RESUMO
Nucleosomes block access to DNA methyltransferase, unless they are remodeled by DECREASE in DNA METHYLATION 1 (DDM1LSH/HELLS), a Snf2-like master regulator of epigenetic inheritance. We show that DDM1 promotes replacement of histone variant H3.3 by H3.1. In ddm1 mutants, DNA methylation is partly restored by loss of the H3.3 chaperone HIRA, while the H3.1 chaperone CAF-1 becomes essential. The single-particle cryo-EM structure at 3.2 Å of DDM1 with a variant nucleosome reveals engagement with histone H3.3 near residues required for assembly and with the unmodified H4 tail. An N-terminal autoinhibitory domain inhibits activity, while a disulfide bond in the helicase domain supports activity. DDM1 co-localizes with H3.1 and H3.3 during the cell cycle, and with the DNA methyltransferase MET1Dnmt1, but is blocked by H4K16 acetylation. The male germline H3.3 variant MGH3/HTR10 is resistant to remodeling by DDM1 and acts as a placeholder nucleosome in sperm cells for epigenetic inheritance.
Assuntos
Proteínas de Arabidopsis , Arabidopsis , Metilação de DNA , Histonas , Nucleossomos , Montagem e Desmontagem da Cromatina , DNA , Metilases de Modificação do DNA , Epigênese Genética , Histonas/genética , Nucleossomos/genética , Sêmen , Arabidopsis/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismoRESUMO
Gene regulation arises out of dynamic competition between nucleosomes, transcription factors, and other chromatin proteins for the opportunity to bind genomic DNA. The timescales of nucleosome assembly and binding of factors to DNA determine the outcomes of this competition at any given locus. Here, we review how these properties of chromatin proteins and the interplay between the dynamics of different factors are critical for gene regulation. We discuss how molecular structures of large chromatin-associated complexes, kinetic measurements, and high resolution mapping of protein-DNA complexes in vivo set the boundary conditions for chromatin dynamics, leading to models of how the steady state behaviors of regulatory elements arise.
Assuntos
Cromatina , Nucleossomos , Cromatina/genética , Montagem e Desmontagem da Cromatina , DNA/genética , DNA/metabolismo , Nucleossomos/genética , Fatores de Transcrição/genéticaRESUMO
Elucidating the global and local rules that govern genome-wide, hierarchical chromatin architecture remains a critical challenge. Current high-throughput chromosome conformation capture (Hi-C) technologies have identified large-scale chromatin structural motifs, such as topologically associating domains and looping. However, structural rules at the smallest or nucleosome scale remain poorly understood. Here, we coupled nucleosome-resolved Hi-C technology with simulated annealing-molecular dynamics (SA-MD) simulation to reveal 3D spatial distributions of nucleosomes and their genome-wide orientation in chromatin. Our method, called Hi-CO, revealed distinct nucleosome folding motifs across the yeast genome. Our results uncovered two types of basic secondary structural motifs in nucleosome folding: α-tetrahedron and ß-rhombus analogous to α helix and ß sheet motifs in protein folding. Using mutants and cell-cycle-synchronized cells, we further uncovered motifs with specific nucleosome positioning and orientation coupled to epigenetic features at individual loci. By illuminating molecular-level structure-function relationships in eukaryotic chromatin, our findings establish organizational principles of nucleosome folding.
Assuntos
Cromatina/ultraestrutura , Nucleossomos/ultraestrutura , Cromatina/genética , Cromatina/metabolismo , Montagem e Desmontagem da Cromatina/fisiologia , Cromossomos/metabolismo , Cromossomos/ultraestrutura , Nucleossomos/genética , Nucleossomos/metabolismo , Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/genética , Sítio de Iniciação de TranscriçãoRESUMO
Large-scale chromatin features, such as replication time and accessibility influence the rate of somatic and germline mutations at the megabase scale. This article reviews how local chromatin structures -e.g., DNA wrapped around nucleosomes, transcription factors bound to DNA- affect the mutation rate at a local scale. It dissects how the interaction of some mutagenic agents and/or DNA repair systems with these local structures influence the generation of mutations. We discuss how this local mutation rate variability affects our understanding of the evolution of the genomic sequence, and the study of the evolution of organisms and tumors.
