Publications by authors named "Jens Vilstrup Johansen"

23 Publications

  • Page 1 of 1

Acute Experimental Barrier Injury Triggers Ulcerative Colitis-Specific Innate Hyperresponsiveness and Ulcerative Colitis-Type Microbiome Changes in Humans.

Cell Mol Gastroenterol Hepatol 2021 9;12(4):1281-1296. Epub 2021 Jun 9.

Department of Gastroenterology, Herlev Hospital, University of Copenhagen, Copenhagen, Denmark.

Background And Aims: The trigger hypothesis opens the possibility of anti-flare initiation therapies by stating that ulcerative colitis (UC) flares originate from inadequate responses to acute mucosal injuries. However, experimental evidence is restricted by a limited use of suitable human models. We thus aimed to investigate the acute mucosal barrier injury responses in humans with and without UC using an experimental injury model.

Methods: A standardized mucosal break was inflicted in the sigmoid colon of 19 patients with UC in endoscopic and histological remission and 20 control subjects. Postinjury responses were assessed repeatedly by high-resolution imaging and sampling to perform Geboes scoring, RNA sequencing, and injury niche microbiota 16S ribosomal RNA gene sequencing.

Results: UC patients had more severe endoscopic postinjury inflammation than did control subjects (P < .01), an elevated modified Geboes score (P < .05), a rapid induction of innate response gene sets (P < .05) and antimicrobial peptides (P < .01), and engagement of neutrophils (P < .01). Innate lymphoid cell type 3 (ILC3) markers were increased preinjury (P < .01), and ILC3 activating cytokines were highly induced postinjury, resulting in an increase in ILC3-type cytokine interleukin-17A. Across groups, the postinjury mucosal microbiome had higher bacterial load (P < .0001) and lower α-diversity (P < .05).

Conclusions: UC patients in remission respond to mucosal breaks by an innate hyperresponse engaging resident regulatory ILC3s and a subsequent adaptive activation. The postinjury inflammatory bowel disease-like microbiota diversity decrease is irrespective of diagnosis, suggesting that the dysbiosis is secondary to host injury responses. We provide a model for the study of flare initiation in the search for antitrigger-directed therapies.
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http://dx.doi.org/10.1016/j.jcmgh.2021.06.002DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8455368PMC
June 2021

Effects of a single dose of psilocybin on behaviour, brain 5-HT receptor occupancy and gene expression in the pig.

Eur Neuropsychopharmacol 2021 01 4;42:1-11. Epub 2020 Dec 4.

Neurobiology Research Unit 8057 and The Center for Experimental Medicine Neuropharmacology, Copenhagen University Hospital Rigshospitalet, Blegdamsvej 9, 2100 Copenhagen O, Denmark; Faculty of Health and Medical Sciences, University of Copenhagen, 2200 Copenhagen N, Denmark. Electronic address:

Psilocybin has in some studies shown promise as treatment of major depressive disorder and psilocybin therapy was in 2019 twice designated as breakthrough therapy by the U.S. Food and Drug Administration (FDA). A very particular feature is that ingestion of just a single dose of psilocybin is associated with lasting changes in personality and mood. The underlying molecular mechanism behind its effect is, however, unknown. In a translational pig model, we here present the effects of a single dose of psilocybin on pig behaviour, receptor occupancy and gene expression in the brain. An acute i.v. injection of 0.08 mg/kg psilocybin to awake female pigs induced characteristic behavioural changes in terms of headshakes, scratching and rubbing, lasting around 20 min. A similar dose was associated with a cerebral 5-HT receptor occupancy of 67%, as determined by positron emission tomography, and plasma psilocin levels were comparable to what in humans is associated with an intense psychedelic experience. We found that 19 genes were differentially expressed in prefrontal cortex one day after psilocybin injection, and 3 genes after 1 week. Gene Set Enrichment Analysis demonstrated that multiple immunological pathways were regulated 1 week after psilocybin exposure. This provides a framework for future investigations of the lasting molecular mechanisms induced by a single dose of psilocybin. In the light of an ongoing debate as to whether psilocybin is a safe treatment for depression and other mental illnesses, it is reassuring that our data suggest that any effects on gene expression are very modest.
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http://dx.doi.org/10.1016/j.euroneuro.2020.11.013DOI Listing
January 2021

KDM4A regulates the maternal-to-zygotic transition by protecting broad H3K4me3 domains from H3K9me3 invasion in oocytes.

