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Rabbit Anti-Histone H3, monomethyl (Lys4) ChIP Grade Polyclonal Antibody, Unconjugated

RRID:AB_306847

Antibody ID

AB_306847

Target Antigen

Histone H3 (mono methyl K4) - ChIP Grade bovine, canine, chicken/avian, donkey, drosophila, feline, guinea pig, hamster, horse, human, mouse, other, porcine, rabbit, rat, sheep, simian, xenopus, yeast, reacts with human, cow, indian muntjac, pig (pmid 18502896), plants, tetrahymena sp and xenopus laevisthis antibody reacts with mono methylated k4 within a sequence found in all mammals and a wide range of other species, including s cerevisiae, s pombe, n crassa, aspergillus nidulans, d melanogaster, c reinhardtii, c elegans, arabidopsis, chicken, plants (r acetosa), zebrafish, tobacco and c albicans (see abreview) tetrahymena sp (pubmed 17284592) and xenopus laevis (from pubmed:17440165)the antibody will react with any species where the modification is present

Proper Citation

(Abcam Cat# ab8895, RRID:AB_306847)

Clonality

polyclonal antibody

Comments

validation status unknown, seller recommendations provided in 2012: Blocking/Neutralize; Flow Cytometry; Immunofluorescence; Immunohistochemistry; Immunoprecipitation; Other; Western Blot; Chromatin IP, Chromatin IP/Chromatin IP, Flow Cytometry, Immunocytochemistry/Immunofluorescence, Immunofluorescence, Immunohistochemistry-P, Western Blot

Host Organism

rabbit

Vendor

Abcam

Histone Methylation by SETD1A Protects Nascent DNA through the Nucleosome Chaperone Activity of FANCD2.

  • Higgs MR
  • Mol. Cell
  • 2018 Jul 5

Literature context:


Abstract:

Components of the Fanconi anemia and homologous recombination pathways play a vital role in protecting newly replicated DNA from uncontrolled nucleolytic degradation, safeguarding genome stability. Here we report that histone methylation by the lysine methyltransferase SETD1A is crucial for protecting stalled replication forks from deleterious resection. Depletion of SETD1A sensitizes cells to replication stress and leads to uncontrolled DNA2-dependent resection of damaged replication forks. The ability of SETD1A to prevent degradation of these structures is mediated by its ability to catalyze methylation on Lys4 of histone H3 (H3K4) at replication forks, which enhances FANCD2-dependent histone chaperone activity. Suppressing H3K4 methylation or expression of a chaperone-defective FANCD2 mutant leads to loss of RAD51 nucleofilament stability and severe nucleolytic degradation of replication forks. Our work identifies epigenetic modification and histone mobility as critical regulatory mechanisms in maintaining genome stability by restraining nucleases from irreparably damaging stalled replication forks.

Funding information:
  • NCRR NIH HHS - RR024574(United States)

LSD1 Ablation Stimulates Anti-tumor Immunity and Enables Checkpoint Blockade.

  • Sheng W
  • Cell
  • 2018 Jun 18

Literature context:


Abstract:

Chromatin regulators play a broad role in regulating gene expression and, when gone awry, can lead to cancer. Here, we demonstrate that ablation of the histone demethylase LSD1 in cancer cells increases repetitive element expression, including endogenous retroviral elements (ERVs), and decreases expression of RNA-induced silencing complex (RISC) components. Significantly, this leads to double-stranded RNA (dsRNA) stress and activation of type 1 interferon, which stimulates anti-tumor T cell immunity and restrains tumor growth. Furthermore, LSD1 depletion enhances tumor immunogenicity and T cell infiltration in poorly immunogenic tumors and elicits significant responses of checkpoint blockade-refractory mouse melanoma to anti-PD-1 therapy. Consistently, TCGA data analysis shows an inverse correlation between LSD1 expression and CD8+ T cell infiltration in various human cancers. Our study identifies LSD1 as a potent inhibitor of anti-tumor immunity and responsiveness to immunotherapy and suggests LSD1 inhibition combined with PD-(L)1 blockade as a novel cancer treatment strategy.

Funding information:
  • Intramural NIH HHS - U01-CA84967(United States)
  • NCI NIH HHS - R01 CA118487()
  • NCI NIH HHS - R35 CA210104()
  • NCI NIH HHS - T32 CA207021()

The Epigenetic State of PRDM16-Regulated Enhancers in Radial Glia Controls Cortical Neuron Position.

  • Baizabal JM
  • Neuron
  • 2018 Jun 6

Literature context:


Abstract:

The epigenetic landscape is dynamically remodeled during neurogenesis. However, it is not understood how chromatin modifications in neural stem cells instruct the formation of complex structures in the brain. We report that the histone methyltransferase PRDM16 is required in radial glia to regulate lineage-autonomous and stage-specific gene expression programs that control number and position of upper layer cortical projection neurons. PRDM16 regulates the epigenetic state of transcriptional enhancers to activate genes involved in intermediate progenitor cell production and repress genes involved in cell migration. The histone methyltransferase domain of PRDM16 is necessary in radial glia to promote cortical neuron migration through transcriptional silencing. We show that repression of the gene encoding the E3 ubiquitin ligase PDZRN3 by PRDM16 determines the position of upper layer neurons. These findings provide insights into how epigenetic control of transcriptional enhancers in radial glial determines the organization of the mammalian cerebral cortex.

Funding information:
  • NCI NIH HHS - R01 CA109038-04(United States)

MLL4 Is Required to Maintain Broad H3K4me3 Peaks and Super-Enhancers at Tumor Suppressor Genes.

