Global Chromatin State Research Articles

Short-chain fatty acids activate acetyltransferase p300

Short-chain fatty acids (SCFAs) acetate, propionate, and butyrate are produced in large quantities by the gut microbiome and contribute to a wide array of physiological processes. While the underlying mechanisms are largely unknown, many effects of SCFAs have been traced to changes in the cell's epigenetic state. Here, we systematically investigate how SCFAs alter the epigenome. Using quantitative proteomics of histone modification states, we identified rapid and sustained increases in histone acetylation after the addition of butyrate or propionate, but not acetate. While decades of prior observations would suggest that hyperacetylation induced by SCFAs are due to inhibition of histone deacetylases (HDACs), we found that propionate and butyrate instead activate the acetyltransferase p300. Propionate and butyrate are rapidly converted to the corresponding acyl-CoAs which are then used by p300 to catalyze auto-acylation of the autoinhibitory loop, activating the enzyme for histone/protein acetylation. This data challenges the long-held belief that SCFAs mainly regulate chromatin by inhibiting HDACs, and instead reveals a previously unknown mechanism of HAT activation that can explain how an influx of low levels of SCFAs alters global chromatin states.

Maternal factor NELFA drives a 2C-like state in mouse embryonic stem cells.

Mouse embryonic stem cells (ESCs) sporadically transit into an early embryonic-like state characterized by the expression of 2-cell (2C) stage-restricted transcripts. Here, we identify a maternal factor-negative elongation factor A (NELFA)-whose heterogeneous expression in mouse ESCs is coupled to 2C gene upregulation and expanded developmental potential in vivo. We show that NELFA partners with Top2a in an interaction specific to the 2C-like state, and that it drives the expression of Dux-a key 2C regulator. Accordingly, loss of NELFA and/or Top2a suppressed Dux activation. Further characterization of 2C-like cells uncovered reduced glycolytic activity; remarkably, mere chemical suppression of glycolysis was sufficient to promote a 2C-like fate, obviating the need for genetic manipulation. Global chromatin state analysis on NELFA-induced cells revealed decommissioning of ESC-specific enhancers, suggesting ESC-state impediments to 2C reversion. Our study positions NELFA as one of the earliest drivers of the 2C-like state and illuminates factors and processes that govern this transition.

29 Introduction to functional annotation of animal genomes consortium

Abstract The annotation of the functional components of genomes is required for both basic and applied genomic research, yet farmed animal genomics are deficient in such annotation. This talk will introduce the Functional Annotation of ANimal Genomes (FAANG) consortium, which was initiated in 2014 to address this knowledge deficit. An overarching theme of early FAANG efforts is the collaborative approach required for such comprehensive research, thus another emphasis of this talk will be our community efforts to develop the environment for successful FAANG research. FAANG scientists have created a coordinated data collection and analysis enterprise crucial for success of this global effort, and funding of ~$20 million has been secured for current projects in several species, with an anticipated doubling of this support during 2019. Member FAANG consortium labs are producing genome-wide datasets on RNA expression, DNA methylation, chromatin modification, chromatin accessibility, and chromatin interactions for many agriculturally relevant animal species. The goal of a first-generation global chromatin state map for cattle, chicken, pig, and potentially other species is projected for completion in the next 3–5 years. These data will be used both to better understand animal genome function at the epigenetic level, as well as improve the precision and sensitivity of genomic selection for animal improvement. The functional annotation delivered by the FAANG initiative will add value and utility to the greatly improved genome sequences being established for domesticated animal species.

Chromatin-associated RNA sequencing (ChAR-seq) maps genome-wide RNA-to-DNA contacts.

RNA is a critical component of chromatin in eukaryotes, both as a product of transcription, and as an essential constituent of ribonucleoprotein complexes that regulate both local and global chromatin states. Here, we present a proximity ligation and sequencing method called Chromatin-Associated RNA sequencing (ChAR-seq) that maps all RNA-to-DNA contacts across the genome. Using Drosophila cells, we show that ChAR-seq provides unbiased, de novo identification of targets of chromatin-bound RNAs including nascent transcripts, chromosome-specific dosage compensation ncRNAs, and genome-wide trans-associated RNAs involved in co-transcriptional RNA processing.