Assuntos
Cromatina/genética , Genoma Humano/genética , Mutação/genética , Mapeamento Cromossômico/métodos , DNA/química , Reparo do DNA/genética , Evolução Molecular , Genômica , Mutação em Linhagem Germinativa/genética , Humanos , Mutagênese/genética , Taxa de Mutação , Nucleossomos/genética , Fatores de Transcrição/genéticaRESUMO
DNA N6-adenine methylation (6mA) has recently been described in diverse eukaryotes, spanning unicellular organisms to metazoa. Here, we report a DNA 6mA methyltransferase complex in ciliates, termed MTA1c. It consists of two MT-A70 proteins and two homeobox-like DNA-binding proteins and specifically methylates dsDNA. Disruption of the catalytic subunit, MTA1, in the ciliate Oxytricha leads to genome-wide loss of 6mA and abolishment of the consensus ApT dimethylated motif. Mutants fail to complete the sexual cycle, which normally coincides with peak MTA1 expression. We investigate the impact of 6mA on nucleosome occupancy in vitro by reconstructing complete, full-length Oxytricha chromosomes harboring 6mA in native or ectopic positions. We show that 6mA directly disfavors nucleosomes in vitro in a local, quantitative manner, independent of DNA sequence. Furthermore, the chromatin remodeler ACF can overcome this effect. Our study identifies a diverged DNA N6-adenine methyltransferase and defines the role of 6mA in chromatin organization.
Assuntos
Complexos Multienzimáticos/metabolismo , Nucleossomos/enzimologia , Oxytricha/enzimologia , Proteínas de Protozoários/metabolismo , DNA Metiltransferases Sítio Específica (Adenina-Específica)/metabolismo , Tetrahymena thermophila/enzimologia , Complexos Multienzimáticos/genética , Nucleossomos/genética , Oxytricha/genética , Proteínas de Protozoários/genética , DNA Metiltransferases Sítio Específica (Adenina-Específica)/genética , Tetrahymena thermophila/genéticaRESUMO
Chromatin domains and their associated structures must be faithfully inherited through cellular division to maintain cellular identity. However, accessing the localized strategies preserving chromatin domain inheritance, specifically the transfer of parental, pre-existing nucleosomes with their associated post-translational modifications (PTMs) during DNA replication, is challenging in living cells. We devised an inducible, proximity-dependent labeling system to irreversibly mark replication-dependent H3.1 and H3.2 histone-containing nucleosomes at desired loci in mouse embryonic stem cells so that their fate after DNA replication could be followed. Strikingly, repressed chromatin domains are preserved through local re-deposition of parental nucleosomes. In contrast, nucleosomes decorating active chromatin domains do not exhibit such preservation. Notably, altering cell fate leads to an adjustment of the positional inheritance of parental nucleosomes that reflects the corresponding changes in chromatin structure. These findings point to important mechanisms that contribute to parental nucleosome segregation to preserve cellular identity.
