Unusual nucleosome formation and transcriptome influence by the histone H3mm18 variant
Ontology highlight
ABSTRACT:
ORGANISM(S): Mus musculus
PROVIDER: GSE168238 | GEO | 2022/01/03
REPOSITORIES: GEO
Similar Datasets
Project description:Histone H3mm18 is a non-allelic H3 variant expressed in skeletal muscle and brain in mice. However, its function has remained enigmatic. We found that H3mm18 is incorporated into chromatin in cells with low efficiency, as compared to H3.3. We determined the structures of the nucleosome core particle (NCP) containing H3mm18 by cryo-electron microscopy, which revealed that the entry/exit DNA regions are drastically disordered in the H3mm18 NCP. Consistently, the H3mm18 NCP is substantially unstable in vitro. The forced expression of H3mm18 in mouse myoblast C2C12 cells markedly suppressed muscle differentiation. A transcriptome analysis revealed that the forced expression of H3mm18 affected the expression of multiple genes, and suppressed a group of genes involved in muscle development. These results suggest a novel gene expression regulation system in which the chromatin landscape is altered by the formation of unusual nucleosomes with a histone variant, H3mm18, and provide important insight into understanding transcription regulation by chromatin.
| S-EPMC8855299 | biostudies-literature
Project description:Unusual nucleosome formation and transcriptome influence by the histone H3mm18 variant
| PRJNA706656 | ENA
Project description:Linker histones are involved in the formation of higher-order chromatin structure. Although linker histones have been implicated in the regulation of specific genes, it still remains unclear what their principal binding determinants are and how their repressive function in vitro can be reconciled with presumed broad binding in vivo. We generated a full genome, high resolution binding map of linker histone H1 in Drosophila Kc cells, using DamID. H1 binds at similar levels across much of the genome, both in classical euchromatin and heterochromatin. Strikingly, there are pronounced dips of low H1 occupancy around transcription start sites of active genes and at many distant cis-regulatory sites. H1 dips are not due to lack of nucleosomes. Rather, all regions with low binding of H1 show enrichment of the histone variant H3.3 which itself has been linked to high nucleosome turnover. Upon knockdown of H3.3, we find that H1 levels increase at sites previously not covered with H1 with a concomitant increase in nucleosome repeat length. These changes are independent of transcriptional changes. Our results show that the H3.3 protein counteracts association of H1 at genomic sites with high rates of histone turnover. This antagonism provides a mechanism to keep diverse genomic sites in an open chromatin conformation. For this study, we generated DamID profiles of histone H1 and RpII18 in Drosophila Kc167 cells. Additionally, we generated H1 profiles in cells treated with RNAi against white, H3.3B, or H3.3A and H3.3B. Nucleosome occupancy profiles were generated in untreated cells and cells treated with RNAi against white or H3.3A and H3.3B. Profiles of expression changes were generated for H3.3B RNAi and H3.3A and H3.3B RNAi. DamID experiments for H1 and RpII18 were performed in Drosophila cell cultures. Samples were hybridized to 380k NimbleGen arrays with 300 bp probe spacing. Formaldehyde-assisted isolation of regulatotry elements (FAIRE) was performed in Drosophila Kc167 cells. Samples were hybridized to 380k NimbleGen arrays with 300 bp probe spacing over non-crosslinked genomic DNA. Nucleosome positioning profiles were made by hybridizing mononucleosomal DNA over MNase digested purified genomic DNA on 380k NimbleGen arrays with 10 bp probe spacing. Expression profiles were made as H3.3 RNAi over white RNAi cohybridizations on spotted INDAC long oligo arrays. Every experiment was done in duplicate in the reverse dye orientation.
2010-06-23 | E-GEOD-16882 | biostudies-arrayexpress
Project description:Linker histones are involved in the formation of higher-order chromatin structure. Although linker histones have been implicated in the regulation of specific genes, it still remains unclear what their principal binding determinants are and how their repressive function in vitro can be reconciled with presumed broad binding in vivo. We generated a full genome, high resolution binding map of linker histone H1 in Drosophila Kc cells, using DamID. H1 binds at similar levels across much of the genome, both in classical euchromatin and heterochromatin. Strikingly, there are pronounced dips of low H1 occupancy around transcription start sites of active genes and at many distant cis-regulatory sites. H1 dips are not due to lack of nucleosomes. Rather, all regions with low binding of H1 show enrichment of the histone variant H3.3 which itself has been linked to high nucleosome turnover. Upon knockdown of H3.3, we find that H1 levels increase at sites previously not covered with H1 with a concomitant increase in nucleosome repeat length. These changes are independent of transcriptional changes. Our results show that the H3.3 protein counteracts association of H1 at genomic sites with high rates of histone turnover. This antagonism provides a mechanism to keep diverse genomic sites in an open chromatin conformation. For this study, we generated DamID profiles of histone H1 and RpII18 in Drosophila Kc167 cells. Additionally, we generated H1 profiles in cells treated with RNAi against white, H3.3B, or H3.3A and H3.3B. Nucleosome occupancy profiles were generated in untreated cells and cells treated with RNAi against white or H3.3A and H3.3B. Profiles of expression changes were generated for H3.3B RNAi and H3.3A and H3.3B RNAi.