Insights into Dynamic Mitotic Chromatin Organization Through the NIMA Kinase Suppressor SonC, a Chromatin-Associated Protein Involved in the DNA Damage Response

The nuclear pore complex proteins SonA and SonB, the orthologs of mammalian RAE1 and NUP98, respectively, were identified in Aspergillus nidulans as cold-sensitive suppressors of a temperature-sensitive allele of the essential mitotic NIMA kinase (nimA1). Subsequent analyses found that sonB1 mutants exhibit temperature-dependent DNA damage sensitivity. To understand this pathway further, we performed a genetic screen to isolate additional conditional DNA damage-sensitive suppressors of nimA1. We identified two new alleles of SonA and four intragenic nimA mutations that suppress the temperature sensitivity of the nimA1 mutant. In addition, we identified SonC, a previously unstudied binuclear zinc cluster protein involved with NIMA and the DNA damage response. Like sonA and sonB, sonC is an essential gene. SonC localizes to nuclei and partially disperses during mitosis. When the nucleolar organizer region (NOR) undergoes mitotic condensation and removal from the nucleolus, nuclear SonC and histone H1 localize in a mutually exclusive manner with H1 being removed from the NOR region and SonC being absent from the end of the chromosome beyond the NOR. This region of chromatin is adjacent to a cluster of nuclear pore complexes to which NIMA localizes last during its progression around the nuclear envelope during initiation of mitosis. The results genetically extend the NIMA regulatory system to include a protein with selective large-scale chromatin location observed during mitosis. The data suggest a model in which NIMA and SonC, its new chromatin-associated suppressor, might help to orchestrate global chromatin states during mitosis and the DNA damage response.

Dynamic expression of combinatorial replication-dependent histone variant genes during mouse spermatogenesis

Nucleosomes are basic chromatin structural units that are formed by DNA sequences wrapping around histones. Global chromatin states in different cell types are specified by combinatorial effects of post-translational modifications of histones and the expression of histone variants. During mouse spermatogenesis, spermatogonial stem cells (SSCs) self-renew while undergo differentiation, events that occur in the company of constant re-modeling of chromatin structures. Previous studies have shown that testes contain highly expressed or specific histone variants to facilitate these epigenetic modifications. However, mechanisms of regulating the epigenetic changes and the specific histone compositions of spermatogenic cells are not fully understood. Using real time quantitative RT-PCR, we examined the dynamic expression of replication-dependent histone genes in post-natal mouse testes. It was found that distinct sets of histone genes are expressed in various spermatogenic cells at different stages during spermatogenesis. While gonocyte-enriched testes from mice at 2-dpp (days post partum) express pre-dominantly thirteen histone variant genes, SSC-stage testes at 9-dpp highly express a different set of eight histone genes. During differentiation stage when testes are occupied mostly by spermatocytes and spermatids, another twenty-two histone genes are expressed much higher than the rest, including previously known testis-specific hist1h1t, hist1h2ba and hist1h4c. In addition, histone genes that are pre-dominantly expressed in gonocytes and SSCs are also highly expressed in embryonic stem cells. Several of them were changed when embryoid bodies were formed from ES cells, suggesting their roles in regulating pluripotency of the cells. Further more, differentially expressed histone genes are specifically localized in either SSCs or spermatocytes and spermatids, as demonstrated by in situ hybridization using gene specific probes. Taken together, results presented here revealed that different combinations of histone variant genes are expressed in distinct spermatogenic cell types accompanying the progression of self-renewal and differentiation of SSCs, suggesting a systematic regulatory role histone variants play during spermatogenesis.

The ability of the Su(Hw) protein to create a platform for ORC binding does not depend on the type of surrounding chromatin

DNA replication begins from multiple sites distributed throughout the genome, named replication origins. Despite the increasing amount of data on the properties of replication origins, it is still unknown what factors are the primary determinants of ORC localization. Su(Hw) is a zinc-finger protein responsible for the activity of the best-studied Drosophila insulators. In the present work, we show that the insulator protein Su(Hw) recruits the histon acetyltransferase complex SAGA and chromatin remodeler dSWI/SNF to Su(Hw)-dependent insulators and creates a platform for ORC binding. We have found Su(Hw) to be necessary for chromatin remodeling and ORC recruitment regardless of the surrounding chromatin type. Thus, the global chromatin state does not affect the molecular mechanism underlying ORC positioning in genome; it is rather the DNA-binding proteins that are the key determinants that create the proper chromatin structure for ORC binding. Su(Hw) is the first example of such protein.

Global Chromatin State Analysis Reveals Lineage-Specific Enhancers during the Initiation of Human T helper 1 and T helper 2 Cell Polarization

Naive CD4⁺ T cells can differentiate into specific helper and regulatory T cell lineages in order to combat infection and disease. The correct response to cytokines and a controlled balance of these populations is critical for the immune system and the avoidance of autoimmune disorders. To investigate how early cell-fate commitment is regulated, we generated the first human genome-wide maps of histone modifications that reveal enhancer elements after 72 hr of in vitro polarization toward T helper 1 (Th1) and T helper 2 (Th2) cell lineages. Our analysis indicated that even at this very early time point, cell-specific gene regulation and enhancers were at work directing lineage commitment. Further examination of lineage-specific enhancers identified transcription factors (TFs) with known and unknown T cell roles as putative drivers of lineage-specific gene expression. Lastly, an integrative analysis of immunopathogenic-associated SNPs suggests a role for distal regulatory elements in disease etiology.