  • Dhar SS
  • Mol. Cell
  • 2018 Jun 7

Literature context:


Abstract:

Super-enhancers are large clusters of enhancers that activate gene expression. Broad trimethyl histone H3 lysine 4 (H3K4me3) often defines active tumor suppressor genes. However, how these epigenomic signatures are regulated for tumor suppression is little understood. Here we show that brain-specific knockout of the H3K4 methyltransferase MLL4 (a COMPASS-like enzyme, also known as KMT2D) in mice spontaneously induces medulloblastoma. Mll4 loss upregulates oncogenic Ras and Notch pathways while downregulating neuronal gene expression programs. MLL4 enhances DNMT3A-catalyzed DNA methylation and SIRT1/BCL6-mediated H4K16 deacetylation, which antagonize expression of Ras activators and Notch pathway components, respectively. Notably, Mll4 loss downregulates tumor suppressor genes (e.g., Dnmt3a and Bcl6) by diminishing broad H3K4me3 and super-enhancers and also causes widespread impairment of these epigenomic signatures during medulloblastoma genesis. These findings suggest an anti-tumor role for super-enhancers and provide a unique tumor-suppressive mechanism in which MLL4 is necessary to maintain broad H3K4me3 and super-enhancers at tumor suppressor genes.

Funding information:
  • NCI NIH HHS - P30 CA016672()
  • NCI NIH HHS - R01 CA157919()
  • NCI NIH HHS - R01 CA207098()
  • NCI NIH HHS - R01 CA207109()
  • NIAID NIH HHS - 1R01AI059372(United States)

H3.3K27M mutant proteins reprogram epigenome by sequestering the PRC2 complex to poised enhancers.

  • Fang D
  • Elife
  • 2018 Jun 22

Literature context:


Abstract:

Expression of histone H3.3K27M mutant proteins in human diffuse intrinsic pontine glioma (DIPG) results in a global reduction of tri-methylation of H3K27 (H3K27me3), and paradoxically, H3K27me3 peaks remain at hundreds of genomic loci, a dichotomous change that lacks mechanistic insights. Here, we show that the PRC2 complex is sequestered at poised enhancers, but not at active promoters with high levels of H3.3K27M proteins, thereby contributing to the global reduction of H3K27me3. Moreover, the levels of H3.3K27M proteins are low at the retained H3K27me3 peaks and consequently having minimal effects on the PRC2 activity at these loci. H3K27me3-mediated silencing at specific tumor suppressor genes, including Wilms Tumor 1, promotes proliferation of DIPG cells. These results support a model in which the PRC2 complex is redistributed to poised enhancers in H3.3K27M mutant cells and contributes to tumorigenesis in part by locally enhancing H3K27me3, and hence silencing of tumor suppressor genes.

Funding information:
  • National Institutes of Health - CA204297()
  • NIGMS NIH HHS - GM107466(United States)

PRC1 Fine-tunes Gene Repression and Activation to Safeguard Skin Development and Stem Cell Specification.

  • Cohen I
  • Cell Stem Cell
  • 2018 May 3

Literature context:


Abstract:

Polycomb repressive complexes (PRCs) 1 and 2 are essential chromatin regulators of cell identity. PRC1, a dominant executer of Polycomb-mediated control, functions as multiple sub-complexes that possess catalytic-dependent H2AK119 mono-ubiquitination (H2AK119ub) and catalytic-independent activities. Here, we show that, despite its well-established repressor functions, PRC1 binds to both silent and active genes. Through in vivo loss-of-function studies, we show that global PRC1 function is essential for skin development and stem cell (SC) specification, whereas PRC1 catalytic activity is dispensable. Further dissection demonstrated that both canonical and non-canonical PRC1 complexes bind to repressed genes, marked by H2AK119ub and PRC2-mediated H3K27me3. Interestingly, loss of canonical PRC1, PRC1 catalytic activity, or PRC2 leads to expansion of mechanosensitive Merkel cells in neonatal skin. Non-canonical PRC1 complexes, however, also bind to and promote expression of genes critical for skin development and SC formation. Together, our findings highlight PRC1's diverse roles in executing a precise developmental program.

Funding information:
  • NIAMS NIH HHS - R00 AR057817()
  • NIAMS NIH HHS - R01 AR063724()
  • NINDS NIH HHS - R21 NS055261(United States)

BRG1 governs glucocorticoid receptor interactions with chromatin and pioneer factors across the genome.

  • Hoffman JA
  • Elife
  • 2018 May 24

Literature context:


Abstract:

The Glucocorticoid Receptor (GR) alters transcriptional activity in response to hormones by interacting with chromatin at GR binding sites (GBSs) throughout the genome. Our work in human breast cancer cells identifies three classes of GBSs with distinct epigenetic characteristics and reveals that BRG1 interacts with GBSs prior to hormone exposure. The GBSs pre-occupied by BRG1 are more accessible and transcriptionally active than other GBSs. BRG1 is required for a proper and robust transcriptional hormone response and knockdown of BRG1 blocks recruitment of the pioneer factors FOXA1 and GATA3 to GBSs. Finally, GR interaction with FOXA1 and GATA3 binding sites was restricted to sites pre-bound by BRG1. These findings demonstrate that BRG1 establishes specialized chromatin environments that define multiple classes of GBS. This in turn predicts that GR and other transcriptional activators function via multiple distinct chromatin-based mechanisms to modulate the transcriptional response.

Funding information:
  • National Institute of Environmental Health Sciences - Z01 ES071006-17()
  • NHLBI NIH HHS - HL-69256(United States)

BRD4 Inhibition Is Synthetic Lethal with PARP Inhibitors through the Induction of Homologous Recombination Deficiency.

  • Sun C
  • Cancer Cell
  • 2018 Mar 12

Literature context:


Abstract:

Poly(ADP-ribose) polymerase inhibitors (PARPi) are selectively active in cells with homologous recombination (HR) deficiency (HRD) caused by mutations in BRCA1, BRCA2, and other pathway members. We sought small molecules that induce HRD in HR-competent cells to induce synthetic lethality with PARPi and extend the utility of PARPi. We demonstrated that inhibition of bromodomain containing 4 (BRD4) induced HRD and sensitized cells across multiple tumor lineages to PARPi regardless of BRCA1/2, TP53, RAS, or BRAF mutation status through depletion of the DNA double-stand break resection protein CtIP (C-terminal binding protein interacting protein). Importantly, BRD4 inhibitor (BRD4i) treatment reversed multiple mechanisms of resistance to PARPi. Furthermore, PARPi and BRD4i are synergistic in multiple in vivo models.