Nat Cell Biol 2020 04 30;22(4):380-388. Epub 2020 Mar 30.

DNRF Center for Chromosome Stability (CCS), Department of Cellular and Molecular Medicine, Faculty of Health and Medical Sciences, University of Copenhagen, Copenhagen, Denmark.

The importance of germline-inherited post-translational histone modifications on priming early mammalian development is just emerging. Histone H3 lysine 9 (H3K9) trimethylation is associated with heterochromatin and gene repression during cell-fate change, whereas histone H3 lysine 4 (H3K4) trimethylation marks active gene promoters. Mature oocytes are transcriptionally quiescent and possess remarkably broad domains of H3K4me3 (bdH3K4me3). It is unknown which factors contribute to the maintenance of the bdH3K4me3 landscape. Lysine-specific demethylase 4A (KDM4A) demethylates H3K9me3 at promoters marked by H3K4me3 in actively transcribing somatic cells. Here, we report that KDM4A-mediated H3K9me3 demethylation at bdH3K4me3 in oocytes is crucial for normal pre-implantation development and zygotic genome activation after fertilization. The loss of KDM4A in oocytes causes aberrant H3K9me3 spreading over bdH3K4me3, resulting in insufficient transcriptional activation of genes, endogenous retroviral elements and chimeric transcripts initiated from long terminal repeats during zygotic genome activation. The catalytic activity of KDM4A is essential for normal epigenetic reprogramming and pre-implantation development. Hence, KDM4A plays a crucial role in preserving the maternal epigenome integrity required for proper zygotic genome activation and transfer of developmental control to the embryo.
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http://dx.doi.org/10.1038/s41556-020-0494-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7212036PMC
April 2020

PLZF targets developmental enhancers for activation during osteogenic differentiation of human mesenchymal stem cells.

Elife 2019 01 23;8. Epub 2019 Jan 23.

Biotech Research and Innovation Centre (BRIC), Faculty of Health and Medical Sciences, University of Copenhagen, Copenhagen, Denmark.

The PLZF transcription factor is essential for osteogenic differentiation of hMSCs; however, its regulation and molecular function during this process is not fully understood. Here, we revealed that the locus encoding PLZF, is repressed by Polycomb (PcG) and H3K27me3 in naive hMSCs. At the pre-osteoblast stage of differentiation, the locus lost PcG binding and H3K27me3, gained JMJD3 recruitment, and H3K27ac resulting in high expression of PLZF. Subsequently, PLZF was recruited to osteogenic enhancers, influencing H3K27 acetylation and expression of nearby genes important for osteogenic function. Furthermore, we identified a latent enhancer within the locus itself that became active, gained PLZF, p300 and Mediator binding and looped to the promoter of the nicotinamide N-methyltransferase () gene. The increased expression of NNMT correlated with a decline in SAM levels, which is dependent on PLZF and is required for osteogenic differentiation.
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http://dx.doi.org/10.7554/eLife.40364DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6344081PMC
January 2019

Histone H4K20 methylation mediated chromatin compaction threshold ensures genome integrity by limiting DNA replication licensing.

Nat Commun 2018 09 12;9(1):3704. Epub 2018 Sep 12.

Biotech Research and Innovation Centre (BRIC), Faculty of Health and Medical Sciences, University of Copenhagen, Ole Maaløes Vej 5, 2200, Copenhagen N, Denmark.

The decompaction and re-establishment of chromatin organization immediately after mitosis is essential for genome regulation. Mechanisms underlying chromatin structure control in daughter cells are not fully understood. Here we show that a chromatin compaction threshold in cells exiting mitosis ensures genome integrity by limiting replication licensing in G1 phase. Upon mitotic exit, chromatin relaxation is controlled by SET8-dependent methylation of histone H4 on lysine 20. In the absence of either SET8 or H4K20 residue, substantial genome-wide chromatin decompaction occurs allowing excessive loading of the origin recognition complex (ORC) in the daughter cells. ORC overloading stimulates aberrant recruitment of the MCM2-7 complex that promotes single-stranded DNA formation and DNA damage. Restoring chromatin compaction restrains excess replication licensing and loss of genome integrity. Our findings identify a cell cycle-specific mechanism whereby fine-tuned chromatin relaxation suppresses excessive detrimental replication licensing and maintains genome integrity at the cellular transition from mitosis to G1 phase.
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http://dx.doi.org/10.1038/s41467-018-06066-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6135857PMC
September 2018

Accurate Recycling of Parental Histones Reproduces the Histone Modification Landscape during DNA Replication.

