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Project description:BackgroundH3.3 is an ancient and conserved H3 variant and plays essential roles in transcriptional regulation. HIRA complex, which is composed of HIRA, UBN1 or UBN2, and Cabin1, is a H3.3 specific chaperone complex. However, it still remains largely uncharacterized how HIRA complex specifically recognizes and deposits H3.3 to the chromatin, such as promoters and enhancers.ResultsIn this study, we demonstrate that the UBN1 or UBN2 subunit is mainly responsible for specific recognition and direct binding of H3.3 by the HIRA complex. While the HIRA subunit can enhance the binding affinity of UBN1 toward H3.3, Cabin1 subunit cannot. We also demonstrate that both Ala87 and Gly90 residues of H3.3 are required and sufficient for the specific recognition and binding by UBN1. ChIP-seq studies reveal that two independent HIRA complexes (UBN1-HIRA and UBN2-HIRA) can cooperatively deposit H3.3 to cis-regulatory regions, including active promoters and active enhancers in mouse embryonic stem (mES) cells. Importantly, disruption of histone chaperone activities of UBN1 and UBN2 by FID/AAA mutation results in the defect of H3.3 deposition at promoters of developmental genes involved in neural differentiation, and subsequently causes the failure of activation of these genes during neural differentiation of mES cells.ConclusionTogether, our results provide novel insights into the mechanism by which the HIRA complex specifically recognizes and deposits H3.3 at promoters and enhancers of developmental genes, which plays a critical role in neural differentiation of mES cells.
| S-EPMC6171237 | biostudies-literature
Project description:In this study, we demonstrate that the UBN1 or UBN2 subunit is mainly responsible for specific recognition and direct binding of H3.3 by the HIRA complex, while the HIRA subunit can enhance the binding affinity of UBN1 toward H3.3, but Cabin1 subunit cannot. We also demonstrate that both Ala87 and Gly90 residues of H3.3 are required and sufficient for the specific recognition and binding by UBN1. ChIP-seq studies reveal that two independent HIRA complexes (UBN1-HIRA and UBN2-HIRA) can cooperatively deposit H3.3 to cis-regulatory regions, including active promoters and active enhancers in mouse embryonic stem (mES) cells. Importantly, disruption of histone chaperone activities of UBN1 and UBN2 by FID/AAA mutation results in the defect of H3.3 deposition at promoters of developmental genes involved in neural differentiation, and subsequently causes the failure of activation of these genes during neural differentiation of mES cells. Together, our results provide novel insights into the mechanism by which the HIRA complex specifically recognizes and deposits H3.3 at promoters and enhancers of developmental genes, which plays a critical role in neural differentiation of mES cells
2018-08-23 | GSE117034 | GEO
Project description:In this study, we demonstrate that the UBN1 or UBN2 subunit is mainly responsible for specific recognition and direct binding of H3.3 by the HIRA complex, while the HIRA subunit can enhance the binding affinity of UBN1 toward H3.3, but Cabin1 subunit cannot. We also demonstrate that both Ala87 and Gly90 residues of H3.3 are required and sufficient for the specific recognition and binding by UBN1. ChIP-seq studies reveal that two independent HIRA complexes (UBN1-HIRA and UBN2-HIRA) can cooperatively deposit H3.3 to cis-regulatory regions, including active promoters and active enhancers in mouse embryonic stem (mES) cells. Importantly, disruption of histone chaperone activities of UBN1 and UBN2 by FID/AAA mutation results in the defect of H3.3 deposition at promoters of developmental genes involved in neural differentiation, and subsequently causes the failure of activation of these genes during neural differentiation of mES cells. Together, our results provide novel insights into the mechanism by which the HIRA complex specifically recognizes and deposits H3.3 at promoters and enhancers of developmental genes, which plays a critical role in neural differentiation of mES cells
2018-08-23 | GSE117032 | GEO
Project description:UBN1/2 of HIRA complex is responsible for recognition and deposition of H3.3 at cis-regulatory elements of genes in mouse ES cells
| PRJNA480880 | ENA
Project description:Polycomb repressive complex 2 (PRC2) regulates gene expression during lineage specification through trimethylation of lysine 27 on histone H3 (H3K27me3). In Drosophila, polycomb binding sites are dynamic chromatin regions enriched with the histone variant H3.3. Here, we show that, in mouse embryonic stem cells (ESCs), H3.3 is required for proper establishment of H3K27me3 at the promoters of developmentally regulated genes. Upon H3.3 depletion, these promoters show reduced nucleosome turnover measured by deposition of de novo synthesized histones and reduced PRC2 occupancy. Further, we show H3.3-dependent interaction of PRC2 with the histone chaperone, Hira, and that Hira localization to chromatin requires H3.3. Our data demonstrate the importance of H3.3 in maintaining a chromatin landscape in ESCs that is important for proper gene regulation during differentiation. Moreover, our findings support the emerging notion that H3.3 has multiple functions in distinct genomic locations that are not always correlated with an "active" chromatin state.
