scholarly journals Dynamics of Histone H3 Deposition In Vivo Reveal a Nucleosome Gap-Filling Mechanism for H3.3 to Maintain Chromatin Integrity

2011 ◽  
Vol 44 (6) ◽  
pp. 928-941 ◽  
Author(s):  
Dominique Ray-Gallet ◽  
Adam Woolfe ◽  
Isabelle Vassias ◽  
Céline Pellentz ◽  
Nicolas Lacoste ◽  
...  
Keyword(s):  
Histone H3 ◽  
Gap Filling ◽  
Chromatin Integrity ◽  
Filling Mechanism

scholarly journals Establishment and Maintenance of Chromatin Architecture Are Promoted Independently of Transcription by the Histone Chaperone FACT and H3-K56 Acetylation in Saccharomyces cerevisiae

Genetics ◽  
2019 ◽  
Vol 211 (3) ◽  
pp. 877-892 ◽  
Author(s):  
Laura L. McCullough ◽  
Trang H. Pham ◽  
Timothy J. Parnell ◽  
Zaily Connell ◽  
Mahesh B. Chandrasekharan ◽  
...  
Keyword(s):  
Nucleosome Occupancy ◽  
Histone H3 ◽  
Antisense Transcription ◽  
Histone Chaperone ◽  
Transcript Levels ◽  
Chromatin Architecture ◽  
Transcription Profiles ◽  
Chromatin Integrity

FACT (FAcilitates Chromatin Transcription/Transactions) is a histone chaperone that can destabilize or assemble nucleosomes. Acetylation of histone H3-K56 weakens a histone–DNA contact that is central to FACT activity, suggesting that this modification could affect FACT functions. We tested this by asking how mutations of H3-K56 and FACT affect nucleosome reorganization activity in vitro, and chromatin integrity and transcript output in vivo. Mimics of unacetylated or permanently acetylated H3-K56 had different effects on FACT activity as expected, but the same mutations had surprisingly similar effects on global transcript levels. The results are consistent with emerging models that emphasize FACT’s importance in establishing global chromatin architecture prior to transcription, promoting transitions among different states as transcription profiles change, and restoring chromatin integrity after it is disturbed. Optimal FACT activity required the availability of both modified and unmodified states of H3-K56. Perturbing this balance was especially detrimental for maintaining repression of genes with high nucleosome occupancy over their promoters and for blocking antisense transcription at the +1 nucleosome. The results reveal a complex collaboration between H3-K56 modification status and multiple FACT functions, and support roles for nucleosome reorganization by FACT before, during, and after transcription.

scholarly journals FACT activity and histone H3-K56 acetylation promote optimal establishment of chromatin architecture independent of ongoing transcription inSaccharomyces cerevisiae

2018 ◽  
Author(s):  
Laura McCullough ◽  
Trang H. Pham ◽  
Timothy J. Parnell ◽  
Mahesh B. Chandrasekharan ◽  
David J. Stillman ◽  
...  
Keyword(s):  
Transcription Initiation ◽  
Nucleosome Occupancy ◽  
Histone H3 ◽  
Antisense Transcription ◽  
Transcript Levels ◽  
Chromatin Architecture ◽  
Nucleosome Structure ◽  
Chromatin Integrity

AbstractFACT is a histone chaperone that can destabilize or assemble nucleosomes. Acetylation of histone H3-K56 weakens a histone:DNA contact that is central to FACT activity, suggesting that this modification could affect FACT functions. We tested this by asking how mutations of H3-K56 and FACT affect nucleosome structure, chromatin integrity, and transcription output. Mimics of unacetylated or permanently acetylated H3-K56 had different effects on FACTin vitroandin vivoas expected, but H3-K56 and FACT mutations caused surprisingly similar changes in transcription of individual genes. Notably, neither the changes in transcript levels nor the effects on nucleosome occupancy resulting from mutations conformed to the model that FACT is needed to overcome nucleosomal barriers during transcription initiation or elongation. Instead, the results suggest that both FACT and H3-K56ac are involved in establishing chromatin architecture prior to transcription and restoring it afterwards. They contribute to a process that optimizes transcription frequency, especially at conditionally expressed genes, and restores chromatin integrity after transcription, especially at the +1 nucleosome to block antisense transcription, but FACT appears to be less involved than expected in directly promoting transcription.