Assuntos
Montagem e Desmontagem da Cromatina/genética , Cromatina/genética , Epigênese Genética , Nucleossomos/genética , Animais , Diferenciação Celular/genética , Divisão Celular/genética , Linhagem da Célula/genética , Replicação do DNA/genética , Histonas/genética , Camundongos , Células-Tronco Embrionárias Murinas/metabolismo , Nucleossomos/metabolismo , Processamento de Proteína Pós-Traducional/genéticaRESUMO
Mutation rates along the genome are highly variable and influenced by several chromatin features. Here, we addressed how nucleosomes, the most pervasive chromatin structure in eukaryotes, affect the generation of mutations. We discovered that within nucleosomes, the somatic mutation rate across several tumor cohorts exhibits a strong 10 base pair (bp) periodicity. This periodic pattern tracks the alternation of the DNA minor groove facing toward and away from the histones. The strength and phase of the mutation rate periodicity are determined by the mutational processes active in tumors. We uncovered similar periodic patterns in the genetic variation among human and Arabidopsis populations, also detectable in their divergence from close species, indicating that the same principles underlie germline and somatic mutation rates. We propose that differential DNA damage and repair processes dependent on the minor groove orientation in nucleosome-bound DNA contribute to the 10-bp periodicity in AT/CG content in eukaryotic genomes.
Assuntos
DNA/genética , Mutação em Linhagem Germinativa , Taxa de Mutação , Nucleossomos/genética , Arabidopsis/genética , DNA/química , Sequência Rica em GC , Variação Genética , Conformação de Ácido Nucleico , Nucleossomos/químicaRESUMO
Histones serve to both package and organize DNA within the nucleus. In addition to histone post-translational modification and chromatin remodelling complexes, histone variants contribute to the complexity of epigenetic regulation of the genome. Histone variants are characterized by a distinct protein sequence and a selection of designated chaperone systems and chromatin remodelling complexes that regulate their localization in the genome. In addition, histone variants can be enriched with specific post-translational modifications, which in turn can provide a scaffold for recruitment of variant-specific interacting proteins to chromatin. Thus, through these properties, histone variants have the capacity to endow specific regions of chromatin with unique character and function in a regulated manner. In this Review, we provide an overview of recent advances in our understanding of the contribution of histone variants to chromatin function in mammalian systems. First, we discuss new molecular insights into chaperone-mediated histone variant deposition. Next, we discuss mechanisms by which histone variants influence chromatin properties such as nucleosome stability and the local chromatin environment both through histone variant sequence-specific effects and through their role in recruiting different chromatin-associated complexes. Finally, we focus on histone variant function in the context of both embryonic development and human disease, specifically developmental syndromes and cancer.
Assuntos
Cromatina/metabolismo , Histonas/genética , Histonas/metabolismo , Animais , DNA/metabolismo , Reparo do DNA/genética , Epigênese Genética/genética , Humanos , Chaperonas Moleculares/metabolismo , Nucleossomos/genética , Nucleossomos/metabolismo , Processamento de Proteína Pós-Traducional/genética , Fatores de Transcrição/metabolismo , Transcrição Gênica/fisiologiaRESUMO
In eukaryotic genomes, transcriptional machinery and nucleosomes compete for binding to DNA sequences; thus, a crucial aspect of gene regulatory element function is to modulate chromatin accessibility for transcription factor (TF) and RNA polymerase binding. Recent structural studies have revealed multiple modes of TF engagement with nucleosomes, but how initial "pioneering" results in steady-state DNA accessibility for further TF binding and RNA polymerase II (RNAPII) engagement has been unclear. Even less well understood is how distant sites of open chromatin interact with one another, such as when developmental enhancers activate promoters to release RNAPII for productive elongation. Here, we review evidence for the centrality of the conserved SWI/SNF family of nucleosome remodeling complexes, both in pioneering and in mediating enhancer-promoter contacts. Consideration of the nucleosome unwrapping and ATP hydrolysis activities of SWI/SNF complexes, together with their architectural features, may reconcile steady-state TF occupancy with rapid TF dynamics observed by live imaging.
