FACT Complex Recruitment and the Role of H2A.Z in Transcriptional Regulation [ChIP-seq]
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ABSTRACT:
PROVIDER: PRJNA1272464 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| Other | ||||
| SRR33839305.fastq.gz | Fastqsanger.gz | |||
| SRR33839306.fastq.gz | Fastqsanger.gz | |||
| SRR33839307.fastq.gz | Fastqsanger.gz | |||
| SRR33839308.fastq.gz | Fastqsanger.gz |
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Project description:The FACT complex (FAcilitates Chromatin Transcription) plays a key role in chromatin remodeling during transcription, enhancing DNA accessibility. It is believed to alter chromatin by temporarily displacing histones, enabling RNA polymerase II (Pol II) to engage, and then reinstating the original chromatin structure after transcription is complete. In this study, we show that FACT complex subunit ssrp1 undergoes neddylation in vivo, which might be important for its subcellular localization and its interaction with Pol II. This, in turn, promotes the recruitment of the FACT complex to actively transcribed chromatin, establishing a positive feedback loop. In addition, the FACT complex mediates H3K4me3 while not influencing the transcriptional rate. Lastly, in contrast to its canonical form, ubiquitinated H2A.Z does not impede FACT's binding to chromatin and may alleviate the transcriptional repression associated with canonical H2A ubiquitination.
2025-06-08 | GSE299054 | GEO
Project description:The FACT complex (FAcilitates Chromatin Transcription) plays a key role in chromatin remodeling during transcription, enhancing DNA accessibility. It is believed to alter chromatin by temporarily displacing histones, enabling RNA polymerase II (Pol II) to engage, and then reinstating the original chromatin structure after transcription is complete. In this study, we show that FACT complex subunit ssrp1 undergoes neddylation in vivo, which might be important for its subcellular localization and its interaction with Pol II. This, in turn, promotes the recruitment of the FACT complex to actively transcribed chromatin, establishing a positive feedback loop. In addition, the FACT complex mediates H3K4me3 while not influencing the transcriptional rate. Lastly, in contrast to its canonical form, ubiquitinated H2A.Z does not impede FACT's binding to chromatin and may alleviate the transcriptional repression associated with canonical H2A ubiquitination.
2025-06-08 | GSE299053 | GEO
Project description:FACT Complex Recruitment and the Role of H2A.Z in Transcriptional Regulation [RNA-seq]
| PRJNA1272465 | ENA
Project description:H2A.Z is a highly conserved histone variant involved in several key nuclear processes. It is incorporated into promoters by SWR-C-related chromatin remodeling complexes, but whether it is also actively excluded from non-promoter regions is not clear. Here, we provide genomic and biochemical evidence that RNA polymerase II (RNAPII) elongation-associated histone chaperones FACT and Spt6 both contribute to restricting H2A.Z from intragenic regions. In the absence of FACT or Spt6, the lack of H2A.Z eviction, coupled to its pervasive incorporation by mislocalized SWR-C, alters chromatin composition and contributes to cryptic initiation. Thus, chaperone-mediated H2A.Z removal is crucial for restricting the chromatin signature of gene promoters, which otherwise may license or promote cryptic transcription. We profiled H2A.Z occupancy in S. cerevisiae by ChIP-chip on tiling arrays in different mutants for chromatin remodelers and histone chaperones. In most experiments, H2A.Z ChIP samples (Cy5-labeled) were performed using a polyclonal antibody against H2A.Z and hybridized against H2B ChIP samples (Cy3-labeled), also performed using a polyclonal antibody. Temperature-sensitive mutants for the histone chaperones Spt16 and Spt6 (spt16-197 and spt6-1004 respectively) showed strong H2A.Z localization defects so the role of these factors in H2A.Z localization was further analyzed. H2A.Z ChIP-chip experiments in spt16-197 and spt6-1004 were repeated using different experimental designs [normalized against Input DNA or Mock (IgG) ChIP samples]. The contribution of Spt16 and Spt6 on H2A.Z localization was also confirmed by nuclear depletion of Spt16 and Spt6 using the anchor-away system. H2A.Z ChIP-chip experiments were also performed in sic1Δ and spt16-197/sic1Δ cells in order to rule out any G1-arrest artifact. H2A.Z ChIP-chip experiments were repeated using spike-in controls for normalization, revealing widespread H2A.Z occupancy in Spt16 and Spt6 mutants. In addition, the localization of the chromatin remodeler SWR-C was determined in wild type cells as well as in spt16-197 and spt6-1004 cells. Finally, we also profiled histone H4 occupancy by ChIP-chip in wild type, spt16-197, spt6-1004, spt16-197/htz1Δ and spt6-1004/htz1Δ cells. All ChIP-chip experiments were done in duplicates. Each microarray was normalized using the Lima Loess and replicates were combined using a weighted average method as previously described (Pokholok et al., 2005).
