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BBA - Gene Regulatory Mechanisms (v.1819, #7)

Editorial Board (pp. i).

Chromatin in time and space by Gordon L. Hager; Lyuba Varticovski (pp. 631-631).

The importance of being supercoiled: How DNA mechanics regulate dynamic processes by Laura Baranello; David Levens; Ashutosh Gupta; Fedor Kouzine (pp. 632-638).

Through dynamic changes in structure resulting from DNA–protein interactions and constraints given by the structural features of the double helix, chromatin accommodates and regulates different DNA-dependent processes. All DNA transactions (such as transcription, DNA replication and chromosomal segregation) are necessarily linked to strong changes in the topological state of the double helix known as torsional stress or supercoiling. As virtually all DNA transactions are in turn affected by the torsional state of DNA, these changes have the potential to serve as regulatory signals detected by protein partners. This two-way relationship indicates that DNA dynamics may contribute to the regulation of many events occurring during cell life. In this review we will focus on the role of DNA supercoiling in the cellular processes, with particular emphasis on transcription. Besides giving an overview on the multiplicity of factors involved in the generation and dissipation of DNA torsional stress, we will discuss recent studies which give new insight into the way cells use DNA dynamics to perform functions otherwise not achievable. This article is part of a Special Issue entitled: Chromatin in time and space.► It has been speculated that DNA topology might play a regulatory role in chromosome biology. ► The review focuses on the most recent approaches developed to evaluate this possibility. ► Multiple factors contribute to dynamic changes in DNA topology. ► These changes are involved in the moment-to-moment guidance of crucial processes. ► This real-time regulation is necessary for rapid cellular responses to physiological stimuli.

Keywords: DNA supercoiling; DNA topology; Non-B DNA; Transcription

Perfect and imperfect nucleosome positioning in yeast by Hope A. Cole; V. Nagarajavel; David J. Clark (pp. 639-643).

Numerous studies of nucleosome positioning have shown that nucleosomes almost invariably adopt one of several alternative overlapping positions on a short DNA fragment in vitro. We define such a set of overlapping positions as a “position cluster”, and the 5S RNA gene positioning sequence is presented as an example. The notable exception is the synthetic 601-sequence, which can position a nucleosome perfectly in vitro, though not in vivo. Many years ago, we demonstrated that nucleosome position clusters are present on the CUP1 and HIS3 genes in native yeast chromatin. Recently, using genome-wide paired-end sequencing of nucleosomes, we have shown that position clusters are the general rule in yeast chromatin, not the exception. We argue that, within a cell population, one of several alternative nucleosomal arrays is formed on each gene. We show how position clusters and alternative arrays can give rise to typical nucleosome occupancy profiles, and that position clusters are disrupted by transcriptional activation. The centromeric nucleosome is a rare example of perfect positioning in vivo. It is, however, a special case, since it contains the centromeric histone H3 variant instead of normal H3. Perfect positioning might be due to centromeric sequence-specific DNA binding proteins. Finally, we point out that the existence of position clusters implies that the putative nucleosome code is degenerate. We suggest that degeneracy might be a crucial point in the debate concerning the code. This article is part of a Special Issue entitled: Chromatin in time and space.► A canonical nucleosome chooses one of several alternative overlapping positions. ► Such “position clusters” are observed both in vitro and in vivo, genome-wide. ► Nucleosome position clusters are disrupted on transcriptionally active genes. ► Perfect positioning is rare. The centromeric nucleosome is an example. ► The nucleosome code is degenerate, specifying alternative nucleosomal arrays.

Keywords: S. cerevisiae; Nucleosome; Chromatin remodelling; Nucleosome position; Nucleosome occupancy; Nucleosome code

Chromatin domains, insulators, and the regulation of gene expression by Rodolfo Ghirlando; Keith Giles; Humaira Gowher; Tiaojiang Xiao; Zhixiong Xu; Hongjie Yao; Gary Felsenfeld (pp. 644-651).

The DNA sequence elements called insulators have two basic kinds of properties. Barrier elements block the propagation of heterochromatic structures into adjacent euchromatin. Enhancer blocking elements interfere with interaction between an enhancer and promoter when placed between them. We have dissected a compound insulator element found at the 5′ end of the chicken β-globin locus, which possesses both activities. Barrier insulation is mediated by two kinds of DNA binding proteins: USF1/USF2, a heterodimer which recruits multiple enzyme complexes capable of marking histone on adjacent nucleosomes with ‘activating’ marks, and Vezf1, which protects against DNA methylation. We have found that the heterochromatic region upstream of the insulator element is maintained in its silent state by a dicer-dependent mechanism, suggesting a mechanism for Vezf1 function in the insulator.Enhancer blocking function in the β-globin insulator element is conferred by a binding site for CTCF. Consistent with this property, CTCF binding was found some years ago to be essential for imprinted expression at the Igf2/H19 locus. Work in many laboratories has since demonstrated that CTCF helps stabilize long-range interactions in the nucleus. We have recently shown that in the case of the human insulin locus such an interaction, over a distance of ~300kb, can result in stimulation of a target gene which itself is important for insulin secretion. This article is part of a Special Issue entitled: Chromatin in time and space.► Heterochromatin is blocked by elements that maintain positive histone modifications. ► Heterochromatin formation in vertebrates can involve dicer dependent mechanisms. ► CTCF mediated stabilization of long range contacts affects gene expression. ► The human insulin gene regulates the distant SYT8 gene through physical contact.