Mol Cell 2018 10 23;72(2):239-249.e5. Epub 2018 Aug 23.

Biotech Research and Innovation Centre (BRIC), University of Copenhagen, Faculty of Health and Medical Sciences, University of Copenhagen, 2200 Copenhagen, Denmark; The Novo Nordisk Center for Protein Research (CPR), University of Copenhagen, Faculty of Health and Medical Sciences, University of Copenhagen, 2200 Copenhagen, Denmark. Electronic address:

Chromatin organization is disrupted genome-wide during DNA replication. On newly synthesized DNA, nucleosomes are assembled from new naive histones and old modified histones. It remains unknown whether the landscape of histone post-translational modifications (PTMs) is faithfully copied during DNA replication or the epigenome is perturbed. Here we develop chromatin occupancy after replication (ChOR-seq) to determine histone PTM occupancy immediately after DNA replication and across the cell cycle. We show that H3K4me3, H3K36me3, H3K79me3, and H3K27me3 positional information is reproduced with high accuracy on newly synthesized DNA through histone recycling. Quantitative ChOR-seq reveals that de novo methylation to restore H3K4me3 and H3K27me3 levels occurs across the cell cycle with mark- and locus-specific kinetics. Collectively, this demonstrates that accurate parental histone recycling preserves positional information and allows PTM transmission to daughter cells while modification of new histones gives rise to complex epigenome fluctuations across the cell cycle that could underlie cell-to-cell heterogeneity.
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http://dx.doi.org/10.1016/j.molcel.2018.08.010DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6202308PMC
October 2018

EZH2 is a potential therapeutic target for H3K27M-mutant pediatric gliomas.

Nat Med 2017 Apr 27;23(4):483-492. Epub 2017 Feb 27.

Biotech Research and Innovation Centre (BRIC), University of Copenhagen, Copenhagen, Denmark.

Diffuse intrinsic pontine glioma (DIPG) is an aggressive brain tumor that is located in the pons and primarily affects children. Nearly 80% of DIPGs harbor mutations in histone H3 genes, wherein lysine 27 is substituted with methionine (H3K27M). H3K27M has been shown to inhibit polycomb repressive complex 2 (PRC2), a multiprotein complex responsible for the methylation of H3 at lysine 27 (H3K27me), by binding to its catalytic subunit EZH2. Although DIPGs with the H3K27M mutation show global loss of H3K27me3, several genes retain H3K27me3. Here we describe a mouse model of DIPG in which H3K27M potentiates tumorigenesis. Using this model and primary patient-derived DIPG cell lines, we show that H3K27M-expressing tumors require PRC2 for proliferation. Furthermore, we demonstrate that small-molecule EZH2 inhibitors abolish tumor cell growth through a mechanism that is dependent on the induction of the tumor-suppressor protein p16. Genome-wide enrichment analyses show that the genes that retain H3K27me3 in H3K27M cells are strong polycomb targets. Furthermore, we find a highly significant overlap between genes that retain H3K27me3 in the DIPG mouse model and in human primary DIPGs expressing H3K27M. Taken together, these results show that residual PRC2 activity is required for the proliferation of H3K27M-expressing DIPGs, and that inhibition of EZH2 is a potential therapeutic strategy for the treatment of these tumors.
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http://dx.doi.org/10.1038/nm.4293DOI Listing
April 2017

JMJD-5/KDM8 regulates H3K36me2 and is required for late steps of homologous recombination and genome integrity.

PLoS Genet 2017 02 16;13(2):e1006632. Epub 2017 Feb 16.

Biotech Research and Innovation Centre (BRIC), University of Copenhagen, Copenhagen, Denmark.