Assuntos
Nucleossomos , Fatores de Transcrição , Nucleossomos/genética , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Cromatina , Proteínas de Ligação a DNA/genética , RNA Polimerase II/genética , RNA Polimerase II/metabolismo , Epigênese Genética , Montagem e Desmontagem da CromatinaRESUMO
Pioneer transcription factors (TFs) regulate cell fate by establishing transcriptionally primed and active states. However, cell fate control requires the coordination of both lineage-specific gene activation and repression of alternative-lineage programs, a process that is poorly understood. Here, we demonstrate that the pioneer TF FOXA coordinates with PRDM1 TF to recruit nucleosome remodeling and deacetylation (NuRD) complexes and Polycomb repressive complexes (PRCs), which establish highly occupied, accessible nucleosome conformation with bivalent epigenetic states, thereby preventing precocious and alternative-lineage gene expression during human endoderm differentiation. Similarly, the pioneer TF OCT4 coordinates with PRDM14 to form bivalent enhancers and repress cell differentiation programs in human pluripotent stem cells, suggesting that this may be a common and critical function of pioneer TFs. We propose that pioneer and PRDM TFs coordinate to safeguard cell fate through epigenetic repression mechanisms.
Assuntos
Nucleossomos , Fatores de Transcrição , Humanos , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Nucleossomos/genética , Diferenciação Celular/genética , Proteínas do Grupo Polycomb/metabolismo , Epigênese GenéticaRESUMO
Polycomb repressive complex 1 (PRC1) is a key transcriptional regulator in development via modulating chromatin structure and catalyzing histone H2A ubiquitination at Lys119 (H2AK119ub1). H2AK119ub1 is one of the most abundant histone modifications in mammalian cells. However, the function of H2AK119ub1 in polycomb-mediated gene silencing remains debated. In this study, we reveal that H2AK119ub1 has two distinct roles in gene expression, through differentially modulating chromatin compaction mediated by canonical PRC1 and the linker histone H1. Interestingly, we find that H2AK119ub1 plays a positive role in transcription through interfering with the binding of canonical PRC1 to nucleosomes and therefore counteracting chromatin condensation. Conversely, we demonstrate that H2AK119ub1 facilitates H1-dependent chromatin condensation and enhances the silencing of developmental genes in mouse embryonic stem cells, suggesting that H1 may be one of several possible pathways for H2AK119ub1 in repressing transcription. These results provide insights and molecular mechanisms by which H2AK119ub1 differentially fine-tunes developmental gene expression.
Assuntos
Cromatina , Complexo Repressor Polycomb 1 , Animais , Camundongos , Cromatina/genética , Complexo Repressor Polycomb 1/genética , Complexo Repressor Polycomb 1/metabolismo , Nucleossomos/genética , Ubiquitinação , Expressão Gênica , Mamíferos/metabolismoRESUMO
Polycomb repressive complexes (PRCs) play a key role in gene repression and are indispensable for proper development. Canonical PRC1 forms condensates in vitro and in cells that are proposed to contribute to the maintenance of repression. However, how chromatin and the various subunits of PRC1 contribute to condensation is largely unexplored. Using a reconstitution approach and single-molecule imaging, we demonstrate that nucleosomal arrays and PRC1 act synergistically, reducing the critical concentration required for condensation by more than 20-fold. We find that the exact combination of PHC and CBX subunits determines condensate initiation, morphology, stability, and dynamics. Particularly, PHC2's polymerization activity influences condensate dynamics by promoting the formation of distinct domains that adhere to each other but do not coalesce. Live-cell imaging confirms CBX's role in condensate initiation and highlights PHC's importance for condensate stability. We propose that PRC1 composition can modulate condensate properties, providing crucial regulatory flexibility across developmental stages.