Keywords: CTCF; Vezf1; Insulation; Heterochromatin

The HMGN family of chromatin-binding proteins: Dynamic modulators of epigenetic processes by Jamie E. Kugler; Tao Deng; Michael Bustin (pp. 652-656).

The HMGN family of proteins binds to nucleosomes without any specificity for the underlying DNA sequence. They affect the global and local structure of chromatin, as well as the levels of histone modifications and thus play a role in epigenetic regulation of gene expression. This review focuses on the recent studies that provide new insights on the interactions between HMGN proteins, nucleosomes, and chromatin, and the effects of these interactions on epigenetic and transcriptional regulation. This article is part of a Special Issue entitled: Chromatin in time and space.► HMGN proteins bind to nucleosomes and affect epigenetic processes. ► The mechanism of HMGN binding to the core particle has recently been elucidated. ► HMGN proteins affect global and local chromatin organization. ► Each HMGN variant affects transcription of a separate set of genes.

Keywords: HMGN protein; Epigenetics; Transcription

Complex dynamics of transcription regulation by Diana A. Stavreva; Lyuba Varticovski; Gordon L. Hager (pp. 657-666).

Transcription is a tightly regulated cellular function which can be triggered by endogenous (intrinsic) or exogenous (extrinsic) signals. The development of novel techniques to examine the dynamic behavior of transcription factors and the analysis of transcriptional activity at the single cell level with increased temporal resolution has revealed unexpected elements of stochasticity and dynamics of this process. Emerging research reveals a complex picture, wherein a wide range of time scales and temporal transcription patterns overlap to generate transcriptional programs. The challenge now is to develop a perspective that can guide us to common underlying mechanisms, and consolidate these findings. Here we review the recent literature on temporal dynamics and stochastic gene regulation patterns governed by intrinsic or extrinsic signals, utilizing the glucocorticoid receptor (GR)-mediated transcriptional model to illustrate commonality of these emerging concepts. This article is part of a Special Issue entitled: Chromatin in time and space.► Transcription is a dynamic and stochastic, nevertheless tightly regulated cellular function. ► Intrinsic and extrinsic signals generate diverse transcriptional responses and affect physiology. ► Regulatory mechanisms driving specific transcriptional programs remain to be discovered.

Keywords: Dynamics; Ultradian; Oscillation; Gene pulsing; Intrinsic and extrinsic signals; Transcription bursting

Complexity of RNA polymerase II elongation dynamics by Murali Palangat; Daniel R. Larson (pp. 667-672).

Transcription of protein-coding genes by RNA polymerase II can be regulated at multiple points during the process of RNA synthesis, including initiation, elongation, and termination. In vivo data suggests that elongating polymerases exhibit heterogeneity throughout the gene body, suggestive of changes in elongation rate and/or pausing. Here, we review evidence from a variety of different experimental approaches for understanding regulation of transcription elongation. We compare steady-state measurements of nascent RNA density and polymerase occupancy to time-resolved measurements and point out areas of disagreement. Finally, we discuss future avenues of investigation for understanding this critically important step in gene regulation. This article is part of a Special Issue entitled: Chromatin in time and space.► Heterogeneity in elongation kinetics is due to pausing and changes in velocity. ► Steady-state measures of polymerase occupancy reflect both initiation and elongation. ► Chromatin plays a role in modifying elongation rates. ► Time-resolved measures of pre-mRNA synthesis do not suggest changes in elongation.

Keywords: Transcription; Elongation; Single-molecule; Splicing; Imaging; Polymerase

Co-transcriptional regulation of alternative pre-mRNA splicing by Sanjeev Shukla; Shalini Oberdoerffer (pp. 673-683).

While studies of alternative pre-mRNA splicing regulation have typically focused on RNA-binding proteins and their target sequences within nascent message, it is becoming increasingly evident that mRNA splicing, RNA polymerase II (pol II) elongation and chromatin structure are intricately intertwined. The majority of introns in higher eukaryotes are excised prior to transcript release in a manner that is dependent on transcription through pol II. As a result of co-transcriptional splicing, variations in pol II elongation influence alternative splicing patterns, wherein a slower elongation rate is associated with increased inclusion of alternative exons within mature mRNA. Physiological barriers to pol II elongation, such as repressive chromatin structure, can thereby similarly impact splicing decisions. Surprisingly, pre-mRNA splicing can reciprocally influence pol II elongation and chromatin structure. Here, we highlight recent advances in co-transcriptional splicing that reveal an extensive network of coupling between splicing, transcription and chromatin remodeling complexes. This article is part of a Special Issue entitled: Chromatin in time and space.► The majority of metazoan introns are excised co-transcriptionally. ► Pre-mRNA processing is coordinated through the carboxy-terminal domain of pol II. ► Pol II elongation kinetics influence splicing decisions. ► Exons display distinct chromatin architecture relative to introns. ► Modulation of chromatin can result in altered pre-mRNA splicing.

Keywords: Alternative pre-mRNA splicing; Chromatin; Transcription elongation; RNA polymerase II

Genome-wide studies of the transcriptional regulation by p53 by Mangmang Li; Yunlong He; Xi Feng; Jing Huang (pp. 684-687).

The tumor suppressor p53 is arguably the most important transcription factor that safe-guards the genome. Although it is clear that the transcriptional activity of p53 is required for its tumor suppressive function, the underlying mechanisms are still largely unknown. In the past several years, genome-wide approaches have provided novel insights into the tumor suppressive functions of p53. This mini-review summarizes recent progress in studying these functions using genome-wide approaches, and offers some perspectives on this rapidly expanding field. This article is part of a Special Issue entitled: Chromatin in time and space.► Mechanisms underlying the tumor suppressive functions of p53 are still unclear. ► Binding of p53 at non-promoter regions is not fully understood. ► Dissection of p53 regulated transcriptome is challenging. ► Non-coding RNAs join the family of p53 targets. ► Epigenetic regulation of chromatin binding of p53 has not been fully explored.