The eukaryotic genome is organized in a three-dimensional structure called chromatin, constituted by DNA and associated proteins, the majority of which are histones. Post-translational modifications of histone proteins greatly influence chromatin structure and regulate many DNA-based biological processes. Methylation of lysine 36 of histone 3 (H3K36) is a post-translational modification functionally relevant during early steps of DNA damage repair. Here, we show that the JMJD-5 regulates H3K36 di-methylation and it is required at late stages of double strand break repair mediated by homologous recombination. Loss of jmjd-5 results in hypersensitivity to ionizing radiation and in meiotic defects, and it is associated with aberrant retention of RAD-51 at sites of double strand breaks. Analyses of jmjd-5 genetic interactions with genes required for resolving recombination intermediates (rtel-1) or promoting the resolution of RAD-51 double stranded DNA filaments (rfs-1 and helq-1) suggest that jmjd-5 prevents the formation of stalled postsynaptic recombination intermediates and favors RAD-51 removal. As these phenotypes are all recapitulated by a catalytically inactive jmjd-5 mutant, we propose a novel role for H3K36me2 regulation during late steps of homologous recombination critical to preserve genome integrity.
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http://dx.doi.org/10.1371/journal.pgen.1006632DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5336306PMC
February 2017

Continual removal of H3K9 promoter methylation by Jmjd2 demethylases is vital for ESC self-renewal and early development.

EMBO J 2016 07 6;35(14):1550-64. Epub 2016 Jun 6.

Biotech Research and Innovation Centre (BRIC), University of Copenhagen, Copenhagen, Denmark Centre for Epigenetics, University of Copenhagen, Copenhagen, Denmark The Danish Stem Cell Center (Danstem), Faculty of Health and Medical Sciences, University of Copenhagen, Copenhagen, Denmark

Chromatin-associated proteins are essential for the specification and maintenance of cell identity. They exert these functions through modulating and maintaining transcriptional patterns. To elucidate the functions of the Jmjd2 family of H3K9/H3K36 histone demethylases, we generated conditional Jmjd2a/Kdm4a, Jmjd2b/Kdm4b and Jmjd2c/Kdm4c/Gasc1 single, double and triple knockout mouse embryonic stem cells (ESCs). We report that while individual Jmjd2 family members are dispensable for ESC maintenance and embryogenesis, combined deficiency for specifically Jmjd2a and Jmjd2c leads to early embryonic lethality and impaired ESC self-renewal, with spontaneous differentiation towards primitive endoderm under permissive culture conditions. We further show that Jmjd2a and Jmjd2c both localize to H3K4me3-positive promoters, where they have widespread and redundant roles in preventing accumulation of H3K9me3 and H3K36me3. Jmjd2 catalytic activity is required for ESC maintenance, and increased H3K9me3 levels in knockout ESCs compromise the expression of several Jmjd2a/c targets, including genes that are important for ESC self-renewal. Thus, continual removal of H3K9 promoter methylation by Jmjd2 demethylases represents a novel mechanism ensuring transcriptional competence and stability of the pluripotent cell identity.
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http://dx.doi.org/10.15252/embj.201593317DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4899663PMC
July 2016

Jmjd2/Kdm4 demethylases are required for expression of Il3ra and survival of acute myeloid leukemia cells.

Genes Dev 2016 06 2;30(11):1278-88. Epub 2016 Jun 2.

Biotech Research and Innovation Centre (BRIC), University of Copenhagen, 2200 Copenhagen, Denmark; Centre for Epigenetics, University of Copenhagen, 2200 Copenhagen, Denmark; The Danish Stem Cell Center (DanStem), Faculty of Health and Medical Sciences, University of Copenhagen, 2200 Copenhagen, Denmark.

Acute myeloid leukemias (AMLs) with a rearrangement of the mixed-linage leukemia (MLL) gene are aggressive hematopoietic malignancies. Here, we explored the feasibility of using the H3K9- and H3K36-specific demethylases Jmjd2/Kdm4 as putative drug targets in MLL-AF9 translocated leukemia. Using Jmjd2a, Jmjd2b, and Jmjd2c conditional triple-knockout mice, we show that Jmjd2/Kdm4 activities are required for MLL-AF9 translocated AML in vivo and in vitro. We demonstrate that expression of the interleukin 3 receptor α (Il3ra also known as Cd123) subunit is dependent on Jmjd2/Kdm4 through a mechanism involving removal of H3K9me3 from the promoter of the Il3ra gene. Importantly, ectopic expression of Il3ra in Jmjd2/Kdm4 knockout cells alleviates the requirement of Jmjd2/Kdm4 for the survival of AML cells, showing that Il3ra is a critical downstream target of Jmjd2/Kdm4 in leukemia. These results suggest that the JMJD2/KDM4 proteins are promising drug targets for the treatment of AML.
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http://dx.doi.org/10.1101/gad.280495.116DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4911927PMC
June 2016

An interactive environment for agile analysis and visualization of ChIP-sequencing data.

Nat Struct Mol Biol 2016 Apr 29;23(4):349-57. Epub 2016 Feb 29.