Funding information:
  • NCI NIH HHS - CA986366(United States)
  • NCI NIH HHS - P50 CA083639()

Activation of the Notch Signaling Pathway In Vivo Elicits Changes in CSL Nuclear Dynamics.

  • Gomez-Lamarca MJ
  • Dev. Cell
  • 2018 Mar 12

Literature context:


Abstract:

A key feature of Notch signaling is that it directs immediate changes in transcription via the DNA-binding factor CSL, switching it from repression to activation. How Notch generates both a sensitive and accurate response-in the absence of any amplification step-remains to be elucidated. To address this question, we developed real-time analysis of CSL dynamics including single-molecule tracking in vivo. In Notch-OFF nuclei, a small proportion of CSL molecules transiently binds DNA, while in Notch-ON conditions CSL recruitment increases dramatically at target loci, where complexes have longer dwell times conferred by the Notch co-activator Mastermind. Surprisingly, recruitment of CSL-related corepressors also increases in Notch-ON conditions, revealing that Notch induces cooperative or "assisted" loading by promoting local increase in chromatin accessibility. Thus, in vivo Notch activity triggers changes in CSL dwell times and chromatin accessibility, which we propose confer sensitivity to small input changes and facilitate timely shut-down.

Funding information:
  • Wellcome Trust - CA084179(United Kingdom)
  • Wellcome Trust - R01 CA178974()

A Non-catalytic Function of SETD1A Regulates Cyclin K and the DNA Damage Response.

  • Hoshii T
  • Cell
  • 2018 Feb 22

Literature context:


Abstract:

MLL/SET methyltransferases catalyze methylation of histone 3 lysine 4 and play critical roles in development and cancer. We assessed MLL/SET proteins and found that SETD1A is required for survival of acute myeloid leukemia (AML) cells. Mutagenesis studies and CRISPR-Cas9 domain screening show the enzymatic SET domain is not necessary for AML cell survival but that a newly identified region termed the "FLOS" (functional location on SETD1A) domain is indispensable. FLOS disruption suppresses DNA damage response genes and induces p53-dependent apoptosis. The FLOS domain acts as a cyclin-K-binding site that is required for chromosomal recruitment of cyclin K and for DNA-repair-associated gene expression in S phase. These data identify a connection between the chromatin regulator SETD1A and the DNA damage response that is independent of histone methylation and suggests that targeting SETD1A and cyclin K complexes may represent a therapeutic opportunity for AML and, potentially, for other cancers.

Funding information:
  • NICHD NIH HHS - R01 HD070056-01(United States)

Programming of Schwann Cells by Lats1/2-TAZ/YAP Signaling Drives Malignant Peripheral Nerve Sheath Tumorigenesis.

  • Wu LMN
  • Cancer Cell
  • 2018 Feb 12

Literature context:


Abstract:

Malignant peripheral nerve sheath tumors (MPNSTs) are highly aggressive Schwann cell (SC)-lineage-derived sarcomas. Molecular events driving SC-to-MPNST transformation are incompletely understood. Here, we show that human MPNSTs exhibit elevated HIPPO-TAZ/YAP expression, and that TAZ/YAP hyperactivity in SCs caused by Lats1/2 loss potently induces high-grade nerve-associated tumors with full penetrance. Lats1/2 deficiency reprograms SCs to a cancerous, progenitor-like phenotype and promotes hyperproliferation. Conversely, disruption of TAZ/YAP activity alleviates tumor burden in Lats1/2-deficient mice and inhibits human MPNST cell proliferation. Moreover, genome-wide profiling reveals that TAZ/YAP-TEAD1 directly activates oncogenic programs, including platelet-derived growth factor receptor (PDGFR) signaling. Co-targeting TAZ/YAP and PDGFR pathways inhibits tumor growth. Thus, our findings establish a previously unrecognized convergence between Lats1/2-TAZ/YAP signaling and MPNST pathogenesis, revealing potential therapeutic targets in these untreatable tumors.

Funding information:
  • NHLBI NIH HHS - R01 HL132211()
  • NIA NIH HHS - R01 AG040990(United States)
  • NINDS NIH HHS - R01 NS072427()
  • NINDS NIH HHS - R01 NS075243()
  • NINDS NIH HHS - R01 NS078092()
  • NINDS NIH HHS - R01 NS086219()
  • NINDS NIH HHS - R37 NS096359()

Dual Targeting of Oncogenic Activation and Inflammatory Signaling Increases Therapeutic Efficacy in Myeloproliferative Neoplasms.

  • Kleppe M
  • Cancer Cell
  • 2018 Jan 8

Literature context:


Abstract:

Genetic and functional studies underscore the central role of JAK/STAT signaling in myeloproliferative neoplasms (MPNs). However, the mechanisms that mediate transformation in MPNs are not fully delineated, and clinically utilized JAK inhibitors have limited ability to reduce disease burden or reverse myelofibrosis. Here we show that MPN progenitor cells are characterized by marked alterations in gene regulation through differential enhancer utilization, and identify nuclear factor κB (NF-κB) signaling as a key pathway activated in malignant and non-malignant cells in MPN. Inhibition of BET bromodomain proteins attenuated NF-κB signaling and reduced cytokine production in vivo. Most importantly, combined JAK/BET inhibition resulted in a marked reduction in the serum levels of inflammatory cytokines, reduced disease burden, and reversed bone marrow fibrosis in vivo.