Assuntos
Proteínas de Ciclo Celular , Cromatina , Nucleossomos , Complexo Repressor Polycomb 1 , Complexo Repressor Polycomb 1/metabolismo , Complexo Repressor Polycomb 1/genética , Cromatina/metabolismo , Cromatina/genética , Humanos , Nucleossomos/metabolismo , Nucleossomos/genética , Animais , Imagem Individual de MoléculaRESUMO
RNF168 plays a central role in the DNA damage response (DDR) by ubiquitylating histone H2A at K13 and K15. These modifications direct BRCA1-BARD1 and 53BP1 foci formation in chromatin, essential for cell-cycle-dependent DNA double-strand break (DSB) repair pathway selection. The mechanism by which RNF168 catalyzes the targeted accumulation of H2A ubiquitin conjugates to form repair foci around DSBs remains unclear. Here, using cryoelectron microscopy (cryo-EM), nuclear magnetic resonance (NMR) spectroscopy, and functional assays, we provide a molecular description of the reaction cycle and dynamics of RNF168 as it modifies the nucleosome and recognizes its ubiquitylation products. We demonstrate an interaction of a canonical ubiquitin-binding domain within full-length RNF168, which not only engages ubiquitin but also the nucleosome surface, clarifying how such site-specific ubiquitin recognition propels a signal amplification loop. Beyond offering mechanistic insights into a key DDR protein, our study aids in understanding site specificity in both generating and interpreting chromatin ubiquitylation.
Assuntos
Nucleossomos , Ubiquitina-Proteína Ligases , Nucleossomos/genética , Microscopia Crioeletrônica , Ubiquitina-Proteína Ligases/metabolismo , Ubiquitinação , Histonas/metabolismo , Cromatina/genética , Reparo do DNA , Ubiquitina/metabolismo , Proteína 1 de Ligação à Proteína Supressora de Tumor p53/genética , Dano ao DNARESUMO
Polycomb repressive complex 2 (PRC2) is an epigenetic regulator that trimethylates lysine 27 of histone 3 (H3K27me3) and is essential for embryonic development and cellular differentiation. H3K27me3 is associated with transcriptionally repressed chromatin and is established when PRC2 is allosterically activated upon methyl-lysine binding by the regulatory subunit EED. Automethylation of the catalytic subunit enhancer of zeste homolog 2 (EZH2) stimulates its activity by an unknown mechanism. Here, we show that human PRC2 forms a dimer on chromatin in which an inactive, automethylated PRC2 protomer is the allosteric activator of a second PRC2 that is poised to methylate H3 of a substrate nucleosome. Functional assays support our model of allosteric trans-autoactivation via EED, suggesting a previously unknown mechanism mediating context-dependent activation of PRC2. Our work showcases the molecular mechanism of auto-modification-coupled dimerization in the regulation of chromatin-modifying complexes.
Assuntos
Cromatina , Proteína Potenciadora do Homólogo 2 de Zeste , Histonas , Complexo Repressor Polycomb 2 , Multimerização Proteica , Complexo Repressor Polycomb 2/metabolismo , Complexo Repressor Polycomb 2/genética , Humanos , Cromatina/metabolismo , Cromatina/genética , Histonas/metabolismo , Histonas/genética , Metilação , Proteína Potenciadora do Homólogo 2 de Zeste/metabolismo , Proteína Potenciadora do Homólogo 2 de Zeste/genética , Nucleossomos/metabolismo , Nucleossomos/genética , Regulação Alostérica , Células HEK293 , Ligação ProteicaRESUMO
Nucleosomes, the basic structural units of chromatin, hinder recruitment and activity of various DNA repair proteins, necessitating modifications that enhance DNA accessibility. Poly(ADP-ribosyl)ation (PARylation) of proteins near damage sites is an essential initiation step in several DNA-repair pathways; however, its effects on nucleosome structural dynamics and organization are unclear. Using NMR, cryoelectron microscopy (cryo-EM), and biochemical assays, we show that PARylation enhances motions of the histone H3 tail and DNA, leaving the configuration of the core intact while also stimulating nuclease digestion and ligation of nicked nucleosomal DNA by LIG3. PARylation disrupted interactions between nucleosomes, preventing self-association. Addition of LIG3 and XRCC1 to PARylated nucleosomes generated condensates that selectively partition DNA repair-associated proteins in a PAR- and phosphorylation-dependent manner in vitro. Our results establish that PARylation influences nucleosomes across different length scales, extending from the atom-level motions of histone tails to the mesoscale formation of condensates with selective compositions.