| S-EPMC3838450 | biostudies-literature
Project description:UBN1/2 of HIRA complex is responsible for recognition and deposition of H3.3 at cis-regulatory elements of genes in mouse ES cells [ChIP-seq]
| PRJNA480878 | ENA
Project description:UBN1/2 of HIRA complex is responsible for recognition and deposition of H3.3 at cis-regulatory elements of genes in mouse ES cells [RNA-seq]
| PRJNA480876 | ENA
Project description:The histone chaperone HIRA is involved in depositing histone variant H3.3 into distinct genic regions, including promoters, enhancers, and gene bodies. However, how HIRA deposits H3.3 to these regions remains elusive. Through a short hairpin RNA (shRNA) screening, we identified single-stranded DNA binding protein replication protein A (RPA) as a regulator of the deposition of newly synthesized H3.3 into chromatin. We show that RPA physically interacts with HIRA to form RPA-HIRA-H3.3 complexes, and it co-localizes with HIRA and H3.3 at gene promoters and enhancers. Depletion of RPA1, the largest subunit of the RPA complex, dramatically reduces both HIRA association with chromatin and the deposition of newly synthesized H3.3 at promoters and enhancers and leads to altered transcription at gene promoters. These results support a model whereby RPA, best known for its role in DNA replication and repair, recruits HIRA to promoters and enhancers and regulates deposition of newly synthesized H3.3 to these regulatory elements for gene regulation.
| S-EPMC5460635 | biostudies-literature
Project description:The mammalian HIRA/UBN1/CABIN1/ASF1a (HUCA) histone chaperone complex deposits the histone H3 variant H3.3 into chromatin and is linked to gene activation, repression, and chromatin assembly in diverse cell contexts. We recently reported that a short N-terminal fragment of UBN1 containing amino acids 1-175 is necessary and sufficient for interaction with the WD repeats of HIRA and attributed this interaction to a region from residues 120-175 that is highly conserved with the yeast ortholog Hpc2 and so termed the HRD for Hpc2-related domain. In this report, through a more comprehensive and refined biochemical and mutational analysis, we identify a smaller and more moderately conserved region within residues 41-77 of UBN1, which we term the NHRD, that is essential for interaction with the HIRA WD repeats; we further demonstrate that the HRD is dispensable for this interaction. We employ analytical ultracentrifugation studies to demonstrate that the NHRD of UBN1 and the WD repeats of HIRA form a tight 1:1 complex with a dissociation constant in the nanomolar range. Mutagenesis experiments identify several key residues in the NHRD that are required for interaction with the HIRA WD repeat domain, stability of the HUCA complex in vitro and in vivo, and changes in chromatin organization in primary human cells. Together, these studies implicate the NHRD domain of UBN1 as being an essential region for HIRA interaction and chromatin organization by the HUCA complex.
| S-EPMC3320765 | biostudies-literature
Project description:Actively transcribed genes are enriched with the histone variant H3.3. Although H3.3 deposition has been linked to transcription, mechanisms controlling this process remain elusive. We investigated the role of the histone methyltransferase Wolf-Hirschhorn syndrome candidate 1 (WHSC1) (NSD2/MMSET) in H3.3 deposition into interferon (IFN) response genes. IFN treatment triggered robust H3.3 incorporation into activated genes, which continued even after cessation of transcription. Likewise, UV radiation caused H3.3 deposition in UV-activated genes. However, in Whsc1(-/-) cells IFN- or UV-triggered H3.3 deposition was absent, along with a marked reduction in IFN- or UV-induced transcription. We found that WHSC1 interacted with the bromodomain protein 4 (BRD4) and the positive transcription elongation factor b (P-TEFb) and facilitated transcriptional elongation. WHSC1 also associated with HIRA, the H3.3-specific histone chaperone, independent of BRD4 and P-TEFb. WHSC1 and HIRA co-occupied IFN-stimulated genes and supported prolonged H3.3 incorporation, leaving a lasting transcriptional mark. Our results reveal a previously unrecognized role of WHSC1, which links transcriptional elongation and H3.3 deposition into activated genes through two molecularly distinct pathways.
| S-EPMC3770338 | biostudies-literature