Funding information:
  • NCI NIH HHS - K08 CA181507()
  • NCI NIH HHS - P30 CA008748()
  • NCI NIH HHS - R01 CA151949()
  • NCI NIH HHS - R01 CA173636()
  • NCI NIH HHS - R21 CA167800(United States)
  • NCI NIH HHS - R35 CA197594()
  • NHLBI NIH HHS - K99 HL122503()

IL-10 Signaling Remodels Adipose Chromatin Architecture to Limit Thermogenesis and Energy Expenditure.

  • Rajbhandari P
  • Cell
  • 2018 Jan 11

Literature context:


Abstract:

Signaling pathways that promote adipose tissue thermogenesis are well characterized, but the limiters of energy expenditure are largely unknown. Here, we show that ablation of the anti-inflammatory cytokine IL-10 improves insulin sensitivity, protects against diet-induced obesity, and elicits the browning of white adipose tissue. Mechanistic studies define bone marrow cells as the source of the IL-10 signal and adipocytes as the target cell type mediating these effects. IL-10 receptor alpha is highly enriched in mature adipocytes and is induced in response to differentiation, obesity, and aging. Assay for transposase-accessible chromatin sequencing (ATAC-seq), ChIP-seq, and RNA-seq reveal that IL-10 represses the transcription of thermogenic genes in adipocytes by altering chromatin accessibility and inhibiting ATF and C/EBPβ recruitment to key enhancer regions. These findings expand our understanding of the relationship between inflammatory signaling pathways and adipose tissue function and provide insight into the physiological control of thermogenesis that could inform future therapy.

Funding information:
  • Howard Hughes Medical Institute - (United States)
  • NHLBI NIH HHS - K08 HL128822()
  • NHLBI NIH HHS - P01 HL090553()
  • NIAID NIH HHS - T32 AI007323()
  • NIDDK NIH HHS - F32 DK104484()
  • NIDDK NIH HHS - F32 DK109601()
  • NIDDK NIH HHS - P30 DK063491()
  • NIGMS NIH HHS - R01 GM086372()
  • NIGMS NIH HHS - T32 GM008042()

Aberrant Activation of a Gastrointestinal Transcriptional Circuit in Prostate Cancer Mediates Castration Resistance.

  • Shukla S
  • Cancer Cell
  • 2017 Dec 11

Literature context:


Abstract:

Prostate cancer exhibits a lineage-specific dependence on androgen signaling. Castration resistance involves reactivation of androgen signaling or activation of alternative lineage programs to bypass androgen requirement. We describe an aberrant gastrointestinal-lineage transcriptome expressed in ∼5% of primary prostate cancer that is characterized by abbreviated response to androgen-deprivation therapy and in ∼30% of castration-resistant prostate cancer. This program is governed by a transcriptional circuit consisting of HNF4G and HNF1A. Cistrome and chromatin analyses revealed that HNF4G is a pioneer factor that generates and maintains enhancer landscape at gastrointestinal-lineage genes, independent of androgen-receptor signaling. In HNF4G/HNF1A-double-negative prostate cancer, exogenous expression of HNF4G at physiologic levels recapitulates the gastrointestinal transcriptome, chromatin landscape, and leads to relative castration resistance.

Funding information:
  • NICHD NIH HHS - R01 HD52472(United States)

Cohesin Loss Eliminates All Loop Domains.

  • Rao SSP
  • Cell
  • 2017 Oct 5

Literature context:


Abstract:

The human genome folds to create thousands of intervals, called "contact domains," that exhibit enhanced contact frequency within themselves. "Loop domains" form because of tethering between two loci-almost always bound by CTCF and cohesin-lying on the same chromosome. "Compartment domains" form when genomic intervals with similar histone marks co-segregate. Here, we explore the effects of degrading cohesin. All loop domains are eliminated, but neither compartment domains nor histone marks are affected. Loss of loop domains does not lead to widespread ectopic gene activation but does affect a significant minority of active genes. In particular, cohesin loss causes superenhancers to co-localize, forming hundreds of links within and across chromosomes and affecting the regulation of nearby genes. We then restore cohesin and monitor the re-formation of each loop. Although re-formation rates vary greatly, many megabase-sized loops recovered in under an hour, consistent with a model where loop extrusion is rapid.

Funding information:
  • NIGMS NIH HHS - T32 GM008294()

CTCF-Mediated Enhancer-Promoter Interaction Is a Critical Regulator of Cell-to-Cell Variation of Gene Expression.

  • Ren G
  • Mol. Cell
  • 2017 Sep 21

Literature context:


Abstract:

Recent studies indicate that even a homogeneous population of cells display heterogeneity in gene expression and response to environmental stimuli. Although promoter structure critically influences the cell-to-cell variation of gene expression in bacteria and lower eukaryotes, it remains unclear what controls the gene expression noise in mammals. Here we report that CTCF decreases cell-to-cell variation of expression by stabilizing enhancer-promoter interaction. We show that CTCF binding sites are interwoven with enhancers within topologically associated domains (TADs) and a positive correlation is found between CTCF binding and the activity of the associated enhancers. Deletion of CTCF sites compromises enhancer-promoter interactions. Using single-cell flow cytometry and single-molecule RNA-FISH assays, we demonstrate that knocking down of CTCF or deletion of a CTCF binding site results in increased cell-to-cell variation of gene expression, indicating that long-range promoter-enhancer interaction mediated by CTCF plays important roles in controlling the cell-to-cell variation of gene expression in mammalian cells.

Dynamic Control of X Chromosome Conformation and Repression by a Histone H4K20 Demethylase.