scholarly journals The Trithorax group protein ASH1 requires a combination of BAH domain and AT hooks, but not the SET domain, for mitotic chromatin binding and survival

Chromosoma ◽  
2021 ◽  
Author(s):  
Philipp A. Steffen ◽  
Christina Altmutter ◽  
Eva Dworschak ◽  
Sini Junttila ◽  
Attila Gyenesei ◽  
...  
Keyword(s):  
Transcriptional Profiling ◽  
Histone H3 ◽  
Chromatin Binding ◽  
Set Domain ◽  
Trithorax Group ◽  
Trxg Protein ◽  
Group Protein ◽  
Bah Domain ◽  
Mitotic Chromatin

AbstractThe Drosophila Trithorax group (TrxG) protein ASH1 remains associated with mitotic chromatin through mechanisms that are poorly understood. ASH1 dimethylates histone H3 at lysine 36 via its SET domain. Here, we identify domains of the TrxG protein ASH1 that are required for mitotic chromatin attachment in living Drosophila. Quantitative live imaging demonstrates that ASH1 requires AT hooks and the BAH domain but not the SET domain for full chromatin binding in metaphase, and that none of these domains are essential for interphase binding. Genetic experiments show that disruptions of the AT hooks and the BAH domain together, but not deletion of the SET domain alone, are lethal. Transcriptional profiling demonstrates that intact ASH1 AT hooks and the BAH domain are required to maintain expression levels of a specific set of genes, including several involved in cell identity and survival. This study identifies in vivo roles for specific ASH1 domains in mitotic binding, gene regulation, and survival that are distinct from its functions as a histone methyltransferase.

α2-Adrenoceptor agonist dexmedetomidine protects septic acute kidney injury through increasing BMP-7 and inhibiting HDAC2 and HDAC5

2012 ◽  
Vol 303 (10) ◽  
pp. F1443-F1453 ◽  
Author(s):  
Chung-Hsi Hsing ◽  
Chiou-Feng Lin ◽  
Edmund So ◽  
Ding-Ping Sun ◽  
Tai-Chi Chen ◽  
...  
Keyword(s):  
Acute Kidney Injury ◽  
Histone Deacetylases ◽  
Trichostatin A ◽  
Histone H3 ◽  
Cecal Ligation ◽  
Kidney Injury ◽  
Protective Effects ◽  
Tnf Α

Bone morphogenetic protein (BMP)-7 protects sepsis-induced acute kidney injury (AKI). Dexmedetomidine (DEX), an α2-adrenoceptor (α2-AR) agonist, has anti-inflammatory effects. We investigated the protective effects of DEX on sepsis-induced AKI and the expression of BMP-7 and histone deacetylases (HDACs). In vitro , the effects of DEX or trichostatin A (TSA, an HDAC inhibitor) on TNF-α, monocyte chemotactic protein (MCP-1), BMP-7, and HDAC mRNA expression in LPS-stimulated rat renal tubular epithelial NRK52E cells, was determined using real-time PCR. In vivo, mice were intraperitoneally injected with DEX (25 μg/kg) or saline immediately and 12 h after cecal ligation and puncture (CLP) surgery. Twenty-four hours after CLP, we examined kidney injury and renal TNF-α, MCP-1, BMP-7, and HDAC expression. Survival was monitored for 120 h. LPS increased HDAC2, HDAC5, TNF-α, and MCP-1 expression, but decreased BMP-7 expression in NRK52E cells. DEX treatment decreased the HDAC2, HDAC5, TNF-α, and MCP-1 expression, but increased BMP-7 and acetyl histone H3 expression, whose effects were blocked by yohimbine, an α2-AR antagonist. With DEX treatment, the LPS-induced TNF-α expression and cell death were attenuated in scRNAi-NRK52E but not BMP-7 RNAi-NRK52E cells. In CLP mice, DEX treatment increased survival and attenuated AKI. The expression of HDAC2, HDAC5, TNF-α, and MCP-1 mRNA in the kidneys of CLP mice was increased, but BMP-7 was decreased. However, DEX treatment reduced those changes. DEX reduces sepsis-induced AKI by decreasing TNF-α and MCP-1 and increasing BMP-7, which is associated with decreasing HDAC2 and HDAC5, as well as increasing acetyl histone H3.