Keywords: p53; Genome-wide; Cancer; Epigenetics; Chromatin; Histone

The dual lives of bidirectional promoters by Clay Wakano; Jung S. Byun; Li-Jun Di; Kevin Gardner (pp. 688-693).

The sequencing of the human genome led to many insights into gene organization and structure. One interesting observation was the high frequency of bidirectional promoters characterized by two protein encoding genes whose promoters are arranged in a divergent or “head-to-head” configuration with less than 2000 base pairs of intervening sequence. Computational estimates published by various groups indicate that nearly 10% of the coding gene promoters are arranged in such a manner and the extent of this bias is a unique feature of mammalian genomes. Moreover, as a class, head-to-head promoters appear to be enriched in specific categories of gene function. Here we review the structure, composition, genomic properties and functional classifications of genes controlled by bidirectional promoters and explore the biological implication of these features. This article is part of a Special Issue entitled: Chromatin in time and space.► Bidirectional promoters encompass approximately 10% of the human genome. ► Functional classes cover DNA repair, chromatin stability, maintenance and assembly. ► CtBP binds to multiple transcription factors that target bidirectional promoters. ► CtBP may link bidirectional promoters to multiple epigenetic regulators in cancer. ► CtBP assembly at bidirectional promoters may drive multiple hallmarks of cancer.

Keywords: Promoter; Chromatin; Transcription; Gene regulation

Isolation and characterization of transcription fidelity mutants by Jeffrey N. Strathern; Ding Jun Jin; Donald L. Court; Mikhail Kashlev (pp. 694-699).

Accurate transcription is an essential step in maintaining genetic information. Error-prone transcription has been proposed to contribute to cancer, aging, adaptive mutagenesis, and mutagenic evolution of retroviruses and retrotransposons. The mechanisms controlling transcription fidelity and the biological consequences of transcription errors are poorly understood. Because of the transient nature of mRNAs and the lack of reliable experimental systems, the identification and characterization of defects that increase transcription errors have been particularly challenging. In this review we describe novel genetic screens for the isolation of fidelity mutants in both Saccharomyces cerevisiae and Escherichia coli RNA polymerases. We obtained and characterized two distinct classes of mutants altering NTP misincorporation and transcription slippage both in vivo and in vitro. Our study not only validates the genetic schemes for the isolation of RNA polymerase mutants that alter fidelity, but also sheds light on the mechanism of transcription accuracy. This article is part of a Special Issue entitled: Chromatin in time and space.► Accurate RNA synthesis is necessary for preservation of biological information. ► Studying transcription fidelity is a challenge due to the transient nature of mRNAs. ► Genetic schemes for the isolation of fidelity RNA polymerase mutants are developed. ► Biochemical assays for detection of transcription errors are designed. ► The consequences of transcription errors by the fidelity mutants are discussed.

Keywords: Abbreviations; RNAP; E. coli; RNA polymerase; Pol II; yeast RNA polymeraseRNA polymerase; Transcription fidelity; Slippage; NTP misincorporation

Coupling polymerase pausing and chromatin landscapes for precise regulation of transcription by Daniel A. Gilchrist; Karen Adelman (pp. 700-706).

Altering gene expression in response to stimuli is a pivotal mechanism through which organisms execute developmental programs and respond to changes in their environment. Packaging of promoter DNA into chromatin can greatly impact the ability of RNA polymerase II to access and transcribe a gene. Promoter chromatin environments thus play a central role in establishing transcriptional output appropriate for specific environmental conditions or developmental states. Recent genomic studies have illuminated general principles of chromatin organization and deepened our understanding of how promoter sequence and nucleosome architecture may impact gene expression. Concurrently, pausing of polymerase during early elongation has been recognized as an important event influencing transcription of genes within stimulus-responsive networks. Promoters regulated by pausing are now recognized to possess a distinct chromatin architecture that may facilitate the plasticity of gene expression in response to signaling events. Here we review advances in understanding chromatin and pausing, and explore how coupling Pol II pausing to distinct promoter architectures may help organisms achieve flexible yet precise transcriptional control. This article is part of a Special Issue entitled: Chromatin in time and space.►Pol II pausing and promoter chromatin cooperate to precisely regulate transcription. ►Pausing is coupled to specialized promoter nucleosome architectures. ►Distinct chromatin landscapes are enriched in paused promoters. ►Promoter architecture and chromatin landscapes may enable cell-type specific pausing.

Keywords: Chromatin; Nucleosome; Pausing; Transcription

Multiple modes of chromatin remodeling by Forkhead box proteins by Avin S. Lalmansingh; Sudipan Karmakar; Yetao Jin; Akhilesh K. Nagaich (pp. 707-715).