  • Brejc K
  • Cell
  • 2017 Sep 21

Literature context:


Abstract:

Chromatin modification and higher-order chromosome structure play key roles in gene regulation, but their functional interplay in controlling gene expression is elusive. We have discovered the machinery and mechanism underlying the dynamic enrichment of histone modification H4K20me1 on hermaphrodite X chromosomes during C. elegans dosage compensation and demonstrated H4K20me1's pivotal role in regulating higher-order chromosome structure and X-chromosome-wide gene expression. The structure and the activity of the dosage compensation complex (DCC) subunit DPY-21 define a Jumonji demethylase subfamily that converts H4K20me2 to H4K20me1 in worms and mammals. Selective inactivation of demethylase activity eliminates H4K20me1 enrichment in somatic cells, elevates X-linked gene expression, reduces X chromosome compaction, and disrupts X chromosome conformation by diminishing the formation of topologically associating domains (TADs). Unexpectedly, DPY-21 also associates with autosomes of germ cells in a DCC-independent manner to enrich H4K20me1 and trigger chromosome compaction. Our findings demonstrate the direct link between chromatin modification and higher-order chromosome structure in long-range regulation of gene expression.

Non-coding Transcription Instructs Chromatin Folding and Compartmentalization to Dictate Enhancer-Promoter Communication and T Cell Fate.

  • Isoda T
  • Cell
  • 2017 Sep 21

Literature context:


Abstract:

It is now established that Bcl11b specifies T cell fate. Here, we show that in developing T cells, the Bcl11b enhancer repositioned from the lamina to the nuclear interior. Our search for factors that relocalized the Bcl11b enhancer identified a non-coding RNA named ThymoD (thymocyte differentiation factor). ThymoD-deficient mice displayed a block at the onset of T cell development and developed lymphoid malignancies. We found that ThymoD transcription promoted demethylation at CTCF bound sites and activated cohesin-dependent looping to reposition the Bcl11b enhancer from the lamina to the nuclear interior and to juxtapose the Bcl11b enhancer and promoter into a single-loop domain. These large-scale changes in nuclear architecture were associated with the deposition of activating epigenetic marks across the loop domain, plausibly facilitating phase separation. These data indicate how, during developmental progression and tumor suppression, non-coding transcription orchestrates chromatin folding and compartmentalization to direct with high precision enhancer-promoter communication.

Funding information:
  • NIAID NIH HHS - P01 AI102853()
  • NIAID NIH HHS - R01 AI082850()
  • NIAID NIH HHS - R01 AI100880()
  • NIAID NIH HHS - R01 AI109599()
  • NIDDK NIH HHS - U54 DK107977()

Enhancer Reprogramming Promotes Pancreatic Cancer Metastasis.

  • Roe JS
  • Cell
  • 2017 Aug 24

Literature context:


Abstract:

Pancreatic ductal adenocarcinoma (PDA) is one of the most lethal human malignancies, owing in part to its propensity for metastasis. Here, we used an organoid culture system to investigate how transcription and the enhancer landscape become altered during discrete stages of disease progression in a PDA mouse model. This approach revealed that the metastatic transition is accompanied by massive and recurrent alterations in enhancer activity. We implicate the pioneer factor FOXA1 as a driver of enhancer activation in this system, a mechanism that renders PDA cells more invasive and less anchorage-dependent for growth in vitro, as well as more metastatic in vivo. In this context, FOXA1-dependent enhancer reprogramming activates a transcriptional program of embryonic foregut endoderm. Collectively, our study implicates enhancer reprogramming, FOXA1 upregulation, and a retrograde developmental transition in PDA metastasis.

Myc Regulates Chromatin Decompaction and Nuclear Architecture during B Cell Activation.

  • Kieffer-Kwon KR
  • Mol. Cell
  • 2017 Aug 17

Literature context:


Abstract:

50 years ago, Vincent Allfrey and colleagues discovered that lymphocyte activation triggers massive acetylation of chromatin. However, the molecular mechanisms driving epigenetic accessibility are still unknown. We here show that stimulated lymphocytes decondense chromatin by three differentially regulated steps. First, chromatin is repositioned away from the nuclear periphery in response to global acetylation. Second, histone nanodomain clusters decompact into mononucleosome fibers through a mechanism that requires Myc and continual energy input. Single-molecule imaging shows that this step lowers transcription factor residence time and non-specific collisions during sampling for DNA targets. Third, chromatin interactions shift from long range to predominantly short range, and CTCF-mediated loops and contact domains double in numbers. This architectural change facilitates cognate promoter-enhancer contacts and also requires Myc and continual ATP production. Our results thus define the nature and transcriptional impact of chromatin decondensation and reveal an unexpected role for Myc in the establishment of nuclear topology in mammalian cells.

A UTX-MLL4-p300 Transcriptional Regulatory Network Coordinately Shapes Active Enhancer Landscapes for Eliciting Transcription.

  • Wang SP
  • Mol. Cell
  • 2017 Jul 20

Literature context:


Abstract:

Enhancer activation is a critical step for gene activation. Here we report an epigenetic crosstalk at enhancers between the UTX (H3K27 demethylase)-MLL4 (H3K4 methyltransferase) complex and the histone acetyltransferase p300. We demonstrate that UTX, in a demethylase activity-independent manner, facilitates conversion of inactive enhancers in embryonic stem cells to an active (H3K4me1+/H3K27ac+) state by recruiting and coupling the enzymatic functions of MLL4 and p300. Loss of UTX leads to attenuated enhancer activity, characterized by reduced levels of H3K4me1 and H3K27ac as well as impaired transcription. The UTX-MLL4 complex enhances p300-dependent H3K27 acetylation through UTX-dependent stimulation of p300 recruitment, while MLL4-mediated H3K4 monomethylation, reciprocally, requires p300 function. Importantly, MLL4-generated H3K4me1 further enhances p300-dependent transcription. This work reveals a previously unrecognized cooperativity among enhancer-associated chromatin modulators, including a unique function for UTX, in establishing an "active enhancer landscape" and defines a detailed mechanism for the joint deposition of H3K4me1 and H3K27ac.

Funding information:
  • NCI NIH HHS - R01 CA129325()
  • NCI NIH HHS - R01 CA178765()
  • NIDDK NIH HHS - R01 DK071900()

The Transcription Factor T-bet Limits Amplification of Type I IFN Transcriptome and Circuitry in T Helper 1 Cells.