Biotech Research and Innovation Centre (BRIC), University of Copenhagen, Copenhagen, Denmark.

To empower experimentalists with a means for fast and comprehensive chromatin immunoprecipitation sequencing (ChIP-seq) data analyses, we introduce an integrated computational environment, EaSeq. The software combines the exploratory power of genome browsers with an extensive set of interactive and user-friendly tools for genome-wide abstraction and visualization. It enables experimentalists to easily extract information and generate hypotheses from their own data and public genome-wide datasets. For demonstration purposes, we performed meta-analyses of public Polycomb ChIP-seq data and established a new screening approach to analyze more than 900 datasets from mouse embryonic stem cells for factors potentially associated with Polycomb recruitment. EaSeq, which is freely available and works on a standard personal computer, can substantially increase the throughput of many analysis workflows, facilitate transparency and reproducibility by automatically documenting and organizing analyses, and enable a broader group of scientists to gain insights from ChIP-seq data.
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http://dx.doi.org/10.1038/nsmb.3180DOI Listing
April 2016

SWI/SNF Subunits SMARCA4, SMARCD2 and DPF2 Collaborate in MLL-Rearranged Leukaemia Maintenance.

PLoS One 2015 16;10(11):e0142806. Epub 2015 Nov 16.

Biotech Research and Innovation Centre (BRIC), University of Copenhagen, Ole Maaløes Vej 5, 2200 Copenhagen, Denmark.

Alterations in chromatin structure caused by deregulated epigenetic mechanisms collaborate with underlying genetic lesions to promote cancer. SMARCA4/BRG1, a core component of the SWI/SNF ATP-dependent chromatin-remodelling complex, has been implicated by its mutational spectrum as exerting a tumour-suppressor function in many solid tumours; recently however, it has been reported to sustain leukaemogenic transformation in MLL-rearranged leukaemia in mice. Here we further explore the role of SMARCA4 and the two SWI/SNF subunits SMARCD2/BAF60B and DPF2/BAF45D in leukaemia. We observed the selective requirement for these proteins for leukaemic cell expansion and self-renewal in-vitro as well as in leukaemia. Gene expression profiling in human cells of each of these three factors suggests that they have overlapping functions in leukaemia. The gene expression changes induced by loss of the three proteins demonstrate that they are required for the expression of haematopoietic stem cell associated genes but in contrast to previous results obtained in mouse cells, the three proteins are not required for the expression of c-MYC regulated genes.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0142806PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4646637PMC
June 2016

Allele-Specific DNA Methylation Detection by Pyrosequencing®.

Methods Mol Biol 2015 ;1315:271-89

Department of Hematology, Rigshospitalet, Blegdamsvej 9, 2100-DK, Copenhagen, Denmark,

DNA methylation is an epigenetic modification that plays important roles in healthy as well as diseased cells, by influencing the transcription of genes. In spite the fact that human somatic cells are diploid, most of the currently available methods for the study of DNA methylation do not provide information on the methylation status of individual alleles of genes. This information may be of importance in many situations. In particular, in cancer both alleles of tumour suppressor genes generally need to be inactivated for a phenotypic effect to be observed. Here, we present a simple and cost-effective protocol for allele-specific DNA methylation detection based on Pyrosequencing(®) of methylation-specific PCR (MSP) products including a single nucleotide polymorphism (SNP) within the amplicon.
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http://dx.doi.org/10.1007/978-1-4939-2715-9_20DOI Listing
March 2016

Cyclin F suppresses B-Myb activity to promote cell cycle checkpoint control.

Nat Commun 2015 Jan 5;6:5800. Epub 2015 Jan 5.

Biotech Research and Innovation Centre (BRIC), University of Copenhagen, Ole Maaløes Vej 5, 2200 Copenhagen N, Denmark.

Cells respond to DNA damage by activating cell cycle checkpoints to delay proliferation and facilitate DNA repair. Here, to uncover new checkpoint regulators, we perform RNA interference screening targeting genes involved in ubiquitylation processes. We show that the F-box protein cyclin F plays an important role in checkpoint control following ionizing radiation. Cyclin F-depleted cells initiate checkpoint signalling after ionizing radiation, but fail to maintain G2 phase arrest and progress into mitosis prematurely. Importantly, cyclin F suppresses the B-Myb-driven transcriptional programme that promotes accumulation of crucial mitosis-promoting proteins. Cyclin F interacts with B-Myb via the cyclin box domain. This interaction is important to suppress cyclin A-mediated phosphorylation of B-Myb, a key step in B-Myb activation. In summary, we uncover a regulatory mechanism linking the F-box protein cyclin F with suppression of the B-Myb/cyclin A pathway to ensure a DNA damage-induced checkpoint response in G2.
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http://dx.doi.org/10.1038/ncomms6800DOI Listing
January 2015

Loss of PRDM11 promotes MYC-driven lymphomagenesis.