2015-05-12 | E-GEOD-62880 | biostudies-arrayexpress
Project description:H2A.Z is a highly conserved histone variant involved in several key nuclear processes. It is incorporated into promoters by SWR-C-related chromatin remodeling complexes, but whether it is also actively excluded from non-promoter regions is not clear. Here, we provide genomic and biochemical evidence that RNA polymerase II (RNAPII) elongation-associated histone chaperones FACT and Spt6 both contribute to restricting H2A.Z from intragenic regions. In the absence of FACT or Spt6, the lack of H2A.Z eviction, coupled to its pervasive incorporation by mislocalized SWR-C, alters chromatin composition and contributes to cryptic initiation. Thus, chaperone-mediated H2A.Z removal is crucial for restricting the chromatin signature of gene promoters, which otherwise may license or promote cryptic transcription.
2015-05-12 | GSE62880 | GEO
Project description:The H2A.Z histone variant plays major roles in the control of gene expression. In human, H2A.Z is encoded by two genes expressing two isoforms, H2A.Z.1 and H2A.Z.2 differing by three amino acids. Here, we undertook an integrated analysis of the independent and interdependent functions of these two isoforms in gene expression using endogenously-tagged isoform. RNA-Seq analysis following depletion of either isoform or both together in untransformed cells showed that they can regulate both distinct and overlapping sets of genes positively or negatively in a context-dependent manner. Our data revealed that the two isoforms have similar or antagonistic function depending on the gene. H2A.Z.1 and H2A.Z.2 can replace each other at Transcription Start Sites, providing a molecular explanation for this interplay. Mass spectrometry analysis showed that H2A.Z.1 and H2A.Z.2 have specific interactors, PHF14 and associated proteins for H2A.Z.1 and the histone deacetylase SIRT1 for H2A.Z.2. Moreover, we show that PHF14 and SIRT1 mediate the functional antagonism between H2A.Z.1 and H2A.Z.2. In conclusion, we propose a model in which the balance between H2A.Z.1 and H2A.Z.2 at promoters is critically important to regulate specific gene expression and depends on the recruitment of specific proteins. Our work thus provides an additional layer of complexity to the control of gene expression by histone variants.
2020-02-27 | GSE131578 | GEO
Project description:A cascade of histone acetylation events with subsequent incorporation of a histone H2A variant plays an essential part in transcription regulation in various model organisms. A key player in this cascade is the chromatin remodellling complex SWR1, which replaces the canonical histone H2A with its variant H2A.Z. Transcriptional regulation of polycistronic transcription units in the unicellular parasite Trypanosoma brucei has been shown to be highly dependent on acetylation of H2A.Z, which is mediated by the histone-acetyltransferase HAT2. The chromatin remodellling complex, which mediates H2A.Z incorporation is not known and an SWR1 orthologue in trypanosomes has not yet been reported. In this study, we identified and characterised an SWR1-like remodelller complex in T. brucei that is responsible for Pol II-dependent transcriptional regulation. Bioinformatic analysis of potential SNF2 DEAD/Box helicases, the key component of SWR1 complexes, identified a 1211 amino acids-long protein that exhibits key structural characteristics of the SWR1 subfamily. Systematic protein-protein interaction analysis revealed the existence of a novel complex exhibiting key features of an SWR1-like chromatin remodelller. RNAi-mediated depletion of the ATPase subunit of this complex resulted in a significant reduction of H2A.Z incorporation at transcription start sites and a subsequent decrease of steady-state mRNA levels. Furthermore, depletion of SWR1 and RNA-polymerase II (Pol II) caused massive chromatin condensation. The potential function of several proteins associated with the SWR1-like complex and with HAT2, the key factor of H2A.Z incorporation, is discussed.
