Histone H3.3 K27M Accelerates Spontaneous Brainstem Glioma and Drives Restricted Changes in Bivalent Gene Expression

Larson, Jon D.; Kasper, Lawryn H.; Paugh, Barbara S.; Jin, H; Wu, Gang; Kwon, Chang‐Hyuk; Fan, Yiping; Shaw, Timothy I.; Silveira, André B.; Qu, Chunxu; Xu, Raymond; Zhu, Xiaoyan; Zhang, Junyuan; Russell, H. R.; Peters, Jennifer L.; Finkelstein, David; Xu, Beisi; Lin, Tong; Tinkle, Christopher L.; Patay, Zoltán; Onar‐Thomas, Arzu; Pounds, Stanley; McKinnon, Peter J.; Ellison, David W.; Zhang, Jinghui; Baker, Suzanne J.

doi:10.1016/j.ccell.2018.11.015

Cited by 214 publications

(212 citation statements)

References 58 publications

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“…Though some H3K27me3 peaks disappeared completely in EZHIP and H3.3 K27M cells, we noted a striking difference in distribution of H3K27me3 among the peaks that remained; while H3K27me3 is usually found in broad domains in wildtype cells, we noted that the peak width decreased substantially in cells expressing EZHIP or H3.3 K27M ( Figure 4A, B). Similar changes in H3K27me3 ChIP-seq profiles have been observed in DIPG cells containing the H3 K27M mutant histone 8, [15][16][17][18][19] .…”

Section: Ezhip Expression Leads To K27m-like Genomic Distribution Ofsupporting

confidence: 73%

“…We plotted the log2 expression profile of genes in PFA and supratentorial ependymomas to identify gene expression changes possibly caused by EZHIP expression ( Figure 5D, S5A) Using STRING analysis, we found enrichment of several gene ontology terms related to organism development and cell differentiation ( Figure S5B). Earlier studies found that K27Mcontaining DIPGs exhibit global reduction in H3K27me3 and retention of H3K27me3 peaks at many genomic loci 8, [15][16][17][18][19] . The CDKN2A tumor suppressor gene is one locus that is silenced in a H3K27me3-dependent manner in K27M-containing DIPG cell lines and in H3.3 K27M glioma model systems 18,41,42 .…”

Section: Ependymomas Expressing Ezhip Display Aberrant Silencing Of Cmentioning

confidence: 93%

“…The H3 K27M oncohistone binds to and inhibits the catalytic subunit (EZH2) of the Polycomb Repressive Complex 2 (PRC2), a conserved protein complex involved in gene silencing [8][9][10][11][12][13][14] . Genome-wide profiling of H3K27me3 in H3 K27Mcontaining DIPG tumors and transgenic cell lines revealed a non-uniform reduction of H3K27me3 and, surprisingly, locus-specific K27M-dependent retention of this histone modification 8, [15][16][17][18][19] . While mechanistic questions related to H3 K27M remain, the unprecedented finding that oncohistones act as enzyme inhibitors and alter global levels of histone modifications in cells indicates a direct effect of chromatin misregulation driven by histone mutations in tumorigenesis.…”

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confidence: 98%

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PFA ependymoma-associated protein EZHIP inhibits PRC2 activity through a H3 K27M-like mechanism

Jain

Truman

Lund

et al. 2019

Preprint

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Polycomb group (PcG) proteins are essential for development and are frequently misregulated in human cancers. Polycomb Repressive Complexes (PRC1, PRC2) function in a collaborative epigenetic cross-talk with H3K27me3 to initiate and maintain transcriptional silencing. Diffuse intrinsic pontine gliomas (DIPGs) have extremely low H3K27me3 levels mediated by H3 K27M oncohistone. Posterior fossa type A (PFA) ependymomas also exhibit very low H3K27 methylation but lack the K27M oncohistone. Instead, PFA tumors express high levels of EZHIP (Enhancer of Zeste Homologs Inhibitory Protein, also termed CXORF67). We find that a highly conserved sequence within the C-terminus of EZHIP is necessary and sufficient to inhibit the catalytic activity of PRC2 in vitro and in vivo. Our biochemical experiments indicate that EZHIP directly interacts with the active site of the EZH2 subunit in a mechanism that is remarkably similar to the K27M oncohistone. Furthermore, expression of H3 K27M or EZHIP in cells promote similar chromatin profiles: loss of broad H3K27me3 domains, but retention of H3K27me3 at the sites of PRC2 recruitment. Importantly, we find that H3K27me3-mediated allosteric activation of PRC2 substantially increases the inhibition potential of EZHIP and H3 K27M, providing a potential mechanism for loss of H3K27me3 spreading from CpG islands in vivo. Our data indicate that PFA ependymoma and DIPG are driven in part by the action of peptidyl PRC2 inhibitors-the K27M oncohistone and the EZHIP 'oncohistone-mimic'-that dysregulate gene silencing to promote tumorigenesis. Covalent modifications to both DNA and histone proteins allow chromatin to act as a dynamic information hub that integrates diverse biochemical stimuli to regulate genomic DNA access to the transcription machinery and ultimately establish and maintain cellular phenotypes. There is increasing appreciation that aberrant chromatin modifications are involved in the pathogenesis of cancer. Nowhere is this better supported than with the groundbreaking discoveries of highfrequency, somatic mutations in histones that are drivers of oncogenesis.