Blood 2015 Feb 12;125(8):1272-81. Epub 2014 Dec 12.

Biotech Research and Innovation Centre and Centre for Epigenetics, University of Copenhagen, Copenhagen, Denmark;

The PR-domain (PRDM) family of genes encodes transcriptional regulators, several of which are deregulated in cancer. By using a functional screening approach, we sought to identify novel tumor suppressors among the PRDMs. Here we demonstrate oncogenic collaboration between depletion of the previously uncharacterized PR-domain family member Prdm11 and overexpression of MYC. Overexpression of PRDM11 inhibits proliferation and induces apoptosis. Prdm11 knockout mice are viable, and loss of Prdm11 accelerates MYC-driven lymphomagenesis in the Eµ-Myc mouse model. Moreover, we show that patients with PRDM11-deficient diffuse large B-cell lymphomas (DLBCLs) have poorer overall survival and belong to the nongerminal center B-cell-like subtype. Mechanistically, genome-wide mapping of PRDM11 binding sites coupled with transcriptome sequencing in human DLBCL cells evidenced that PRDM11 associates with transcriptional start sites of target genes and regulates important oncogenes such as FOS and JUN. Hence, we characterize PRDM11 as a putative novel tumor suppressor that controls the expression of key oncogenes, and we add new mechanistic insight into B-cell lymphomagenesis.
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http://dx.doi.org/10.1182/blood-2014-03-560805DOI Listing
February 2015

Gene silencing triggers polycomb repressive complex 2 recruitment to CpG islands genome wide.

Mol Cell 2014 Aug 3;55(3):347-60. Epub 2014 Jul 3.

Biotech Research and Innovation Centre (BRIC), University of Copenhagen, Ole Maaløes Vej 5, 2200 Copenhagen, Denmark; Centre for Epigenetics, University of Copenhagen, Ole Maaløes Vej 5, 2200 Copenhagen, Denmark; The Danish Stem Cell Center (DanStem), Blegdamsvej 3, 2200 Copenhagen, Denmark. Electronic address:

Polycomb group (PcG) proteins are required for normal differentiation and development and are frequently deregulated in cancer. PcG proteins are involved in gene silencing; however, their role in initiation and maintenance of transcriptional repression is not well defined. Here, we show that knockout of the Polycomb repressive complex 2 (PRC2) does not lead to significant gene expression changes in mouse embryonic stem cells (mESCs) and that it is dispensable for initiating silencing of target genes during differentiation. Transcriptional inhibition in mESCs is sufficient to induce genome-wide ectopic PRC2 recruitment to endogenous PcG target genes found in other tissues. PRC2 binding analysis shows that it is restricted to nucleosome-free CpG islands (CGIs) of untranscribed genes. Our results show that it is the transcriptional state that governs PRC2 binding, and we propose that it binds by default to nontranscribed CGI genes to maintain their silenced state and to protect cell identity.
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http://dx.doi.org/10.1016/j.molcel.2014.06.005DOI Listing
August 2014

The histone lysine demethylase JMJD3/KDM6B is recruited to p53 bound promoters and enhancer elements in a p53 dependent manner.

PLoS One 2014 5;9(5):e96545. Epub 2014 May 5.

Biotech Research and Innovation Centre (BRIC), University of Copenhagen, Copenhagen, Denmark; Centre for Epigenetics, University of Copenhagen, Copenhagen, Denmark; The Danish Stem Cell Center (DanStem), University of Copenhagen, Copenhagen, Denmark.