Assuntos
Nucleossomos , Poli ADP Ribosilação , Nucleossomos/genética , Poli ADP Ribosilação/genética , Poli(ADP-Ribose) Polimerases/metabolismo , Microscopia Crioeletrônica , Condensados Biomoleculares , Reparo do DNA , Histonas/genética , Histonas/metabolismo , DNA/genética , DNA/metabolismo , Dano ao DNA , Poli(ADP-Ribose) Polimerase-1/metabolismoRESUMO
In this issue of Molecular Cell, Engeholm et al.1 present cryo-EM structures of the chromatin remodeler Chd1 bound to a hexasome-nucleosome complex, an intermediate state during transcription either with or without FACT to restore the missing H2A-H2B dimer. These two binding modes reveal how Chd1 and FACT cooperate in nucleosome re-establishment during transcription.
Assuntos
Microscopia Crioeletrônica , Proteínas de Ligação a DNA , Nucleossomos , Nucleossomos/metabolismo , Nucleossomos/genética , Proteínas de Ligação a DNA/metabolismo , Proteínas de Ligação a DNA/genética , Montagem e Desmontagem da Cromatina , Histonas/metabolismo , Histonas/genética , Humanos , Proteínas de Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/genética , DNA Helicases/metabolismo , DNA Helicases/genética , Ligação Proteica , Transcrição Gênica , Proteínas de Grupo de Alta Mobilidade/metabolismo , Proteínas de Grupo de Alta Mobilidade/genética , Fatores de Elongação da Transcrição/metabolismo , Fatores de Elongação da Transcrição/genética , Fatores de Elongação da Transcrição/químicaRESUMO
Mammalian gene expression is controlled by transcription factors (TFs) that engage sequence motifs in a chromatinized genome, where nucleosomes can restrict DNA access. Yet, how nucleosomes affect individual TFs remains unclear. Here, we measure the ability of over one hundred TF motifs to recruit TFs in a defined chromosomal locus in mouse embryonic stem cells. This identifies a set sufficient to enable the binding of TFs with diverse tissue specificities, functions, and DNA-binding domains. These chromatin-competent factors are further classified when challenged to engage motifs within a highly phased nucleosome. The pluripotency factors OCT4-SOX2 preferentially engage non-nucleosomal and entry-exit motifs, but not nucleosome-internal sites, a preference that also guides binding genome wide. By contrast, factors such as BANP, REST, or CTCF engage throughout, causing nucleosomal displacement. This supports that TFs vary widely in their sensitivity to nucleosomes and that genome access is TF specific and influenced by nucleosome position in the cell.
Assuntos
Células-Tronco Embrionárias Murinas , Nucleossomos , Fatores de Transcrição , Nucleossomos/metabolismo , Nucleossomos/genética , Animais , Camundongos , Fatores de Transcrição/metabolismo , Fatores de Transcrição/genética , Células-Tronco Embrionárias Murinas/metabolismo , Sítios de Ligação , Ligação Proteica , Genoma/genética , Fator 3 de Transcrição de Octâmero/metabolismo , Fator 3 de Transcrição de Octâmero/genética , Cromatina/metabolismo , Cromatina/genética , Fatores de Transcrição SOXB1/genética , Fatores de Transcrição SOXB1/metabolismo , Montagem e Desmontagem da CromatinaRESUMO
Alternative transcription start sites can affect transcript isoform diversity and translation levels. In a recently described form of gene regulation, coordinated transcriptional and translational interference results in transcript isoform-dependent changes in protein expression. Specifically, a long undecoded transcript isoform (LUTI) is transcribed from a gene-distal promoter, interfering with expression of the gene-proximal promoter. Although transcriptional and chromatin features associated with LUTI expression have been described, the mechanism underlying LUTI-based transcriptional interference is not well understood. Using an unbiased genetic approach followed by functional genomics, we uncovered that the Swi/Snf chromatin remodeling complex is required for co-transcriptional nucleosome remodeling that leads to LUTI-based repression. We identified genes with tandem promoters that rely on Swi/Snf function for transcriptional interference during protein folding stress, including LUTI-regulated genes. This study provides clear evidence for Swi/Snf playing a direct role in gene repression via a cis transcriptional interference mechanism.