Genome-wide Chromatin State Transitions Associated with Developmental and Environmental Cues

Differences in chromatin organization are key to the multiplicity of cell states that arise from a single genetic background, yet the landscapes of in vivo tissues remain largely uncharted. Here, we mapped chromatin genome-wide in a large and diverse collection of human tissues and stem cells. The maps yield unprecedented annotations of functional genomic elements and their regulation across developmental stages, lineages, and cellular environments. They also reveal global features of the epigenome, related to nuclear architecture, that also vary across cellular phenotypes. Specifically, developmental specification is accompanied by progressive chromatin restriction as the default state transitions from dynamic remodeling to generalized compaction. Exposure to serum in vitro triggers a distinct transition that involves de novo establishment of domains with features of constitutive heterochromatin. We describe how these global chromatin state transitions relate to chromosome and nuclear architecture, and discuss their implications for lineage fidelity, cellular senescence, and reprogramming.

HMGA regulates the global chromatin state and neurogenic potential in neocortical precursor cells

Neural precursor cells (NPCs) in the mouse neocortex generate various neuronal and glial cell types in a developmental stage–dependent manner. Most NPCs lose their neurogenic potential during development, although the underlying mechanisms of this process are not fully understood. We found that the chromatin of mouse NPCs gradually becomes more condensed and less dynamic on a global scale during neocortical development. Furthermore, we found high mobility group A (HMGA) proteins to be essential for the open chromatin state of NPCs at early developmental stages. Knockdown of HMGA proteins in early-stage NPCs reduced their neurogenic potential. Conversely, overexpression of HMGA proteins conferred neurogenic potential on late-stage NPCs, an effect that was antagonized by coexpression of a histone H1 mutant that inhibits chromatin opening. Thus, HMGA proteins contribute to the neurogenic potential of NPCs in the early stages of neocortical development, possibly through induction of an open chromatin state.

Chromatin States Accurately Classify Cell Differentiation Stages

Gene expression is controlled by the concerted interactions between transcription factors and chromatin regulators. While recent studies have identified global chromatin state changes across cell-types, it remains unclear to what extent these changes are co-regulated during cell-differentiation. Here we present a comprehensive computational analysis by assembling a large dataset containing genome-wide occupancy information of 5 histone modifications in 27 human cell lines (including 24 normal and 3 cancer cell lines) obtained from the public domain, followed by independent analysis at three different representations. We classified the differentiation stage of a cell-type based on its genome-wide pattern of chromatin states, and found that our method was able to identify normal cell lines with nearly 100% accuracy. We then applied our model to classify the cancer cell lines and found that each can be unequivocally classified as differentiated cells. The differences can be in part explained by the differential activities of three regulatory modules associated with embryonic stem cells. We also found that the “hotspot” genes, whose chromatin states change dynamically in accordance to the differentiation stage, are not randomly distributed across the genome but tend to be embedded in multi-gene chromatin domains, and that specialized gene clusters tend to be embedded in stably occupied domains.

MeCP2: Phosphorylated Locally, Acting Globally

In this issue of Neuron, Greenberg and colleagues revise our understanding of how activity-dependent MeCP2 phosphorylation regulates distinct aspects of brain development and circuit function. The study also suggests a prominent role for MeCP2 in the regulation of global chromatin state invivo.

Open chromatin in pluripotency and reprogramming

Pluripotent stem cells can be derived from embryos or induced from adult cells by reprogramming. They are unique among stem cells in that they can give rise to all cell types of the body. Recent findings indicate that a particularly 'open' chromatin state contributes to maintenance of pluripotency. Two principles are emerging: specific factors maintain a globally open chromatin state that is accessible for transcriptional activation; and other chromatin regulators contribute locally to the silencing of lineage-specific genes until differentiation is triggered. These same principles may apply during reacquisition of an open chromatin state upon reprogramming to pluripotency, and during de-differentiation in cancer.