  • Iwata S
  • Immunity
  • 2017 Jun 20

Literature context:


Abstract:

Host defense requires the specification of CD4+ helper T (Th) cells into distinct fates, including Th1 cells that preferentially produce interferon-γ (IFN-γ). IFN-γ, a member of a large family of anti-pathogenic and anti-tumor IFNs, induces T-bet, a lineage-defining transcription factor for Th1 cells, which in turn supports IFN-γ production in a feed-forward manner. Herein, we show that a cell-intrinsic role of T-bet influences how T cells perceive their secreted product in the environment. In the absence of T-bet, IFN-γ aberrantly induced a type I IFN transcriptomic program. T-bet preferentially repressed genes and pathways ordinarily activated by type I IFNs to ensure that its transcriptional response did not evoke an aberrant amplification of type I IFN signaling circuitry, otherwise triggered by its own product. Thus, in addition to promoting Th1 effector commitment, T-bet acts as a repressor in differentiated Th1 cells to prevent abberant autocrine type I IFN and downstream signaling.

Funding information:
  • Intramural NIH HHS - ZIA AI000579-27()
  • Intramural NIH HHS - ZIA AR041159-09()
  • Intramural NIH HHS - ZIA AR041167-09()

A Methyl-Balanced Diet Prevents CRF-Induced Prenatal Stress-Triggered Predisposition to Binge Eating-like Phenotype.

  • Schroeder M
  • Cell Metab.
  • 2017 Jun 6

Literature context:


Abstract:

Binge eating (BE) is a common aberrant form of eating behavior, characterized by overconsumption of food in a brief period of time. Recurrent episodes of BE constitute the BE disorder, which mostly affects females and is associated with early-life adversities. Here, we show that corticotropin releasing factor (CRF)-induced prenatal stress (PNS) in late gestation predisposes female offspring to BE-like behavior that coincides with hypomethylation of hypothalamic miR-1a and downstream dysregulation of the melanocortin system through Pax7/Pax3. Moreover, exposing the offspring to a methyl-balanced diet during adolescence prevents the dysregulation and predisposition from being triggered. We demonstrate that gestational programming, per se, will not lead to BE-like behavior, but pre-existing alterations due to prenatal programming are revealed only when challenged during adolescence. We provide experimental evidence for long-term epigenetic abnormalities stemming from PNS in predisposing female offspring to BE disorder as well as a potential non-invasive prevention strategy.

The Dynamic Epigenetic Landscape of the Retina During Development, Reprogramming, and Tumorigenesis.

  • Aldiri I
  • Neuron
  • 2017 May 3

Literature context:


Abstract:

In the developing retina, multipotent neural progenitors undergo unidirectional differentiation in a precise spatiotemporal order. Here we profile the epigenetic and transcriptional changes that occur during retinogenesis in mice and humans. Although some progenitor genes and cell cycle genes were epigenetically silenced during retinogenesis, the most dramatic change was derepression of cell-type-specific differentiation programs. We identified developmental-stage-specific super-enhancers and showed that most epigenetic changes are conserved in humans and mice. To determine how the epigenome changes during tumorigenesis and reprogramming, we performed integrated epigenetic analysis of murine and human retinoblastomas and induced pluripotent stem cells (iPSCs) derived from murine rod photoreceptors. The retinoblastoma epigenome mapped to the developmental stage when retinal progenitors switch from neurogenic to terminal patterns of cell division. The epigenome of retinoblastomas was more similar to that of the normal retina than that of retina-derived iPSCs, and we identified retina-specific epigenetic memory.

Funding information:
  • Howard Hughes Medical Institute - P30 CA021765()
  • NCI NIH HHS - R01 CA168875()
  • NEI NIH HHS - R01 EY014867()
  • NEI NIH HHS - R01 EY018599()
  • NEI NIH HHS - R01 EY023619()

Mll3 and Mll4 Facilitate Enhancer RNA Synthesis and Transcription from Promoters Independently of H3K4 Monomethylation.

  • Dorighi KM
  • Mol. Cell
  • 2017 May 18

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Abstract:

Monomethylation of histone H3 at lysine 4 (H3K4me1) and acetylation of histone H3 at lysine 27 (H3K27ac) are correlated with transcriptionally engaged enhancer elements, but the functional impact of these modifications on enhancer activity is not well understood. Here we used CRISPR/Cas9 genome editing to separate catalytic activity-dependent and independent functions of Mll3 (Kmt2c) and Mll4 (Kmt2d, Mll2), the major enhancer H3K4 monomethyltransferases. Loss of H3K4me1 from enhancers in Mll3/4 catalytically deficient cells causes partial reduction of H3K27ac, but has surprisingly minor effects on transcription from either enhancers or promoters. In contrast, loss of Mll3/4 proteins leads to strong depletion of enhancer Pol II occupancy and eRNA synthesis, concomitant with downregulation of target genes. Interestingly, downregulated genes exhibit reduced polymerase levels in gene bodies, but not at promoters, suggestive of pause-release defects. Altogether, our results suggest that enhancer H3K4me1 provides only a minor contribution to the long-range coactivator function of Mll3/4.

Dynamic Rewiring of Promoter-Anchored Chromatin Loops during Adipocyte Differentiation.

  • Siersbæk R
  • Mol. Cell
  • 2017 May 4

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Abstract:

Interactions between transcriptional promoters and their distal regulatory elements play an important role in transcriptional regulation; however, the extent to which these interactions are subject to rapid modulations in response to signals is unknown. Here, we use promoter capture Hi-C to demonstrate a rapid reorganization of promoter-anchored chromatin loops within 4 hr after inducing differentiation of 3T3-L1 preadipocytes. The establishment of new promoter-enhancer loops is tightly coupled to activation of poised (histone H3 lysine 4 mono- and dimethylated) enhancers, as evidenced by the acquisition of histone H3 lysine 27 acetylation and the binding of MED1, SMC1, and P300 proteins to these regions, as well as to activation of target genes. Intriguingly, formation of loops connecting activated enhancers and promoters is also associated with extensive recruitment of corepressors such as NCoR and HDACs, indicating that this class of coregulators may play a previously unrecognized role during enhancer activation.

