and forms abortive Top2‐DNA cleavage complexes (Top2ccs) on chromatin bridges. Top2ccs are then processed by the proteasome to promote localization of the DNA damage sensor protein Rad17 to Top2‐generated double‐strand DNA ends on DNA knots. In turn, Rad17 promotes (...) local recruitment of the MRN protein complex and downstream ATM‐Chk2‐INCENP signaling to delay abscission and prevent chromatin bridge breakage in cytokinesis. (shrink)

How fast? How strong? How many? So what? Why do numbers matter in biology? Chromatin binding proteins are forever in motion, exchanging rapidly between bound and free pools. How do regulatory systems whose components are in constant flux ensure stability and flexibility? This review explores the application of quantitative and mathematical approaches to mechanisms of epigenetic regulation. We discuss methods for measuring kinetic parameters and protein quantities in living cells, and explore the insights that have been gained by quantifying (...) and modelling dynamics of chromatin binding proteins. (shrink)

Fluorescence microscopy has provided a route to qualitatively analyze features of nuclear structures and chromatin domains with increasing resolution. However, it is becoming increasingly important to develop tools for quantitative analysis. Here, we present an automated method to quantitatively determine the enrichment of several endogenous factors, immunostained in pericentric heterochromatin domains in mouse cells. We show that this method permits an unbiased characterization of changes in the enrichment of several factors with statistical significance from a large number of nuclei. (...) Furthermore, the nuclei can be sorted according to the enrichment value of these factors. This method should prove useful to monitor events related to changes in the amount, rather than the presence or absence, of any factor. By adapting a few parameters, it could be extended to other nuclear structures and the benefit of using available software will permit its use in many biological labs. (shrink)

Although the promyelocytic leukemia (PML) protein is renowned for regulating a wide range of cellular processes and as an essential component of PML nuclear bodies (PML‐NBs), the mechanisms through which it exerts its broad physiological impact are far from fully elucidated. Here, we review recent studies supporting an emerging view that PML's pleiotropic effects derive, at least partially, from its role in regulating histone H3.3 chromatin assembly, a critical epigenetic mechanism. These studies suggest that PML maintains heterochromatin organization by (...) restraining H3.3 incorporation. Examination of PML's contribution to H3.3 chromatin assembly in the context of the cell cycle and PML‐NB assembly suggests that PML represses heterochromatic H3.3 deposition during S phase and that transcription and SUMOylation regulate PML's recruitment to heterochromatin. Elucidating PML’ s contributions to H3.3‐mediated epigenetic regulation will provide insight into PML's expansive influence on cellular physiology and open new avenues for studying oncogenesis linked to PML malfunction. (shrink)

Recently, there has been a convergence of fields studying the processing of DNA, such as transcription, replication, and repair. This convergence has been centered around the packaging of DNA in chromatin. Chromatin structure affects all aspects of DNA processing because it modulates access of proteins to DNA. Therefore, a central theme has become the mechanism(s) for accessing DNA in chromatin. It seems likely that mechanisms involved in one of these processes may also be used in others. For (...) example, the discovery of transcriptional coactivators with histone acetyltransferase activity and chromatin remodeling complexes has provided possible mechanisms required for efficient repair of DNA in chromatin. BioEssays 21:596–603, 1999. © 1999 John Wiley & Sons, Inc. (shrink)

In this review, we discuss a novel on‐site remodeling function that is mediated by the H2A‐ubiquitin binding protein ZRF1. ZRF1 facilitates the remodeling of multiprotein complexes at chromatin and lies at the heart of signaling processes that occur at DNA damage sites and during transcriptional activation. In nucleotide excision repair ZRF1 remodels E3 ubiquitin ligase complexes at the damage site. During embryonic stem cell differentiation, it contributes to retinoic acid‐mediated gene activation by altering the subunit composition of the Mediator (...) complex. We postulate that ZRF1 operates in conjunction with cellular remodeling machines and suggest that on‐site remodeling might be a hallmark of many chromatin‐associated signaling pathways. We discuss yet unexplored functions of ZRF1‐mediated remodeling in replication and double strand break repair. In conclusion, we postulate that on‐site remodeling of multiprotein complexes is essential for the timing of chromatin signaling processes. (shrink)

Bone‐marrow mesenchymal stem cell (BM‐MSC) proliferation and lineage commitment are under the coordinated control of metabolism and epigenetics; the MSC niche contains low oxygen, which is an important determinant of the cellular metabolic state. In turn, metabolism drives stem cell fate decisions via alterations of the chromatin landscape. Due to the fundamental role of BM‐MSCs in the development of adipose tissue, bones and cartilage, age‐associated changes in metabolism and the epigenome perturb the balance between stem cell proliferation and differentiation (...) leading to stem cell depletion, fat accumulation and bone‐quality related diseases. Therefore, understanding the dynamics of the metabolism‐chromatin interplay is crucial for maintaining the stem cell pool and delaying the development and progression of ageing. This review summarizes the current knowledge on the role of metabolism in stem cell identity and highlights the impact of the metabolic inputs on the epigenome, with regards to stemness and pluripotency. (shrink)

The bulk of the DNA in eukaryotic chromatin behaves as if it is topologically relaxed; however, a subfraction can be shown to be under suercoil tension. Endonuclease S1 cuts at specific hypersentive sites in chromatin (in the promoter regions of active genes) and this enzyme cuts in the same region in supercoiled plasmids, but not in relaxed or linearized molecules. A subfraction of the minichromosomes formed after SV40 infection or microinjection of plasmid DNA into oocytes contains supercoil tension (...) and this subfraction is thought to play an important role in transcription. (shrink)

The “Encyclopedia of DNA Elements” (ENCODE) project was launched by the US National Human Genome Research Institute in the aftermath of the Human Genome Project (HGP). It aimed to systematically map the human transcriptome, and held the promise that identifying potential regulatory regions and transcription factor binding sites would help address some of the perplexing results of the HGP. Its initial results published in 2012 produced a flurry of high-impact publications as well as criticisms. Here we put the results of (...) ENCODE and the work on epigenomics that followed in a broad theoretical and historical context, focusing on three strands of research. The first is the history of thinking about the organization of genomes, both physical and regulatory. The second is the history of ideas about gene regulation, primarily in eukaryotes. Finally, and connecting these two issues, we suggest how to think about the role of genetic material in physiology and development. (shrink)

Polycomb group (PcG) proteins maintain the expression state of PcG‐responsive genes during development of multicellular organisms. Recent observations suggest that “the H3K27me3 modification” acts to maintain Polycomb repressive complex (PRC) 2, the enzyme that creates this modification, on replicating chromatin. This could in turn promote propagation of H3K27me3 on newly replicated daughter chromatin, and promote recruitment of PRC1. Other work suggests that PRC1‐class complexes can be maintained on replicating chromatin, at least in vitro, independently of H3K27me3. Thus, (...) histone modifications and PcG proteins themselves may both be maintained through replication. (shrink)

Editor's suggested further reading in BioEssays Epigenetics meets mathematics: Towards a quantitative understanding of chromatin biology Abstract.