The JmjC domain-containing protein JMJD3/KDM6B catalyses the demethylation of H3K27me3 and H3K27me2. JMJD3 appears to be highly regulated at the transcriptional level and is upregulated in response to diverse stimuli such as differentiation inducers and stress signals. Accordingly, JMJD3 has been linked to the regulation of different biological processes such as differentiation of embryonic stem cells, inflammatory responses in macrophages, and induction of cellular senescence via regulation of the INK4A-ARF locus. Here we show here that JMJD3 interacts with the tumour suppressor protein p53. We find that the interaction is dependent on the p53 tetramerization domain. Following DNA damage, JMJD3 is transcriptionally upregulated and by performing genome-wide mapping of JMJD3, we demonstrate that it binds genes involved in basic cellular processes, as well as genes regulating cell cycle, response to stress and apoptosis. Moreover, we find that JMJD3 binding sites show significant overlap with p53 bound promoters and enhancer elements. The binding of JMJD3 to p53 target sites is increased in response to DNA damage, and we demonstrate that the recruitment of JMJD3 to these sites is dependent on p53 expression. Therefore, we propose a model in which JMJD3 is recruited to p53 responsive elements via its interaction with p53 and speculate that JMJD3 could act as a fail-safe mechanism to remove low levels of H3K27me3 and H3K27me2 to allow for efficient acetylation of H3K27.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0096545PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4010471PMC
July 2015

A screen identifies the oncogenic micro-RNA miR-378a-5p as a negative regulator of oncogene-induced senescence.

PLoS One 2014 20;9(3):e91034. Epub 2014 Mar 20.

Biotech Research and Innovation Centre, University of Copenhagen, Copenhagen, Denmark; Centre for Epigenetics, University of Copenhagen, Copenhagen, Denmark; The Danish Stem Cell Center (DanStem), University of Copenhagen, Copenhagen, Denmark.

Oncogene-induced senescence (OIS) can occur in response to hyperactive oncogenic signals and is believed to be a fail-safe mechanism protecting against tumorigenesis. To identify new factors involved in OIS, we performed a screen for microRNAs that can overcome or inhibit OIS in human diploid fibroblasts. This screen led to the identification of miR-378a-5p and in addition several other miRNAs that have previously been shown to play a role in senescence. We show that ectopic expression of miR-378a-5p reduces the expression of several senescence markers, including p16(INK4A) and senescence-associated β-galactosidase. Moreover, cells with ectopic expression of miR-378a-5p retain proliferative capacity even in the presence of an activated Braf oncogene. Finally, we identified several miR-378a-5p targets in diploid fibroblasts that might explain the mechanism by which the microRNA can delay OIS. We speculate that miR-378a-5p might positively influence tumor formation by delaying OIS, which is consistent with a known pro-oncogenic function of this microRNA.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0091034PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3961217PMC
December 2014

Fbxl10/Kdm2b recruits polycomb repressive complex 1 to CpG islands and regulates H2A ubiquitylation.

Mol Cell 2013 Mar 7;49(6):1134-46. Epub 2013 Feb 7.

Biotech Research and Innovation Centre, University of Copenhagen, Ole Maaløes Vej 5, 2200 Copenhagen, Denmark.

Polycomb repressive complex 1 (PRC1) catalyzes lysine 119 monoubiquitylation on H2A (H2AK119ub1) and regulates pluripotency in embryonic stem cells (ESCs). However, the mechanisms controlling the binding of PRC1 to genomic sites and its catalytic activity are poorly understood. Here, we show that Fbxl10 interacts with Ring1B and Nspc1, forming a noncanonical PRC1 that is required for H2AK119ub1 in mouse ESCs. Genome-wide analyses reveal that Fbxl10 preferentially binds to CpG islands and colocalizes with Ring1B on Polycomb target genes. Notably, Fbxl10 depletion causes a decrease in Ring1B binding to target genes and a major loss of H2AK119ub1. Furthermore, genetic analyses demonstrate that Fbxl10 DNA binding capability and integration into PRC1 are required for H2AK119 ubiquitylation. ESCs lacking Fbxl10, like previously characterized Polycomb mutants, cannot differentiate properly. These results demonstrate that Fbxl10 has a key role in regulating Ring1B recruitment to its target genes and H2AK119 ubiquitylation in ESCs.
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http://dx.doi.org/10.1016/j.molcel.2013.01.016DOI Listing
March 2013

A genetic screen identifies BRCA2 and PALB2 as key regulators of G2 checkpoint maintenance.

EMBO Rep 2011 Jul 1;12(7):705-12. Epub 2011 Jul 1.

Biotech Research and Innovation Centre, University of Copenhagen, Ole Maaløes Vej 5, Copenhagen N 2200, Denmark.