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Section: Ezhip Expression Leads To K27m-like Genomic Distribution Ofsupporting

confidence: 73%

Section: Ependymomas Expressing Ezhip Display Aberrant Silencing Of Cmentioning

confidence: 93%

mentioning

confidence: 98%

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PFA ependymoma-associated protein EZHIP inhibits PRC2 activity through a H3 K27M-like mechanism

Jain

Truman

Lund

et al. 2019

Preprint

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“…These compounds, however, have proved challenging to adapt for clinical use. The relatively nonspecific approach of inhibiting genome-wide transcription may also impart off-target consequences given the comparatively restricted transcriptomic changes observed in H3K27M-mutant tumors (21). In this study, we set out to identify specific epigenetic regulators which represent critical cellular dependencies within the context of an H3K27M+ transformed cell state , to understand how they might mediate the altered developmental phenotype observed in DMG cell populations, and to determine whether these represented vulnerabilities that might be amenable to disruption using clinically relevant compounds.…”

Section: Introductionmentioning

confidence: 99%

Super elongation complex as a targetable dependency in diffuse midline glioma

Dahl

Danis

Balakrishnan

et al. 2020

Preprint

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AbstractMutations in the histone 3 gene (H3K27M) are the eponymous drivers in diffuse intrinsic pontine gliomas (DIPGs) and other diffuse midline gliomas (DMGs), aggressive pediatric brain cancers for which no curative therapy currently exists. The salient molecular consequence of these recurrent oncohistones is a global loss of repressive H3K27me3 residues and broad epigenetic dysregulation. In order to identify specific, therapeutically targetable epigenetic dependencies within this disease context, we performed an shRNA screen targeting 408 genes classified as epigenetic/chromatin-associated molecules in patient-derived DMG cultures. This approach identified AFF4, the scaffold protein of the super elongation complex (SEC), as a previously-undescribed dependency in DMG. Interrogation of SEC function demonstrated a key role for maintaining DMG cell viability and clonogenic potential while promoting self-renewal of DMG tumor stem cells. Small-molecule inhibition of the SEC with the highly-specific, clinically relevant CDK9 inhibitors atuveciclib and AZD4573 restores regulatory RNA polymerase II pausing, promotes cellular differentiation, and leads to potent anti-tumor effect both in vitro and in patient-derived xenograft models. These studies present a biologic rationale for translational exploration of CDK9 inhibition as a promising therapeutic approach in a disease which currently has no effective medical therapies.

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“…This approach could be particularly amenable to studying other poorly-druggable oncogenes involving transcriptional and epigenetic remodeling factors, including fusion oncogenes (e.g. MLL-fusions(23) and NUP98-fusions(24)) in hematopoietic malignancies, but also in others cancers, including ependymoma(25) and H3.3 mutants glioma (26).…”

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confidence: 99%

Screening of clustered regulatory elements reveals functional cooperating dependencies in Leukemia

Benbarche

Lopez

Salataj

et al. 2019

Preprint

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In the recent years, massively parallel sequencing approaches identified hundreds of mutated genes in cancer(1) providing an unprecedented amount of information about mechanisms of cancer cell maintenance and progression. However, while (it is widely accepted that) transformation processes result from oncogenic cooperation between deregulated genes and pathways, the functional characterization of candidate key players is mostly performed at the single gene level which is generally inadequate to identify these oncogene circuitries. In addition, studies aimed at depicting oncogenic cooperation involve the generation of challenging mouse models or the deployment of tedious screening pipelines. Genome wide mapping of epigenomic modifications on histone tails or binding of factors such as MED1 and BRD4 allowed identification of clusters of regulatory elements, also termed Super-Enhancers (SE)(2). Functional annotation of these regions revealed their high relevance during normal tissue development and cancer ontogeny(3). An interesting paradigm of the tumorigenic function of these SE regions comes from ETO2-GLIS2-driven acute megakaryoblastic leukemia (AMKL) in which the fusion protein ETO2-GLIS2 is sufficient to promote an aberrant transcriptional network by the rewiring of SE regions(4). We thus hypothesized that important regulatory regions could control simultaneously expression of genes cooperating in functional modules to promote cancer development. In an effort to identify such modules, we deployed a genome-wide CRISPRi-based screening approach and nominated SE regions that are functionally linked to leukemia maintenance. In particular, we pinpointed a novel SE region regulating the expression of both tyrosine kinases KIT and PDGFRA. Whereas the inhibition of each kinase alone affected modestly cancer cell growth, combined inhibition of both receptors synergizes to impair leukemia cell growth and survival. Our results demonstrate that genome-wide screening of regulatory DNA elements can identify co-regulated genes collaborating to promote cancer and could open new avenues to the concept of combined gene inhibition upon single hit targeting.Super-Enhancers (SE) are clusters of regulatory elements characterized by high intensity of enhancer-related histone tail modifications such as histone H3 lysin 27 acetylation or lysine 4 mono-methylation (H3K27ac and H3K4me1 respectively) and binding of enhancerassociated factors such as the mediator complex, in particular MED1, or bromodomaincontaining proteins such as BRD4(5). These clustered regulatory regions shape the transcriptional identity of specific tissues and cell types and their landscape often shifts in disease conditions particularly in cancer cells where they can control oncogene expression (6, 7). Whether these regions control expression of one or several genes and what are the genes directly controlled by these regions is mainly unknown. Additionally, whether these regions contribute to transformation or disease progression remains to be characterized. We hy...

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Histone H3.3 K27M Accelerates Spontaneous Brainstem Glioma and Drives Restricted Changes in Bivalent Gene Expression

Cited by 214 publications

References 58 publications

PFA ependymoma-associated protein EZHIP inhibits PRC2 activity through a H3 K27M-like mechanism

PFA ependymoma-associated protein EZHIP inhibits PRC2 activity through a H3 K27M-like mechanism

Super elongation complex as a targetable dependency in diffuse midline glioma

Screening of clustered regulatory elements reveals functional cooperating dependencies in Leukemia

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