Genomic Characterization of Murine Monocytes Reveals C/EBPβ Transcription Factor Dependence of Ly6C- Cells.

  • Mildner A
  • Immunity
  • 2017 May 16

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Abstract:

Monocytes are circulating, short-lived mononuclear phagocytes, which in mice and man comprise two main subpopulations. Murine Ly6C+ monocytes display developmental plasticity and are recruited to complement tissue-resident macrophages and dendritic cells on demand. Murine vascular Ly6C- monocytes patrol the endothelium, act as scavengers, and support vessel wall repair. Here we characterized population and single cell transcriptomes, as well as enhancer and promoter landscapes of the murine monocyte compartment. Single cell RNA-seq and transplantation experiments confirmed homeostatic default differentiation of Ly6C+ into Ly6C- monocytes. The main two subsets were homogeneous, but linked by a more heterogeneous differentiation intermediate. We show that monocyte differentiation occurred through de novo enhancer establishment and activation of pre-established (poised) enhancers. Generation of Ly6C- monocytes involved induction of the transcription factor C/EBPβ and C/EBPβ-deficient mice lacked Ly6C- monocytes. Mechanistically, C/EBPβ bound the Nr4a1 promoter and controlled expression of this established monocyte survival factor.

Funding information:
  • Intramural NIH HHS - ZIA BC010876-03(United States)

A Metabolic Function for Phospholipid and Histone Methylation.

  • Ye C
  • Mol. Cell
  • 2017 Apr 20

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Abstract:

S-adenosylmethionine (SAM) is the methyl donor for biological methylation modifications that regulate protein and nucleic acid functions. Here, we show that methylation of a phospholipid, phosphatidylethanolamine (PE), is a major consumer of SAM. The induction of phospholipid biosynthetic genes is accompanied by induction of the enzyme that hydrolyzes S-adenosylhomocysteine (SAH), a product and inhibitor of methyltransferases. Beyond its function for the synthesis of phosphatidylcholine (PC), the methylation of PE facilitates the turnover of SAM for the synthesis of cysteine and glutathione through transsulfuration. Strikingly, cells that lack PE methylation accumulate SAM, which leads to hypermethylation of histones and the major phosphatase PP2A, dependency on cysteine, and sensitivity to oxidative stress. Without PE methylation, particular sites on histones then become methyl sinks to enable the conversion of SAM to SAH. These findings reveal an unforeseen metabolic function for phospholipid and histone methylation intrinsic to the life of a cell.

Funding information:
  • NCI NIH HHS - P30 CA142543()
  • NIGMS NIH HHS - R01 GM094314()

YAP is essential for mechanical force production and epithelial cell proliferation during lung branching morphogenesis.

  • Lin C
  • Elife
  • 2017 Mar 21

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Abstract:

Branching morphogenesis is a fundamental program for tissue patterning. We show that active YAP, a key mediator of Hippo signaling, is distributed throughout the murine lung epithelium and loss of epithelial YAP severely disrupts branching. Failure to branch is restricted to regions where YAP activity is removed. This suggests that YAP controls local epithelial cell properties. In support of this model, mechanical force production is compromised and cell proliferation is reduced in Yap mutant lungs. We propose that defective force generation and insufficient epithelial cell number underlie the branching defects. Through genomic analysis, we also uncovered a feedback control of pMLC levels, which is critical for mechanical force production, likely through the direct induction of multiple regulators by YAP. Our work provides a molecular pathway that could control epithelial cell properties required for proper morphogenetic movement and pattern formation.

Funding information:
  • NINDS NIH HHS - K99 NS097627()

PHB Associates with the HIRA Complex to Control an Epigenetic-Metabolic Circuit in Human ESCs.

  • Zhu Z
  • Cell Stem Cell
  • 2017 Feb 2

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Abstract:

The chromatin landscape and cellular metabolism both contribute to cell fate determination, but their interplay remains poorly understood. Using genome-wide siRNA screening, we have identified prohibitin (PHB) as an essential factor in self-renewal of human embryonic stem cells (hESCs). Mechanistically, PHB forms protein complexes with HIRA, a histone H3.3 chaperone, and stabilizes the protein levels of HIRA complex components. Like PHB, HIRA is required for hESC self-renewal. PHB and HIRA act together to control global deposition of histone H3.3 and gene expression in hESCs. Of particular note, PHB and HIRA regulate the chromatin architecture at the promoters of isocitrate dehydrogenase genes to promote transcription and, thus, production of α-ketoglutarate, a key metabolite in the regulation of ESC fate. Our study shows that PHB has an unexpected nuclear role in hESCs that is required for self-renewal and that it acts with HIRA in chromatin organization to link epigenetic organization to a metabolic circuit.

Funding information:
  • NIAID NIH HHS - U01 AI095611(United States)

CpG Island Hypermethylation Mediated by DNMT3A Is a Consequence of AML Progression.

  • Spencer DH
  • Cell
  • 2017 Feb 23

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Abstract:

DNMT3A mutations occur in ∼25% of acute myeloid leukemia (AML) patients. The most common mutation, DNMT3AR882H, has dominant negative activity that reduces DNA methylation activity by ∼80% in vitro. To understand the contribution of DNMT3A-dependent methylation to leukemogenesis, we performed whole-genome bisulfite sequencing of primary leukemic and non-leukemic cells in patients with or without DNMT3AR882 mutations. Non-leukemic hematopoietic cells with DNMT3AR882H displayed focal methylation loss, suggesting that hypomethylation antedates AML. Although virtually all AMLs with wild-type DNMT3A displayed CpG island hypermethylation, this change was not associated with gene silencing and was essentially absent in AMLs with DNMT3AR882 mutations. Primary hematopoietic stem cells expanded with cytokines were hypermethylated in a DNMT3A-dependent manner, suggesting that hypermethylation may be a response to, rather than a cause of, cellular proliferation. Our findings suggest that hypomethylation is an initiating phenotype in AMLs with DNMT3AR882, while DNMT3A-dependent CpG island hypermethylation is a consequence of AML progression.