2022-08-12 | PXD026796 | Pride
Project description:The H2A.Z histone variant plays major roles in the control of gene expression. In human, H2A.Z is encoded by two genes expressing two isoforms, H2A.Z.1 and H2A.Z.2 differing by three amino acids. Here, we undertook an integrated analysis of the independent and interdependent functions of these two isoforms in gene expression using endogenously-tagged isoform. RNA-Seq analysis following depletion of either isoform or both together in untransformed cells showed that they can regulate both distinct and overlapping sets of genes positively or negatively in a context-dependent manner. Our data revealed that the two isoforms have similar or antagonistic function depending on the gene. H2A.Z.1 and H2A.Z.2 can replace each other at Transcription Start Sites, providing a molecular explanation for this interplay. Mass spectrometry analysis showed that H2A.Z.1 and H2A.Z.2 have specific interactors, PHF14 and associated proteins for H2A.Z.1 and the histone deacetylase SIRT1 for H2A.Z.2. Moreover, we show that PHF14 and SIRT1 mediate the functional antagonism between H2A.Z.1 and H2A.Z.2. In conclusion, we propose a model in which the balance between H2A.Z.1 and H2A.Z.2 at promoters is critically important to regulate specific gene expression and depends on the recruitment of specific proteins. Our work thus provides an additional layer of complexity to the control of gene expression by histone variants.
2020-02-27 | GSE131574 | GEO
Project description:The H2A.Z histone variant plays major roles in the control of gene expression. In human, H2A.Z is encoded by two genes expressing two isoforms, H2A.Z.1 and H2A.Z.2 differing by three amino acids. Here, we undertook an integrated analysis of the independent and interdependent functions of these two isoforms in gene expression using endogenously-tagged isoform. RNA-Seq analysis following depletion of either isoform or both together in untransformed cells showed that they can regulate both distinct and overlapping sets of genes positively or negatively in a context-dependent manner. Our data revealed that the two isoforms have similar or antagonistic function depending on the gene. H2A.Z.1 and H2A.Z.2 can replace each other at Transcription Start Sites, providing a molecular explanation for this interplay. Mass spectrometry analysis showed that H2A.Z.1 and H2A.Z.2 have specific interactors, PHF14 and associated proteins for H2A.Z.1 and the histone deacetylase SIRT1 for H2A.Z.2. Moreover, we show that PHF14 and SIRT1 mediate the functional antagonism between H2A.Z.1 and H2A.Z.2. In conclusion, we propose a model in which the balance between H2A.Z.1 and H2A.Z.2 at promoters is critically important to regulate specific gene expression and depends on the recruitment of specific proteins. Our work thus provides an additional layer of complexity to the control of gene expression by histone variants.
2020-02-27 | GSE131576 | GEO
Project description:The H2A.Z histone variant plays major roles in the control of gene expression. In human, H2A.Z is encoded by two genes expressing two isoforms, H2A.Z.1 and H2A.Z.2 differing by three amino acids. Here, we undertook an integrated analysis of the independent and interdependent functions of these two isoforms in gene expression using endogenously-tagged isoform. RNA-Seq analysis following depletion of either isoform or both together in untransformed cells showed that they can regulate both distinct and overlapping sets of genes positively or negatively in a context-dependent manner. Our data revealed that the two isoforms have similar or antagonistic function depending on the gene. H2A.Z.1 and H2A.Z.2 can replace each other at Transcription Start Sites, providing a molecular explanation for this interplay. Mass spectrometry analysis showed that H2A.Z.1 and H2A.Z.2 have specific interactors, PHF14 and associated proteins for H2A.Z.1 and the histone deacetylase SIRT1 for H2A.Z.2. Moreover, we show that PHF14 and SIRT1 mediate the functional antagonism between H2A.Z.1 and H2A.Z.2. In conclusion, we propose a model in which the balance between H2A.Z.1 and H2A.Z.2 at promoters is critically important to regulate specific gene expression and depends on the recruitment of specific proteins. Our work thus provides an additional layer of complexity to the control of gene expression by histone variants.
2020-02-27 | GSE131571 | GEO