Forkhead box (FOX) proteins represent a large family of transcriptional regulators unified by their DNA binding domain (DBD) known as a ‘forkhead’ or ‘winged helix’ domain. Over 40 FOX genes have been identified in the mammalian genome. FOX proteins share significant sequence similarities in the DBD which allow them to bind to a consensus DNA response element. However, their modes of action are quite diverse as they regulate gene expression by acting as pioneer factors, transcription factors, or both. This review focuses on the mechanisms of chromatin remodeling with an emphasis on three sub-classes—FOXA, FOXO, and FOXP members. FOXA proteins serve as pioneer factors to open up local chromatin structure and thereby increase accessibility of chromatin to factors regulating transcription. FOXP proteins, in contrast, function as classic transcription factors to recruit a variety of chromatin modifying enzymes to regulate gene expression. FOXO proteins represent a hybrid subclass having dual roles as pioneering factors and transcription factors. A subset of FOX proteins interacts with condensed mitotic chromatin and may function as ‘bookmarking’ agents to maintain transcriptional competence at specific genomic sites. The overall diversity in chromatin remodeling function by FOX proteins is related to unique structural motifs present within the DBD flanking regions that govern selective interactions with core histones and/or chromatin coregulatory proteins. This article is part of a Special Issue entitled: Chromatin in time and space.► Different chromatin remodeling mechanisms by Forkhead box proteins are reviewed. ► They carry out diverse remodeling as pioneer factors, transcription factors or both. ► They possess a conserved Forkhead box domain giving specificity in DNA recognition. ► N- and C-termini regions flanking DBD provide functional diversity. ► Unique flanking motifs specify interactions with histones and chromatin coregulators.

Keywords: Forkhead box protein; Chromatin remodeling; FOXA1; FOXP3; FOXO1; Pioneer factor

Chromatin remodeling during glucocorticoid receptor regulated transactivation by Heather A. King; Kevin W. Trotter; Trevor K. Archer (pp. 716-726).

Steroid hormone receptor (SR) signaling leads to widespread changes in gene expression, and aberrant SR signaling can lead to malignancies including breast, prostate, and lung cancers. Chromatin remodeling is an essential component of SR signaling, and defining the process of chromatin and nucleosome remodeling during signaling is critical to the continued development of related therapies. The glucocorticoid receptor (GR) is a key SR that activates numerous promoters including the well defined MMTV promoter. The activation of MMTV by GR provides an excellent model for teasing apart the sequence of events between hormone treatment and changes in gene expression. Comparing hormone-induced transcription from stably integrated promoters with defined nucleosomal structure to that from transiently expressed, unstructured promoters permits key distinctions between interactions that require remodeling and those that do not. The importance of co-activators and histone modifications prior to remodeling and the formation of the preinitiation complex that follows can also be clarified by defining key transition points in the propagation of hormonal signals. Combined with detailed mapping of proteins along the promoter, a temporal and spatial understanding of the signaling and remodeling processes begins to emerge. In this review, we examine SR signaling with a focus on GR activation of the MMTV promoter. We also discuss the ATP-dependent remodeling complex SWI/SNF, which provides the necessary remodeling activity during GR signaling and interacts with several SRs. BRG1, the central ATPase of SWI/SNF, also interacts with a set of BAF proteins that help determine the specialized function and fine-tuned regulation of BRG1 remodeling activity. BRG1 regulation comes from its own subdomains as well as its interactive partners. In particular, the HSA domain region of BRG1 and unique features of its ATPase homology appear to play key roles in regulating remodeling function. Details of the inter-workings of this chromatin remodeling protein continue to be revealed and promise to improve our understanding of the mechanism of chromatin remodeling during steroid hormone signaling. This article is part of a Special Issue entitled: Chromatin in time and space.► Chromatin remodeling is an essential part of steroid hormone signaling. ► The glucocorticoid receptor is a particularly well explored member of the steroid receptor super family. ► The hormone inducible MMTV promoter provides a model for signal responsive remodeling. ► BRG1 mediated transactivation depends upon the makeup of the BAF complex. ► BRG1 activity is also regulated by its own subdomains and their interacting partners.

Keywords: Chromatin; Transcription; Steroid receptor; BRG1; MMTV; Chromatin remodeling

Epigenetic regulation of adipogenesis by histone methylation by Kai Ge (pp. 727-732).

Histone methylation is implicated in both gene activation and repression, depending on the specific lysine residue that gets methylated. Recent years have witnessed an explosive expansion of the list of remarkably site-specific histone methyltransferases and demethylases, which greatly facilitates the study on the biological functions of histone methylation in gene expression and cell differentiation in mammalian cells. Adipogenesis represents an excellent model system to understand transcriptional and epigenetic regulation of gene expression and cell differentiation. While transcriptional regulation of adipogenesis has been extensively studied, the roles of epigenetic mechanisms in particular histone methylation in regulation of adipogenesis have just begun to be understood. This review will summarize the recent progress on epigenetic regulation of adipogenesis by histone methylation, with a focus on histone H3K4 and H3K27. The available evidence suggests that site-specific histone methylations play critical roles in adipogenesis and control the expression of both positive and negative master regulators of adipogenesis. This article is part of a Special Issue entitled: Chromatin in time and space.► Histone methylations regulate gene expression and cell differentiation. ► Histone methylations are regulated by methyltransferases and demethylases. ► This review focuses on regulation of adipogenesis by histone methylation. ► H3K4 and H3K27 methylations control expression of master regulators of adipogenesis.

Keywords: Histone methylation; Adipogenesis; MLL4; PTIP; Ezh2; PPARγ

Roles for histone H3K4 methyltransferase activities during immunoglobulin class-switch recombination by Jeremy A. Daniel; André Nussenzweig (pp. 733-738).

