The HMG-box module in FACT is critical for suppressing epigenetic variegation of heterochromatin in fission yeast
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PROVIDER: PRJNA1057767 | ENA |
REPOSITORIES: ENA
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Project description:Chromatin condensation state is the key for retrieving genetic information. High-mobility group protein (HMG) proteins exhibit DNA-binding and bending activities, playing an important role in the regulation of chromatin structure. We have shown that nucleosomes tightly packaged into heterochromatin undergo considerable dynamic histone H2A-H2B maintenance via the direct interaction between HP1/Swi6 and facilitate chromatin transcription (FACT), which is composed of the Spt16/Pob3 heterodimer and Nhp6. In this study, we analyzed the role of Nhp6, an HMG box protein, in the FACT at heterochromatin. Pob3 mutant strains showed derepressed heterochromatin-dependent gene silencing, whereas Nhp6 mutant strains did not show significant defects in chromatin regulation or gene expression, suggesting that these two modules play different roles in chromatin regulation. We expressed a protein fusing Nhp6 to the C-terminus of Pob3, which mimics the multicellular FACT component Ssrp1. The chromatin-binding activity of FACT increased with the number of Nhp6 fused to Pob3, and the heterochromatin formation rate was promoted more strongly. Furthermore, we demonstrated that this promotion of heterochromatinization inhibited the heterochromatic variegation caused by epe1+ disruption. Heterochromatic variegation can be observed in a variety of regulatory steps; however, when it is caused by fluctuations in chromatin arrangement, it can be eliminated through the strong recruitment of the FACT complex.
2024-06-12 | GSE252092 | GEO
Project description:To assess the mechanisms by which FACT depletion leads to increased sensitivity of cells to be reprogrammed, we measured the chromatin accessibility landscape using ATAC-seq following mock treatment, SSRP1 knockdown, or SUPT16H knockdown in human fibroblasts and mock, hmg-3 or hmg-4 knockdown in whole worms, and differential gene expression in hmg-3 knockout mutants or following mock, hmg-4, or spt-16 knockdown by RNAseq.
2018-08-29 | GSE98758 | GEO
Project description:Position-effect variegation (PEV) is the stochastic transcriptional silencing of a gene positioned adjacent to heterochromatin. white-mottled X-chromosomal inversions in Drosophila are classic PEV models that show variegation of the eye color gene white due to its relocation next to pericentric heterochromatin. To obtain insight into the mechanism of PEV, we constructed detailed binding maps of Heterochromatin Protein 1 (HP1), a major component of heterochromatin, on white-mottled chromosomes. We find that HP1 invades euchromatin across the inversion breakpoints over ~175kb and ~30kb, causing de novo association of HP1 with 20 genes. However, HP1 binding levels in these regions show substantial local variation; white is most strongly bound by HP1 and is one of only two genes that are substantially repressed by heterochromatin. HP1 binding to the invaded region is exceptionally sensitive to the dosage of the histone methyltransferase Su(var)3-9, indicating that the de novo formed heterochromatin is relatively unstable. Our molecular maps demonstrate that heterochromatin can invade a normally euchromatic region, yet the strength of HP1 binding and effects on gene expression are highly dependent on local context. Keywords: DamID, gene expression, genetic modification.
2009-02-18 | GSE12395 | GEO
Project description:FACT mediates cohesin function on chromatin
| PRJNA485963 | ENA
Project description:Cohesin is a key regulator of genome architecture with roles in sister chromatid cohesion and the organisation of higher-order structures during interphase and mitosis. The recruitment and mobility of cohesin complexes on DNA are restricted by nucleosomes. Here we show that cohesin role in chromosome organization requires the histone chaperone FACT. Depletion of FACT in metaphase cells affects cohesin stability on chromatin reducing its accumulation at pericentric regions and binding on chromosome arms. Using Hi-C, we show that cohesin-dependent TAD (Topological Associated Domains)-like structures in G1 and metaphase chromosomes are disrupted in the absence of FACT. Surprisingly, sister chromatid cohesion is intact in FACT-depleted cells, although chromosome segregation failure is observed. Our results uncover a role for FACT in genome organisation by facilitating cohesin dependent compartmentalization of chromosomes into loop domains.
2019-10-11 | GSE118534 | GEO
Project description:HMGB1 and HMGB2 are members of the High Mobility Group (HMG) protein superfamily that contain a DNA binding domain (HMG-Box), and their overexpression has been related to main cancer hallmarks, tumor progression, metastasis formation and bad prognosis. We have used yeast two hybrid and affinity purification-mass spectrometry to explore HMGB1 and HMGB2 protein interactions in prostate and ovarian epithelial cells.
2018-05-07 | PXD007867 | Pride
Project description:Protein-protein interaction analysis for RNA Polymerase II and its elongation factors including: the PAF Complex, the FACT Complex, casein kinase II, DSIF, and TFIIF. These studies also characterized a number of casein kinase II phosphorylation sites on the subunits of the PAF Complex.
2015-07-20 | MSV000079197 | MassIVE
Project description:Histone chaperone FACT is commonly expressed and essential for the viability of transformed but not normal cells and its expression levels correlate with poor prognosis in cancer patients. FACT binds several components of nucleosomes and has been viewed as a factor destabilizing nucleosomes to facilitate RNA polymerase passage. To connect FACT’s role in transcription with the viability of tumor cells, we analyzed genome-wide FACT binding to chromatin in conjunction with transcription in mouse and human cells with different degrees of FACT dependence. While genomic distribution and density of FACT correlated with the intensity of transcription, FACT knockout or knockdown was unexpectedly accompanied by the elevation, rather than suppression, of transcription and with the destabilization of chromatin. These data suggest that the real function of FACT is to stabilize and reassemble nucleosomes disturbed by transcription. This function is apparently vital for tumor cells because malignant transformation is accompanied by chromatin destabilization.
2019-06-12 | GSE132576 | GEO
Project description:FACT has been identified as a histone chaperone that enables transcription through chromatin in vitro, but its role in regulating chromatin structure and transcription in vivo remains unclear. In this study, we have investigated the function and molecular mechanism of FACT in unprecedented detail, by using a rapid depletion system in combination with high-resolution genomic analyses. We show that acute depletion of FACT leads to changes in 3D chromatin structure and a concomitant multilayered transcriptional defect, including loss of promoter-proximal pausing, deregulated elongation and increased drop-off of RNA Pol II. Integration of in vitro transcription assays shows that FACT stimulates RNA Pol II pausing by stabilizing nucleosomal architecture, while simultaneously promoting transcription through the +1 nucleosome. In addition to providing detailed mechanistic insight into the seemingly contradictory functions of FACT and the regulation of promoter-proximal pausing, our study indicates a direct coupling of chromatin structure and transcription.
2024-05-30 | GSE241421 | GEO
Project description:This SuperSeries is composed of the SubSeries listed below. FACT (Facilitates Chromatin Transcription) is an evolutionarily conserved histone chaperone that was initially identified as an activity capable of promoting RNA polymerase II transcription through nucleosomes in vitro. In this report, we describe a global analysis of FACT function in Pol II transcription in Drosophila. We present evidence that loss of FACT has a dramatic impact on Pol II elongation-coupled processes including H3K4 and H3K36 methylation, consistent with a role for FACT in coordinating histone modification and chromatin architecture during Pol II transcription. Importantly, we identify a role for FACT in the maintenance of promoter proximal Pol II pausing, a key step in transcription activation in higher eukaryotes. These findings bring to light a broader role for FACT in the regulation of Pol II transcription.
2019-06-14 | GSE129236 | GEO