2009-10-15 | GSE16882 | GEO
Project description:Nucleosome is the basic structural unit of chromatin, and its dynamics plays critical roles in the regulation of genome functions. However, how the nucleosome structure is regulated by histone variants in vivo is still largely uncharacterized. Here, by employing Micrococcal nuclease (MNase) digestion of crosslinked chromatin followed by chromatin immunoprecipitation (ChIP) and paired-end sequencing (MNase-X-ChIP-seq), we mapped genome-wide unwrapping states of nucleosomes containing histone variant H2A.Z in mouse embryonic stem (ES) cells. We found that H2A.Z is enriched with unwrapped nucleosomes. Interestingly, the function of +1 H2A.Z nucleosome in transcriptional regulation is correlated with the unwrapping states. We further showed that H2A.Z nucleosomes adjacent the CTCF binding sites (CBS) may adopt an open conformation. We confirmed the unwrapping state of H2A.Z nucleosomes under native condition by re-ChIP of H2A.Z after CTCF CUT&RUN in mouse ES cells. Importantly, we found that depletion of H2A.Z results in increased CTCF binding, indicating dynamic competition between the unwrapped H2A.Z nucleosomal intermediates and CTCF at the CBS. Taken together, our results showed that histone variant H2A.Z regulates transcription and CTCF binding through modulating the nucleosome unwrapping.
2020-04-27 | GSE146082 | GEO
Project description:In this work we have studied the properties of the novel mouse histone variant H2AL2. H2AL2 was used to reconstitute nucleosomes and the structural and functional properties of these particles were studied by a combination of biochemical approaches, atomic force microscopy (AFM) and electron cryo-microscopy. DNase I and hydroxyl radical footprinting as well as micrococcal and exonuclease III digestion demonstrated an altered structure of the H2AL2 nucleosomes all over the nucleosomal DNA length. Restriction nuclease accessibility experiments revealed that the interactions of the H2AL2 histone octamer with the ends of the nucleosomal DNA are highly perturbed. AFM imaging showed that the H2AL2 histone octamer was complexed with only approximately 130 bp of DNA. H2AL2 reconstituted trinucleosomes exhibited a type of a 'beads on a string' structure, which was quite different from the equilateral triangle 3D organization of conventional H2A trinucleosomes. The presence of H2AL2 affected both the RSC and SWI/SNF remodeling and mobilization of the variant particles. These unusual properties of the H2AL2 nucleosomes suggest a specific role of H2AL2 during mouse spermiogenesis.
| S-EPMC2724287 | biostudies-literature
Project description:Maintaining the dynamic structure of chromatin is critical for regulating the cellular processes that require access to the DNA template, such as DNA damage repair, transcription, and replication. Histone chaperones and ATP-dependent chromatin remodeling factors facilitate transitions in chromatin structure by assembling and positioning nucleosomes through a variety of enzymatic activities. SMARCAD1 is a unique chromatin remodeler that combines the ATP-dependent ability to exchange histones, with the chaperone-like activity of nucleosome deposition. We have shown previously that phosphorylated SMARCAD1 exhibits reduced binding to nucleosomes. However, it is unknown how phosphorylation affects SMARCAD1’s ability to perform its various enzymatic activities. Here we use mutational analysis, activity assays, and mass spectrometry, to probe SMARCAD1 regulation and to investigate the role of its flexible N-terminal region. We show that phosphorylation affects SMARCAD1 binding to nucleosomes and DNA, as well as ATP hydrolysis and histone exchange. Conversely, we report that phosphorylation is inconsequential for histone binding and nucleosome assembly. In addition, the SMARCAD1 N-terminal region is revealed to be critical for nucleosome assembly and histone exchange. Together, this work examines the intricacies of how phosphorylation governs SMARCAD1 activity and provides insight into its complex regulation and diverse activities.Maintaining the dynamic structure of chromatin is critical for regulating the cellular processes that require access to the DNA template, such as DNA damage repair, transcription, and replication. Histone chaperones and ATP-dependent chromatin remodeling factors facilitate transitions in chromatin structure by assembling and positioning nucleosomes through a variety of enzymatic activities. SMARCAD1 is a unique chromatin remodeler that combines the ATP-dependent ability to exchange histones, with the chaperone-like activity of nucleosome deposition. We have shown previously that phosphorylated SMARCAD1 exhibits reduced binding to nucleosomes. However, it is unknown how phosphorylation affects SMARCAD1’s ability to perform its various enzymatic activities. Here we use mutational analysis, activity assays, and mass spectrometry, to probe SMARCAD1 regulation and to investigate the role of its flexible N-terminal region. We show that phosphorylation affects SMARCAD1 binding to nucleosomes and DNA, as well as ATP hydrolysis and histone exchange. Conversely, we report that phosphorylation is inconsequential for histone binding and nucleosome assembly. In addition, the SMARCAD1 N-terminal region is revealed to be critical for nucleosome assembly and histone exchange. Together, this work examines the intricacies of how phosphorylation governs SMARCAD1 activity and provides insight into its complex regulation and diverse activities.