Germ-line transcription of an antigen receptor gene segment is an essential feature of the targeting mechanism for DNA double-strand break formation during physiological DNA rearrangements in lymphocytes. Alterations in chromatin structure have long been postulated to regulate accessibility of recombinase activities for lymphocytes to generate antibody diversity; however, whether or not germ-line transcripts are the cause or the effect of chromatin changes at antigen receptor loci is still not clear. Methylation of histone H3 at lysine 4 is one of the most well-studied histone post-translational modifications yet we have only recently begun to understand the significance of the MLL-like H3K4 methyltransferase activities in lymphocyte function. While it is clear during lymphocyte development that H3K4me3 plays a critical role in targeting and stimulating RAG1/2 recombinase activity for V(D)J recombination, recent work suggests roles for this histone mark and different MLL-like complexes in mature B cells during immunoglobulin class-switch recombination. In this review, we focus our discussion to advances on how MLL-like complexes and H3K4 methylation may function during the germ-line transcription and recombinase targeting steps of class-switch recombination. This article is part of a Special Issue entitled: Chromatin in time and space.

Keywords: Histone; Methyltransferase; Lymphocyte; Recombination

Allele-specific gene expression and epigenetic modifications and their application to understanding inheritance and cancer by Maxwell P. Lee (pp. 739-742).

Epigenetic information is characterized by its plasticity during development and differentiation as well as its stable transmission during mitotic cell divisions in somatic tissues. This duality contrasts to genetic information, which is essentially static and identical in every cell in an organism with only a few exceptions such as immunoglobulin genes in lymphocytes. Epigenetics is traditionally perceived as a means to regulate gene expression without a change in DNA sequence. This, however, does not exclude a potential role for genetic variations in providing differential backgrounds on which epigenetic modulations and their regulatory consequences are achieved. An effective approach to investigating the interplay between genetic variations and epigenetic variations is through allele-specific analysis of epigenetics and gene expression. Such studies have generated many new insights into functions of genetic variations, mechanisms of gene expression regulation, and the role of mutations and epigenetic alterations in human cancer. This article is part of a Special Issue entitled: Chromatin in time and space.► I summarize the current research on allele-specific gene expression and epigenetic modifications. ► I discuss how allele-specific features can be applied to study inheritance. ► I provide some examples on how allele-specific analyses can help gain new insights into better understanding of cancer.

Keywords: Allele-specific gene expression; Allele-specific methylation; Allele-specific chromatin; Polymorphism; Inheritance; Cancer

γ-H2AX and other histone post-translational modifications in the clinic by Christophe E. Redon; Urbain Weyemi; Palak R. Parekh; Dejun Huang; Allison S. Burrell; William M. Bonner (pp. 743-756).

Chromatin is a dynamic complex of DNA and proteins that regulates the flow of information from genome to end product. The efficient recognition and faithful repair of DNA damage, particularly double-strand damage, is essential for genomic stability and cellular homeostasis. Imperfect repair of DNA double-strand breaks (DSBs) can lead to oncogenesis. The efficient repair of DSBs relies in part on the rapid formation of foci of phosphorylated histone H2AX (γ-H2AX) at each break site, and the subsequent recruitment of repair factors. These foci can be visualized with appropriate antibodies, enabling low levels of DSB damage to be measured in samples obtained from patients. Such measurements are proving useful to optimize treatments involving ionizing radiation, to assay in vivo the efficiency of various drugs to induce DNA damage, and to help diagnose patients with a variety of syndromes involving elevated levels of γ-H2AX. We will survey the state of the art of utilizing γ-H2AX in clinical settings. We will also discuss possibilities with other histone post-translational modifications. The ability to measure in vivo the responses of individual patients to particular drugs and/or radiation may help optimize treatments and improve patient care. This article is part of a Special Issue entitled: Chromatin in time and space.► We review the uses of γ-H2AX and other modified histone species in the clinic. ► γ-H2AX is a useful radiation biodosimeter. ► γ-H2AX can be used to assay DNA damage during chemotherapy. ► γ-H2AX can be used to optimize cancer treatments and other procedures. ► Uses of other modified histone species are discussed.

Keywords: γ-H2AX focus; DNA double-strand break; Ionizing radiation; Biodosimeter; Chemotherapy; Clinical study

The ghosts in the machine: DNA methylation and the mystery of differentiation by Victorino Briones; Kathrin Muegge (pp. 757-762).

Methylation regulates DNA by altering chromatin and limiting accessibility of transcription factors and RNA polymerase. In this way, DNA methylation controls gene expression and plays a role in ES cell regulation, tissue differentiation and the development of the organism. In abnormal circumstances methylation can also induce diseases and promote cancer progression. Chromatin remodeling proteins such as the SNF2 family member Lsh regulates genome-wide cytosine methylation patterns during mammalian development. Lsh promotes methylation by targeting and repressing repeat sequences that are imbedded in heterochromatin. Lsh also regulates cytosine methylation at unique loci. Alterations in histone modifications (such as H3K4me3, histone acetylation, H3K27me3 and H2Aub) can be associated with DNA methylation changes making Lsh-mediated cytosine methylation part of a larger epigenetic network defining gene expression and cellular differentiation during development. This article is part of a Special Issue entitled: Chromatin in time and space.► Cytosine methylation pattern are critical for cellular differentiation and development. ► Lsh is a SNF2 homologue that regulates cytosine methylation in mammals. ► Lsh controls DNA methylation and histone modifications at repeat elements and developmental genes. ► Abnormal chromatin organization and aberrant gene expression in the absence of Lsh lead to early lethality. ► The Lsh−/− mouse can serve as a model to investigate the biologic function of cytosine methylation.

Keywords: DNA methylation; SNF2; Embryogenesis

CpG methylation recruits sequence specific transcription factors essential for tissue specific gene expression by Raghunath Chatterjee; Charles Vinson (pp. 763-770).