A Cellular Memory of Developmental History Generates Phenotypic Diversity in C. elegans

Early life experiences have a major impact on adult phenotypes [1-3]. However, the mechanisms by which animals retain a cellular memory of early experience are not well understood. Here we show that adult wild-type Caenorhabditis elegans that transiently pass through the stress-resistant dauer larval stage exhibit distinct gene expression profiles and life history traits, as compared to adult animals that bypassed this stage. Using chromatin immunoprecipitation experiments coupled with massively parallel sequencing, we found that genome-wide levels of specific histone tail modifications are markedly altered in postdauer animals. Mutations in subsets of genes implicated in chromatin remodeling abolish, or alter, the observed changes in gene expression and life history traits in postdauer animals. Modifications to the epigenome as a consequence of early experience may contribute in part to a memory of early experience and generate phenotypic variation in an isogenic population.

Chromatinisation of herpesvirus genomes

The double-stranded DNA genomes of herpesviruses exist in at least three alternative global chromatin states characterised by distinct nucleosome content. When encapsidated in virus particles, the viral DNA is devoid of any nucleosomes. In contrast, within latently infected nuclei herpesvirus genomes are believed to form regular nucleosomal structures resembling cellular chromatin. Finally, during productive infection nuclear viral DNA appears to adopt a state of intermediate chromatin formation with irregularly spaced nucleosomes. Nucleosome occupancy coupled with posttranslational histone modifications and other epigenetic marks may contribute significantly to the extent and timing of transcription from the viral genome and, consequently, to the outcome of infection. Recent research has provided first insights into the viral and cellular mechanisms that either maintain individual herpesvirus chromatin states or mediate transition between them. Here, we summarise and discuss both early work and new developments pointing towards common principles pertinent to the dynamic structure and epigenetic regulation of herpesvirus chromatin. Special emphasis is given to the emerging similarities in nucleosome assembly and disassembly processes on herpes simplex virus type 1 and human cytomegalovirus genomes over the course of the viral productive replication cycle and during the switch between latent and lytic infectious stages.

GlcNAcylation of a histone methyltransferase in retinoic-acid-induced granulopoiesis

The post-translational modifications of histone tails generate a 'histone code' that defines local and global chromatin states. The resultant regulation of gene function is thought to govern cell fate, proliferation and differentiation. Reversible histone modifications such as methylation are under mutual controls to organize chromosomal events. Among the histone modifications, methylation of specific lysine and arginine residues seems to be critical for chromatin configuration and control of gene expression. Methylation of histone H3 lysine 4 (H3K4) changes chromatin into a transcriptionally active state. Reversible modification of proteins by beta-N-acetylglucosamine (O-GlcNAc) in response to serum glucose levels regulates diverse cellular processes. However, the epigenetic impact of protein GlcNAcylation is unknown. Here we report that nuclear GlcNAcylation of a histone lysine methyltransferase (HKMT), MLL5, by O-GlcNAc transferase facilitates retinoic-acid-induced granulopoiesis in human HL60 promyelocytes through methylation of H3K4. MLL5 is biochemically identified in a GlcNAcylation-dependent multi-subunit complex associating with nuclear retinoic acid receptor RARalpha (also known as RARA), serving as a mono- and di-methyl transferase to H3K4. GlcNAcylation at Thr 440 in the MLL5 SET domain evokes its H3K4 HKMT activity and co-activates RARalpha in target gene promoters. Increased nuclear GlcNAcylation by means of O-GlcNAc transferase potentiates retinoic-acid-induced HL60 granulopoiesis and restores the retinoic acid response in the retinoic-acid-resistant HL60-R2 cell line. Thus, nuclear MLL5 GlcNAcylation triggers cell lineage determination of HL60 through activation of its HKMT activity.

Allele-Specific Chromatin Immunoprecipitation Studies Show Genetic Influence on Chromatin State in Human Genome

Several recent studies have shown a genetic influence on gene expression variation, including variation between the two chromosomes within an individual and variation between individuals at the population level. We hypothesized that genetic inheritance may also affect variation in chromatin states. To test this hypothesis, we analyzed chromatin states in 12 lymphoblastoid cells derived from two Centre d'Etude du Polymorphisme Humain families using an allele-specific chromatin immunoprecipitation (ChIP-on-chip) assay with Affymetrix 10K SNP chip. We performed the allele-specific ChIP-on-chip assays for the 12 lymphoblastoid cells using antibodies targeting at RNA polymerase II and five post-translation modified forms of the histone H3 protein. The use of multiple cell lines from the Centre d'Etude du Polymorphisme Humain families allowed us to evaluate variation of chromatin states across pedigrees. These studies demonstrated that chromatin state clustered by family. Our results support the idea that genetic inheritance can determine the epigenetic state of the chromatin as shown previously in model organisms. To our knowledge, this is the first demonstration in humans that genetics may be an important factor that influences global chromatin state mediated by histone modification, the hallmark of the epigenetic phenomena.