Funding information:
  • NCI NIH HHS - K08 CA166229()
  • NCI NIH HHS - K08 CA190815()
  • NCI NIH HHS - P01 CA101937()
  • NCI NIH HHS - P30 CA091842()
  • NCI NIH HHS - R35 CA197561()

Epigenetic Activation of WNT5A Drives Glioblastoma Stem Cell Differentiation and Invasive Growth.

  • Hu B
  • Cell
  • 2016 Nov 17

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Abstract:

Glioblastoma stem cells (GSCs) are implicated in tumor neovascularization, invasiveness, and therapeutic resistance. To illuminate mechanisms governing these hallmark features, we developed a de novo glioblastoma multiforme (GBM) model derived from immortalized human neural stem/progenitor cells (hNSCs) to enable precise system-level comparisons of pre-malignant and oncogene-induced malignant states of NSCs. Integrated transcriptomic and epigenomic analyses uncovered a PAX6/DLX5 transcriptional program driving WNT5A-mediated GSC differentiation into endothelial-like cells (GdECs). GdECs recruit existing endothelial cells to promote peritumoral satellite lesions, which serve as a niche supporting the growth of invasive glioma cells away from the primary tumor. Clinical data reveal higher WNT5A and GdECs expression in peritumoral and recurrent GBMs relative to matched intratumoral and primary GBMs, respectively, supporting WNT5A-mediated GSC differentiation and invasive growth in disease recurrence. Thus, the PAX6/DLX5-WNT5A axis governs the diffuse spread of glioma cells throughout the brain parenchyma, contributing to the lethality of GBM.

Funding information:
  • NINDS NIH HHS - R56 NS094589(United States)

Transcriptional Networks Controlled by NKX2-1 in the Development of Forebrain GABAergic Neurons.

  • Sandberg M
  • Neuron
  • 2016 Sep 21

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Abstract:

The embryonic basal ganglia generates multiple projection neurons and interneuron subtypes from distinct progenitor domains. Combinatorial interactions of transcription factors and chromatin are thought to regulate gene expression. In the medial ganglionic eminence, the NKX2-1 transcription factor controls regional identity and, with LHX6, is necessary to specify pallidal projection neurons and forebrain interneurons. Here, we dissected the molecular functions of NKX2-1 by defining its chromosomal binding, regulation of gene expression, and epigenetic state. NKX2-1 binding at distal regulatory elements led to a repressed epigenetic state and transcriptional repression in the ventricular zone. Conversely, NKX2-1 is required to establish a permissive chromatin state and transcriptional activation in the sub-ventricular and mantle zones. Moreover, combinatorial binding of NKX2-1 and LHX6 promotes transcriptionally permissive chromatin and activates genes expressed in cortical migrating interneurons. Our integrated approach provides a foundation for elucidating transcriptional networks guiding the development of the MGE and its descendants.

Pioneering Activity of the C-Terminal Domain of EBF1 Shapes the Chromatin Landscape for B Cell Programming.

  • Boller S
  • Immunity
  • 2016 Mar 15

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Abstract:

Lymphopoiesis requires the activation of lineage-specific genes embedded in naive, inaccessible chromatin or in primed, accessible chromatin. The mechanisms responsible for de novo gain of chromatin accessibility, known as "pioneer" function, remain poorly defined. Here, we showed that the EBF1 C-terminal domain (CTD) is required for the regulation of a specific gene set involved in B cell fate decision and differentiation, independently of activation and repression functions. Using genome-wide analysis of DNaseI hypersensitivity and DNA methylation in multipotent Ebf1(-/-) progenitors and derivative EBF1wt- or EBF1ΔC-expressing cells, we found that the CTD promoted chromatin accessibility and DNA demethylation in previously naive chromatin. The CTD allowed EBF1 to bind at inaccessible genomic regions that offer limited co-occupancy by other transcription factors, whereas the CTD was dispensable for EBF1 binding at regions that are occupied by multiple transcription factors. Thus, the CTD enables EBF1 to confer permissive lineage-specific changes in progenitor chromatin landscape.

Registered report: Oncometabolite 2-hydroxyglutarate is a competitive inhibitor of α-ketoglutarate-dependent dioxygenases.

  • Evans B
  • Elife
  • 2015 Jul 31

Literature context:


Abstract:

The Reproducibility Project: Cancer Biology seeks to address growing concerns about reproducibility in scientific research by conducting replications of selected experiments from a number of high-profile papers in the field of cancer biology. The papers, which were published between 2010 and 2012, were selected on the basis of citations and Altmetric scores (Errington et al., 2014). This Registered report describes the proposed replication plan of key experiments from 'Oncometabolite 2-hydroxyglutarate is a competitive inhibitor of α-ketoglutarate-dependent dioxygenases' by Xu and colleagues, published in Cancer Cell in 2011 (Xu et al., 2011). The key experiments being replicated include Supplemental Figure 3I, which demonstrates that transfection with mutant forms of IDH1 increases levels of 2-hydroxyglutarate (2-HG), Figures 3A and 8A, which demonstrate changes in histone methylation after treatment with 2-HG, and Figures 3D and 7B, which show that mutant IDH1 can effect the same changes as treatment with excess 2-HG. The Reproducibility Project: Cancer Biology is a collaboration between the Center for Open Science and Science Exchange, and the results of the replications will be published by eLife.

Funding information:
  • NINDS NIH HHS - N01NS02331(United States)