scholarly journals The chromatin-binding protein HMGN3 stimulates histone acetylation and transcription across the Glyt1 gene

2012 ◽  
Vol 442 (3) ◽  
pp. 495-505 ◽  
Author(s):  
Gráinne Barkess ◽  
Yuri Postnikov ◽  
Chrisanne D. Campos ◽  
Shivam Mishra ◽  
Gokula Mohan ◽  
...  
Keyword(s):  
Histone Modifications ◽  
Splice Variants ◽  
Binding Protein ◽  
Histone H3 ◽  
Camp Response Element Binding ◽  
Element Binding Protein ◽  
H3k14 Acetylation ◽  
H3 Acetylation

HMGNs are nucleosome-binding proteins that alter the pattern of histone modifications and modulate the binding of linker histones to chromatin. The HMGN3 family member exists as two splice forms, HMGN3a which is full-length and HMGN3b which lacks the C-terminal RD (regulatory domain). In the present study, we have used the Glyt1 (glycine transporter 1) gene as a model system to investigate where HMGN proteins are bound across the locus in vivo, and to study how the two HMGN3 splice variants affect histone modifications and gene expression. We demonstrate that HMGN1, HMGN2, HMGN3a and HMGN3b are bound across the Glyt1 gene locus and surrounding regions, and are not enriched more highly at the promoter or putative enhancer. We conclude that the peaks of H3K4me3 (trimethylated Lys4 of histone H3) and H3K9ac (acetylated Lys9 of histone H3) at the active Glyt1a promoter do not play a major role in recruiting HMGN proteins. HMGN3a/b binding leads to increased H3K14 (Lys14 of histone H3) acetylation and stimulates Glyt1a expression, but does not alter the levels of H3K4me3 or H3K9ac enrichment. Acetylation assays show that HMGN3a stimulates the ability of PCAF [p300/CREB (cAMP-response-element-binding protein)-binding protein-associated factor] to acetylate nucleosomal H3 in vitro, whereas HMGN3b does not. We propose a model where HMGN3a/b-stimulated H3K14 acetylation across the bodies of large genes such as Glyt1 can lead to more efficient transcription elongation and increased mRNA production.

scholarly journals Regulation of the Dot1 histone H3K79 methyltransferase by histone H4K16 acetylation

Science ◽  
2021 ◽  
Vol 371 (6527) ◽  
pp. eabc6663
Author(s):  
Marco Igor Valencia-Sánchez ◽  
Pablo De Ioannes ◽  
Miao Wang ◽  
David M. Truong ◽  
Rachel Lee ◽  
...  
Keyword(s):  
Histone H3 ◽  
Histone H4 ◽  
Histone H2b ◽  
Epigenetic State ◽  
Optimal Maintenance ◽  
H4k16 Acetylation ◽  
H3k79 Methylation ◽  
Regulate Gene

Dot1 (disruptor of telomeric silencing-1), the histone H3 lysine 79 (H3K79) methyltransferase, is conserved throughout evolution, and its deregulation is found in human leukemias. Here, we provide evidence that acetylation of histone H4 allosterically stimulates yeast Dot1 in a manner distinct from but coordinating with histone H2B ubiquitination (H2BUb). We further demonstrate that this stimulatory effect is specific to acetylation of lysine 16 (H4K16ac), a modification central to chromatin structure. We provide a mechanism of this histone cross-talk and show that H4K16ac and H2BUb play crucial roles in H3K79 di- and trimethylation in vitro and in vivo. These data reveal mechanisms that control H3K79 methylation and demonstrate how H4K16ac, H3K79me, and H2BUb function together to regulate gene transcription and gene silencing to ensure optimal maintenance and propagation of an epigenetic state.