2025-05-07 | PXD055413 | Pride
Project description:BAF and PBAF are mammalian SWI/SNF family chromatin remodeling complexes that possess multiple histone/DNA-binding subunits and create nucleosome-depleted/free regions for transcription activation. Despite previous structural studies and recent advance of SWI/SNF family complexes, it remains incompletely understood how PBAF-nucleosome complex is organized. Here we determined structure of 13-subunit human PBAF in complex with acetylated nucleosome in ADP-BeF3-bound state. Four PBAF-specific subunits work together with nine BAF/PBAF-shared subunits to generate PBAF-specific modular organization, distinct from that of BAF at various regions. PBAF-nucleosome structure reveals six histone-binding domains and four DNA-binding domains/modules, the majority of which directly bind histone/DNA. This multivalent nucleosome-binding pattern, not observed in previous studies, suggests that PBAF may integrate comprehensive chromatin information to target genomic loci for function. Our study reveals molecular organization of subunits and histone/DNA-binding domains/modules in PBAF-nucleosome complex and provides structural insights into PBAF-mediated nucleosome association complimentary to the recently reported PBAF-nucleosome structure.
2023-03-11 | PXD036708 | Pride
Project description:Maintaining the dynamic structure of chromatin is critical for regulating the cellular processes that require access to the DNA template, such as DNA damage repair, transcription, and replication. Histone chaperones and ATP-dependent chromatin remodeling factors facilitate transitions in chromatin structure by assembling and positioning nucleosomes through a variety of enzymatic activities. SMARCAD1 is a unique chromatin remodeler that combines the ATP-dependent ability to exchange histones, with the chaperone-like activity of nucleosome deposition. We have shown previously that phosphorylated SMARCAD1 exhibits reduced binding to nucleosomes. However, it is unknown how phosphorylation affects SMARCAD1’s ability to perform its various enzymatic activities. Here we use mutational analysis, activity assays, and mass spectrometry, to probe SMARCAD1 regulation and to investigate the role of its flexible N-terminal region. We show that phosphorylation affects SMARCAD1 binding to nucleosomes and DNA, as well as ATP hydrolysis and histone exchange. Conversely, we report that phosphorylation is inconsequential for histone binding and nucleosome assembly. In addition, the SMARCAD1 N-terminal region is revealed to be critical for nucleosome assembly and histone exchange. Together, this work examines the intricacies of how phosphorylation governs SMARCAD1 activity and provides insight into its complex regulation and diverse activities.
2025-05-07 | PXD055412 | Pride
Project description:Maintaining the dynamic structure of chromatin is critical for regulating the cellular processes that require access to the DNA template, such as DNA damage repair, transcription, and replication. Histone chaperones and ATP-dependent chromatin remodeling factors facilitate transitions in chromatin structure by assembling and positioning nucleosomes through a variety of enzymatic activities. SMARCAD1 is a unique chromatin remodeler that combines the ATP-dependent ability to exchange histones, with the chaperone-like activity of nucleosome deposition. We have shown previously that phosphorylated SMARCAD1 exhibits reduced binding to nucleosomes. However, it is unknown how phosphorylation affects SMARCAD1’s ability to perform its various enzymatic activities. Here we use mutational analysis, activity assays, and mass spectrometry, to probe SMARCAD1 regulation and to investigate the role of its flexible N-terminal region. We show that phosphorylation affects SMARCAD1 binding to nucleosomes and DNA, as well as ATP hydrolysis and histone exchange. Conversely, we report that phosphorylation is inconsequential for histone binding and nucleosome assembly. In addition, the SMARCAD1 N-terminal region is revealed to be critical for nucleosome assembly and histone exchange. Together, this work examines the intricacies of how phosphorylation governs SMARCAD1 activity and provides insight into its complex regulation and diverse activities.
2025-05-07 | PXD055414 | Pride