Assuntos
Montagem e Desmontagem da Cromatina , Proteínas Cromossômicas não Histona , Nucleossomos , Regiões Promotoras Genéticas , Proteínas de Saccharomyces cerevisiae , Saccharomyces cerevisiae , Fatores de Transcrição , Transcrição Gênica , Fatores de Transcrição/metabolismo , Fatores de Transcrição/genética , Proteínas de Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismo , Proteínas Cromossômicas não Histona/genética , Proteínas Cromossômicas não Histona/metabolismo , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Nucleossomos/metabolismo , Nucleossomos/genética , Regulação Fúngica da Expressão Gênica , Sítio de Iniciação de Transcrição , Cromatina/metabolismo , Cromatina/genéticaRESUMO
Heterochromatin enforces transcriptional gene silencing and can be epigenetically inherited, but the underlying mechanisms remain unclear. Here, we show that histone deacetylation, a conserved feature of heterochromatin domains, blocks SWI/SNF subfamily remodelers involved in chromatin unraveling, thereby stabilizing modified nucleosomes that preserve gene silencing. Histone hyperacetylation, resulting from either the loss of histone deacetylase (HDAC) activity or the direct targeting of a histone acetyltransferase to heterochromatin, permits remodeler access, leading to silencing defects. The requirement for HDAC in heterochromatin silencing can be bypassed by impeding SWI/SNF activity. Highlighting the crucial role of remodelers, merely targeting SWI/SNF to heterochromatin, even in cells with functional HDAC, increases nucleosome turnover, causing defective gene silencing and compromised epigenetic inheritance. This study elucidates a fundamental mechanism whereby histone hypoacetylation, maintained by high HDAC levels in heterochromatic regions, ensures stable gene silencing and epigenetic inheritance, providing insights into genome regulatory mechanisms relevant to human diseases.
Assuntos
Montagem e Desmontagem da Cromatina , Epigênese Genética , Inativação Gênica , Heterocromatina , Histona Desacetilases , Histonas , Nucleossomos , Heterocromatina/metabolismo , Heterocromatina/genética , Nucleossomos/metabolismo , Nucleossomos/genética , Histonas/metabolismo , Histonas/genética , Acetilação , Histona Desacetilases/metabolismo , Histona Desacetilases/genética , Humanos , Histona Acetiltransferases/metabolismo , Histona Acetiltransferases/genética , AnimaisRESUMO
RNA polymerases must initiate and pause within a complex chromatin environment, surrounded by nucleosomes and other transcriptional machinery. This environment creates a spatial arrangement along individual chromatin fibers ripe for both competition and coordination, yet these relationships remain largely unknown owing to the inherent limitations of traditional structural and sequencing methodologies. To address this, we employed long-read chromatin fiber sequencing (Fiber-seq) in Drosophila to visualize RNA polymerase (Pol) within its native chromatin context with single-molecule precision along up to 30 kb fibers. We demonstrate that Fiber-seq enables the identification of individual Pol II, nucleosome, and transcription factor footprints, revealing Pol II pausing-driven destabilization of downstream nucleosomes. Furthermore, we demonstrate pervasive direct distance-dependent transcriptional coupling between nearby Pol II genes, Pol III genes, and transcribed enhancers, modulated by local chromatin architecture. Overall, transcription initiation reshapes surrounding nucleosome architecture and couples nearby transcriptional machinery along individual chromatin fibers.