Post-ovulatory ageing of mouse oocytes affects the distribution of specific spindle-associated proteins and Akt expression levels

2014 ◽  
Vol 26 (4) ◽  
pp. 562 ◽  
Author(s):  
Sandra Cecconi ◽  
Gianna Rossi ◽  
Hamid Deldar ◽  
Valerio Cellini ◽  
Felice Patacchiola ◽  
...  
Keyword(s):  
Histone H3 ◽  
Meiotic Spindle ◽  
Post Translational Modifications ◽  
Expression Levels ◽  
Acetylated Tubulin ◽  
Protein Levels ◽  
Histone 3 ◽  
Phosphorylated Akt ◽  
Associated Proteins

The aim of this study has been to determine the effects of in vivo post-ovulatory ageing (POA) on the distribution of spindle-associated proteins, histone H3/H4 post-translational modifications and on v-akt murine thymoma viral oncogene homolog 1 (Akt) expression levels. To this end, oocytes were retrieved 13, 29 and 33 h after human chorionic gonadotrophin (hCG) treatment. The presence and distribution at the meiotic spindle of acetylated tubulin, γ-tubulin, polo kinase-1 and Ser473/Thr308 phosphorylated Akt (pAkt) as well as histone H3 and H4 acetylation and phosphorylation levels were assayed via immunofluorescence. Akt expression levels were determined via reverse transcription–polymerase chain reaction and western blotting analyses. Spindles from oocytes recovered 13 h and 29 h after hCG treatment showed similar levels of acetylated tubulin but ageing induced: (1) translocation of γ-tubulin from spindle poles to microtubules, (2) absence of Thr308- and Ser473-pAkt in 76% and 30% of oocytes, respectively, and (3) a significant reduction in phosphorylation levels of serine 10 on histone 3. At 29 h, a significant decrease in Akt mRNA, but not in pAkt or Akt protein levels, was recorded. By contrast, protein content significantly decreased 33 h after hCG. We conclude that POA impairs oocyte viability and fertilisability by altering the expression levels and spindle distribution of proteins that are implicated in cell survival and chromosome segregation. Together, these events could play a role in oocyte apoptosis.

Polo-Like Kinase-1 (Plk-1) Inhibitor BI 2536 Induces Mitotic Arrest and Apoptosis in Vivo: First Demonstration of Target Inhibition in the Bone Marrow of AML Patients

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 2641-2641
Author(s):  
Kang-Hun Lee ◽  
Richard F. Schlenk ◽  
Gesine Bug ◽  
Carsten Müller-Tidow ◽  
Ralph M. Waesch ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Clinical Response ◽  
Peripheral Blood ◽  
Histone H3 ◽  
Mitotic Arrest ◽  
Clinical Testing ◽  
Facs Analysis ◽  
M Phase ◽  
Bi 2536