CG methylation is an epigenetically inherited chemical modification of DNA found in plants and animals. In mammals it is essential for accurate regulation of gene expression and normal development. Mammalian genomes are depleted for the CG dinucleotide, a result of the chemical deamination of methyl-cytosine in CG resulting in TpG. Most CG dinucleotides are methylated, but ~15% are unmethylated. Five percent of CGs cluster into ~20,000 regions termed CG islands (CGI) which are generally unmethylated. About half of CGIs are associated with housekeeping genes. In contrast, the gene body, repeats and transposable elements in which CGs are generally methylated. Unraveling the epigenetic machinery operating in normal cells is important for understanding the epigenetic aberrations that are involved in human diseases including cancer. With the advent of high-throughput sequencing technologies, it is possible to identify the CG methylation status of all 30million unique CGs in the human genome, and monitor differences in distinct cell types during differentiation and development. Here we summarize the present understanding of DNA methylation in normal cells and discuss recent observations that CG methylation can have an effect on tissue specific gene expression. We also discuss how aberrant CG methylation can lead to cancer. This article is part of a Special Issue entitled: Chromatin in time and space.► Present understanding of DNA methylation in normal cells. ► A new function of CpG methylation: positive effect on tissue specific gene expression. ► Aberrant CpG methylation and cancer.

Keywords: Epigenetics; DNA methylation; Transcription; CpG island; Cancer; 5-azacytidine

Retrotransposons at Drosophila telomeres: Host domestication of a selfish element for the maintenance of genome integrity by Liang Zhang; Yikang S. Rong (pp. 771-775).

Telomere serves two essential functions for the cell. It prevents the recognition of natural chromosome ends as DNA breaks (the end capping function). It counteracts incomplete end replication by adding DNA to the ends of chromosomes (the end elongation function). In most organisms studied, telomerase fulfills the end elongation function. In Drosophila, however, telomere specific retrotransposons have been coerced into performing this essential function for the host. In this review, we focus our discussion on transposition mechanisms and transcriptional regulation of these transposable elements, and present provocative models for the purpose of spurring new interests in the field. This article is part of a Special Issue entitled: Chromatin in time and space.► Most transposable elements negatively impact genome integrity of the host. ► A group of non-LTR retransposons has been coerced to perform essential telomere maintaining function in Drosophila and other insect. ► How the elements transpose exclusively to the ends of chromosomes is poorly understood. ► We raise provocative models to spur new interests in the field.

Keywords: Telomere elongation; Non-LTR retrotransposon; Drosophila; piRNA; Telomerase recruitment; Telomere capping

Insights into assembly and regulation of centromeric chromatin in Saccharomyces cerevisiae by John S. Choy; Prashant K. Mishra; Wei-Chun Au; Munira A. Basrai (pp. 776-783).

At the core of chromosome segregation is the centromere, which nucleates the assembly of a macromolecular kinetochore (centromere DNA and associated proteins) complex responsible for mediating spindle attachment. Recent advances in centromere research have led to identification of many kinetochore components, such as the centromeric-specific histone H3 variant, CenH3, and its interacting partner, Scm3. Both are essential for chromosome segregation and are evolutionarily conserved from yeast to humans. CenH3 is proposed to be the epigenetic mark that specifies centromeric identity. Molecular mechanisms that regulate the assembly of kinetochores at specific chromosomal sites to mediate chromosome segregation are not fully understood. In this review, we summarize the current literature and discuss results from our laboratory, which show that restricting the localization of budding yeast CenH3, Cse4, to centromeres and balanced stoichiometry between Scm3 and Cse4, contribute to faithful chromosome transmission. We highlight our findings that, similar to other eukaryotic centromeres, budding yeast centromeric histone H4 is hypoacetylated, and we discuss how altered histone acetylation affects chromosome segregation. This article is part of a Special Issue entitled: Chromatin in time and space.► Hypoacetylation of centromeric H4 contributes to chromosome transmission fidelity. ► Altered histone H4 HAT and HDAC activities lead to chromosome missegregation. ► Mislocalization of Cse4 to non-centromeric regions leads to chromosome loss. ► N-terminal domain of Cse4 contributes to its exclusion from non-centromeric regions. ► Misregulation of Scm3/HJURP causes defects in chromosome segregation.

Keywords: Cse4; Histone H4 acetylation; Scm3/HJURP; Kinetochore; CenH3; Chromosome segregation

The consequences of chromosomal aneuploidy on the transcriptome of cancer cells by Thomas Ried; Yue Hu; Michael J. Difilippantonio; B. Michael Ghadimi; Marian Grade; Jordi Camps (pp. 784-793).

Chromosomal aneuploidies are a defining feature of carcinomas, i.e., tumors of epithelial origin. Such aneuploidies result in tumor specific genomic copy number alterations. The patterns of genomic imbalances are tumor specific, and to a certain extent specific for defined stages of tumor development. Genomic imbalances occur already in premalignant precursor lesions, i.e., before the transition to invasive disease, and their distribution is maintained in metastases, and in cell lines derived from primary tumors. These observations are consistent with the interpretation that tumor specific genomic imbalances are drivers of malignant transformation. Naturally, this precipitates the question of how such imbalances influence the expression of resident genes. A number of laboratories have systematically integrated copy number alterations with gene expression changes in primary tumors and metastases, cell lines, and experimental models of aneuploidy to address the question as to whether genomic imbalances deregulate the expression of one or few key genes, or rather affect the cancer transcriptome more globally. The majority of these studies showed that gene expression levels follow genomic copy number. Therefore, gross genomic copy number changes, including aneuploidies of entire chromosome arms and chromosomes, result in a massive deregulation of the transcriptome of cancer cells. This article is part of a Special Issue entitled: Chromatin in time and space.