Abstract Background: BI 2536 is a potent and selective inhibitor of Plk-1, which plays a crucial role in the regulation of mitosis. Inhibition of Plk-1 leads to mitotic arrest and apoptosis. Therefore, Plk-1 inhibitors are currently in clinical testing as anti-proliferative agents in cancer patients. BI 2536, the first specific Plk-1 inhibitor in clinical testing, demonstrated strong anti-proliferative effects on AML cell lines in vitro and in in vivo models. To investigate target inhibition in the malignant cell, bone marrow samples from patients who participated in a Phase I/II study of BI 2536 single-agent therapy in elderly patients with relapsed or refractory AML were analyzed. Methods: Pharmacodynamic analyses including immunocytochemistry (ICC) and flow cytometry (FACS) of bone marrow (BM) were performed to examine cell morphology, phosphorylation of histone H3 (phospho-H3), and induction of apoptosis. Samples were acquired before and 24 h after the first administration of BI 2536. FACS analysis included propidium iodide (PI)-FACS to determine the percentage of cells residing in various cell cycle stages (G0/1-, S- and G2/M-phases) and Annexin V-Cy5-staining for apoptosis. Immunocytochemistry included staining for phosphorylated histone H3 and TUNEL assay for apoptosis. Results: At the time of analysis, data from 28 patients treated at doses in the range of 50 to 400 mg BI 2536 were available. BM taken after BI 2536 administration showed an increase of phospho-H3 positive cells as compared to baseline prior to treatment. There was a trend toward a positive correlation between phospho-H3 increase and increase in dosage of BI 2536 (p=0.16) when treatment at low doses (50 to 60 mg) of BI 2536 were compared to treatment at higher doses (100 to 400 mg). Furthermore, trends were observed that an increase of phospho-H3 is positively correlated with both a higher percentage of cells in G2/M and an increase in apoptotic cells. The typical morphology of cells in mitotic arrest could be demonstrated by ICC. Interestingly, when patients with progressive disease after one cycle (PD) were compared to patients with disease stabilization or response (nonPD), a statistically significant positive correlation between PD and increase in phospho-H3 compared to baseline was found (p=0.03). Also, there was a clear trend for an increase in the percentage of cells in G2/M phase and apoptosis in patients with PD compared to patients with nonPD. Preliminary evaluation of other disease characteristics including karyotype (normal vs complex), secondary AML, complete response in previous treatments, baseline value for blasts in the bone marrow or in the peripheral blood, did not reveal any signs of correlation with the clinical response to BI 2536 treatment. Conclusion: BI 2536 treatment increases the number of cells in G2/M phase (as detected by FACS analysis and phospho-H3 staining) and the number of apoptotic cells within 24 h after administration. In line with the clinical observation of rapid blast reduction both in the BM and the peripheral blood, these findings indicate that BI 2536 induces mitotic arrest and apoptosis of the malignant target cells in AML patients. The biological meaning of the correlation between the BM findings and the clinical response to BI 2536 is unclear, but if substantiated by more data, these results may suggest a predictive value of BM examinations for the response to BI 2536.

scholarly journals A role for the MLL fusion partner ENL in transcriptional elongation and chromatin modification

Blood ◽  
2007 ◽  
Vol 110 (13) ◽  
pp. 4445-4454 ◽  
Author(s):  
Dorothee Mueller ◽  
Christian Bach ◽  
Deniz Zeisig ◽  
Maria-Paz Garcia-Cuellar ◽  
Sara Monroe ◽  
...  
Keyword(s):  
Rna Polymerase Ii ◽  
Chromatin Modification ◽  
Histone H3 ◽  
Elongation Factor ◽  
Polycomb Group ◽  
Fusion Partner ◽  
Transcriptional Elongation ◽  
Associated Proteins ◽  
Group Members

Chimeric proteins joining the histone methyltransferase MLL with various fusion partners trigger distinctive lymphoid and myeloid leukemias. Here, we immunopurified proteins associated with ENL, a protein commonly fused to MLL. Identification of these ENL-associated proteins (EAPs) by mass spectrometry revealed enzymes with a known role in transcriptional elongation (RNA polymerase II C-terminal domain kinase [RNAPolII CTD] positive transcription elongation factor b [pTEFb]), and in chromatin modification (histone-H3 methyltransferase DOT1L) as well as other frequent MLL partners (AF4, AF5q31, and LAF4), and polycomb group members (RING1, CBX8, and BCoR). The composition of EAP was further verified by coimmunoprecipitation, 2-hybrid analysis, pull-down, and colocalization experiments. Purified EAP showed a histone H3 lysine 79–specific methylase activity, displayed a robust RNAPolII CTD kinase function, and counteracted the effect of the pTEFb inhibitor 5,6-dichloro-benzimidazole-riboside. In vivo, an ENL knock-down diminished genome-wide as well as gene-specific H3K79 dimethylation, reduced global run-on elongation, and inhibited transient transcriptional reporter activity. According to structure-function data, DOT1L recruitment was important for transformation by the MLL-ENL fusion derivative. These results suggest a function of ENL in histone modification and transcriptional elongation.

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