Keywords: Aneuploidy; Gene expression; Copy number alterations; Solid tumors; Cell lines

The chromatin backdrop of DNA replication: Lessons from genetics and genome-scale analyses by Amy L. Conner; Mirit I. Aladjem (pp. 794-801).

The entire cellular genome must replicate during each cell cycle, but it is yet unclear how replication proceeds along with chromatin condensation and remodeling while ensuring the fidelity of the replicated genome. Mapping replication initiation sites can provide clues for the coordination of DNA replication and transcription on a whole-genome scale. Here we discuss recent insights obtained from genome-scale analyses of replication initiation sites and transcription in mammalian cells and ask how transcription and chromatin modifications affect the frequency of replication initiation events. We also discuss DNA sequences, such as insulators and replicators, which modulate replication and transcription of target genes, and use genome-wide maps of replication initiation sites to evaluate possible commonalities between replicators and chromatin insulators. This article is part of a Special Issue entitled: Chromatin in time and space.► Genome-scale maps and genetic analyses probe how replication coordinates with transcription. ► Replication starts preferentially at accessible chromatin and at methylated CpG regions. ► Replication starts at regions that straddle but do not overlap transcription start site. ► Replication starts preferentially at moderate, but not highly transcribed, chromatin regions. ► DNA sequences that start replication might colocalize with chromatin insulators.

Keywords: DNA replication; Chromatin; Cell cycle; Transcription

Chromatin changes in the development and pathology of the Fragile X-associated disorders and Friedreich ataxia by Daman Kumari; Rachel Lokanga; Dmitry Yudkin; Xiao-Nan Zhao; Karen Usdin (pp. 802-810).

The Fragile X-associated disorders (FXDs) and Friedreich ataxia (FRDA) are genetic conditions resulting from expansion of a trinucleotide repeat in a region of the affected gene that is transcribed but not translated. In the case of the FXDs, pathology results from expansion of CGG•CCG-repeat tract in the 5′ UTR of the FMR1 gene, while pathology in FRDA results from expansion of a GAA•TTC-repeat in intron 1 of the FXN gene. Expansion occurs during gametogenesis or early embryogenesis by a mechanism that is not well understood. Associated Expansion then produces disease pathology in various ways that are not completely understood either. In the case of the FXDs, alleles with 55–200 repeats express higher than normal levels of a transcript that is thought to be toxic, while alleles with >200 repeats are silenced. In addition, alleles with >200 repeats are associated with a cytogenetic abnormality known as a fragile site, which is apparent as a constriction or gap in the chromatin that is seen when cells are grown in presence of inhibitors of thymidylate synthase. FRDA alleles show a deficit of the FXN transcript. This review will address the role of repeat-mediated chromatin changes in these aspects of FXD and FRDA disease pathology. This article is part of a Special Issue entitled: Chromatin in time and space.► The Fragile X disorders and Friedreich ataxia result from expansion of a tandem repeat tract. ► In all of these diseases the repeat is transcribed but not translated. ► Evidence suggests that repeat-mediated chromatin changes are responsible for disease pathology.

Keywords: Abbreviations; FM; full mutation; FRDA; Friedreich ataxia; FS; fragile site; FXDs; Fragile X associated disorders; FXPOI; Fragile X-associated primary ovarian insufficiency; FXTAS; Fragile X-associated tremor and ataxia syndrome; FXS; Fragile X syndrome; PM; premutation; REDs; Repeat Expansion Diseases; SCA; spinocerebellar ataxiaFriedreich ataxia; Fragile X disorders; Heterochromatin; Repeat-mediated chromatin changes; Gene silencing; Chromosome fragility

Chromatin dynamics in DNA double-strand break repair by Lei Shi; Philipp Oberdoerffer (pp. 811-819).

DNA double-strand breaks (DSBs) occur in the context of a highly organized chromatin environment and are, thus, a significant threat to the epigenomic integrity of eukaryotic cells. Changes in break-proximal chromatin structure are thought to be a prerequisite for efficient DNA repair and may help protect the structural integrity of the nucleus. Unlike most bona fide DNA repair factors, chromatin influences the repair process at several levels: the existing chromatin context at the site of damage directly affects the access and kinetics of the repair machinery; DSB induced chromatin modifications influence the choice of repair factors, thereby modulating repair outcome; lastly, DNA damage can have a significant impact on chromatin beyond the site of damage. We will discuss recent findings that highlight both the complexity and importance of dynamic and tightly orchestrated chromatin reorganization to ensure efficient DSB repair and nuclear integrity. This article is part of a Special Issue entitled: Chromatin in time and space.► Chromatin reorganization is a key aspect of eukaryotic DNA repair. ► DNA break-induced chromatin remodeling affects repair factor access and choice. ► The pre-existing chromatin environment influences DNA repair kinetics and outcome. ► DNA breaks cause epigenomic changes that extend beyond the site of damage.

Keywords: Chromatin; DNA repair; Nuclear reorganization; Histone modification

Hitchhiking on host chromatin: how papillomaviruses persist by Alison A. McBride; Nozomi Sakakibara; Wesley H. Stepp; Moon Kyoo Jang (pp. 820-825).

Persistent viruses need mechanisms to protect their genomes from cellular defenses and to ensure that they are efficiently propagated to daughter host cells. One mechanism by which papillomaviruses achieve this is through the association of viral genomes with host chromatin, mediated by the viral E2 tethering protein. Association of viral DNA with regions of active host chromatin ensures that the virus remains transcriptionally active and is not relegated to repressed heterochromatin. In addition, viral genomes are tethered to specific regions of host mitotic chromosomes to efficiently partition their DNA to daughter cells. Vegetative viral DNA replication also initiates at specific regions of host chromatin, where the viral E1 and E2 proteins initiate a DNA damage response that recruits cellular DNA damage and repair proteins to viral replication foci for efficient viral DNA synthesis. Thus, these small viruses have capitalized on interactions with chromatin to efficiently target their genomes to beneficial regions of the host nucleus. This article is part of a Special Issue entitled: Chromatin in time and space.► Papillomaviruses develop a long term, persistent relationship with their host. ► The papillomavirus life cycle is tightly linked to keratinocyte differentiation. ► Papillomaviruses must overcome intrinsic cellular defenses to establish an efficient infection. ► Papillomaviruses retain their genomes in infected cells by tethering them to host chromatin. ► Papillomaviruses utilize the cellular DNA damage response for their own replication.

Keywords: Virus; HPV; Replication; Chromatin; DNA damage response; ND10

Chromosome dynamics in multichromosome bacteria by Jyoti K. Jha; Jong Hwan Baek; Tatiana Venkova-Canova; Dhruba K. Chattoraj (pp. 826-829).

On the basis of limited information, bacteria were once assumed to have no more than one chromosome. In the era of genomics, it has become clear that some, like eukaryotes, have more than one chromosome. Multichromosome bacteria provide opportunities to investigate how split genomes emerged, whether the individual chromosomes communicate to coordinate their replication and segregation, and what selective advantages split genomes might provide. Our current knowledge of these topics comes mostly from studies in Vibrio cholerae, which has two chromosomes, chr1 and chr2. Chr1 carries out most of the house-keeping functions and is considered the main chromosome, whereas chr2 appears to have originated from a plasmid and has acquired genes of mostly unknown origin and function. Nevertheless, unlike plasmids, chr2 replicates once and only once per cell cycle, like a bona fide chromosome. The two chromosomes replicate and segregate using separate programs, unlike eukaryotic chromosomes. They terminate replication synchronously, suggesting that there might be communication between them. Replication of the chromosomes is affected by segregation genes but in a chromosome specific fashion, a new development in the field of DNA replication control. The split genome allows genome duplication to complete in less time and with fewer replication forks, which could be beneficial for genome maintenance during rapid growth, which is the norm for V. cholerae in broth cultures and in the human host. In the latter, the expression of chr2 genes increases preferentially. Studies of chromosome maintenance in multichromosomal bacteria, although in their infancy, are already broadening our view of chromosome biology. This article is part of a Special Issue entitled: Chromatin in time and space.► Some bacteria, like eukaryotes, have multiple chromosomes ► Some of these chromosomes appear to be of plasmid provenance ► Maintenance mechanisms of individual chromosomes are distinct, not as in eukaryotes

Keywords: Divided genome; Genome evolution; Secondary chromosomes; Genome maintenance; Communication between chromosome replication and segregation; V. cholerae

Architectural organization in E. coli nucleoid by Mirjana Macvanin; Sankar Adhya (pp. 830-835).

In contrast to organized hierarchical structure of eukaryotic chromosome, bacterial chromosomes are believed not to have such structures. The genomes of bacteria are condensed into a compact structure called the nucleoid. Among many architectural, histone-like proteins which associate with the chromosomal DNA is HU which is implicated in folding DNA into a compact structure by bending and wrapping DNA. Unlike the majority of other histone-like proteins, HU is highly conserved in eubacteria and unique in its ability to bind RNA. Furthermore, an HU mutation profoundly alters the cellular transcription profile and consequently has global effects on physiology and the lifestyle of E. coli. Here we provide a short overview of the mechanisms by which the nucleoid is organized into different topological domains. We propose that HU is a major player in creating domain-specific superhelicities and thus influences the transcription profile from the constituent promoters. This article is part of a Special Issue entitled: Chromatin in time and space.► A review on chromosome organization in E. coli nucleoid. ► We provide an overview of the mechanisms by which the nucleoid is organized into different domains. ► We discuss the role of histone-like bacterial protein HU in chromosome organization.

Keywords: Nucleoid; Histone-like protein; HU; Chromosome organization

Targeting epigenetic mediators of gene expression in thoracic malignancies by David S. Schrump (pp. 836-845).

Lung and esophageal cancers and malignant pleural mesotheliomas are highly lethal neoplasms that are leading causes of cancer-related deaths worldwide. Presently, limited information is available pertaining to epigenetic mechanisms mediating initiation and progression of these neoplasms. The following presentation will focus on the potential clinical relevance of epigenomic alterations in thoracic malignancies mediated by DNA methylation, perturbations in the histone code, and polycomb group proteins, as well as ongoing translational efforts to target epigenetic regulators of gene expression for treatment of these neoplasms. This article is part of a Special Issue entitled: Chromatin in time and space.► Overview of clinically relevant alterations in DNA methylation in thoracic malignancies. ► Correlation of histone alterations and polycomb protein expression with patient outcome. ► Review of translational efforts to target the epigenome in thoracic malignancies.

Keywords: Lung cancer; Esophageal cancer; Mesothelioma; Epigenetics; Prognosis; Clinical trials

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BBA - Gene Regulatory Mechanisms (v.1819, #7)