To identify key connections between DNA-damage repair and checkpoint pathways, we performed RNA interference screens for regulators of the ionizing radiation-induced G2 checkpoint, and we identified the breast cancer gene BRCA2. The checkpoint was also abrogated following depletion of PALB2, an interaction partner of BRCA2. BRCA2 and PALB2 depletion led to premature checkpoint abrogation and earlier activation of the AURORA A-PLK1 checkpoint-recovery pathway. These results indicate that the breast cancer tumour suppressors and homologous recombination repair proteins BRCA2 and PALB2 are main regulators of G2 checkpoint maintenance following DNA-damage.
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http://dx.doi.org/10.1038/embor.2011.99DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3128973PMC
July 2011

MMSET is highly expressed and associated with aggressiveness in neuroblastoma.

Cancer Res 2011 Jun 28;71(12):4226-35. Epub 2011 Apr 28.

Biotech Research and Innovation Centre, University of Copenhagen, Denmark.

MMSET (WHSC1/NSD2) is a SET domain-containing histone lysine methyltransferase the expression of which is deregulated in a subgroup of multiple myelomas with the t(4;14)(p16;q32) translocation associated with poor prognosis. Recent studies have shown that MMSET mRNA levels are increased in other tumor types as well. We have carried out immunohistochemical staining of tissue microarrays and found that MMSET protein is frequently and highly expressed in neuroblastoma (MMSET positive in 75% of neuroblastomas, n = 164). The expression level of MMSET in neuroblastomas was significantly associated with poor survival, negative prognostic factors, and metastatic disease. Moreover, a subset of neuroblastomas for which pre- and postchemotherapy biopsies were available displayed a strong decrease in MMSET protein levels after chemotherapy. In agreement with neuroblastomas becoming more differentiated after treatment, we show that retinoic acid-induced differentiation of human neuroblastoma cells in vitro also leads to a strong decrease in MMSET levels. Furthermore, we show that the high levels of MMSET in normal neural progenitor cells are strongly downregulated during differentiation. Importantly, we show that MMSET is required for proliferation of neuroblastoma cells and brain-derived neural stem cells. Taken together, our results suggest that MMSET is implicated in neuroblastomagenesis possibly by supporting proliferation of progenitor cells and negatively regulating their differentiation. In this respect, MMSET might be a strong candidate therapeutic target in a subset of neuroblastomas with unfavorable prognosis.
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http://dx.doi.org/10.1158/0008-5472.CAN-10-3810DOI Listing
June 2011

The histone methyltransferase and putative oncoprotein MMSET is overexpressed in a large variety of human tumors.

Clin Cancer Res 2011 May 8;17(9):2919-33. Epub 2011 Mar 8.

Biotech Research and Innovation Centre, Rigshospitalet, University of Copenhagen, Denmark.

Purpose: Multiple myeloma SET (Suppressor of variegation, Enhancer of zeste, and Trithorax) domain (MMSET) is a histone lysine methyltransferase deregulated in a subgroup of multiple myelomas with the t(4;14)(p16;q32) translocation and poor prognosis. With the aim of understanding, if MMSET can be involved in other types of cancer we investigated the expression of MMSET protein in different types of human tumors.

Experimental Design: A monoclonal antibody against MMSET was developed and immunohistochemical staining of tissue microarrays (TMA) containing a large number of tumor samples (n = 3774) and corresponding normal tissues (n = 904) was carried out. Further validations of MMSET expression were carried out on independent, tumor-specific sets of TMAs for urinary bladder (n = 1293) and colon cancer (n = 1206) with corresponding clinicopathological data and long-term follow-up.

Results: MMSET protein was highly expressed in different tumor types compared to normal counterparts. Particular frequent and/or high MMSET expression was found in carcinomas of the gastrointestinal tract (stomach, colon, anal canal), small cell lung carcinoma, tumors of the urinary bladder, female genitals, and skin. In bladder cancer, MMSET expression correlated with tumor aggressiveness. In contrast, MMSET expression was associated with good prognostic factors in colon cancer and was more pronounced in early stages of colon carcinogenesis (dysplasias) than in adenocarcinomas. However, colon cancer patients with high MMSET levels showed a worse 5-year survival.

Conclusions: Our data suggest that MMSET has a broader role in cancer than previously anticipated, and further analysis might qualify it as a prognostic marker and a target for the development of therapy against several types of cancer.
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http://dx.doi.org/10.1158/1078-0432.CCR-10-1302DOI Listing
May 2011

The histone demethylases JMJD1A and JMJD2B are transcriptional targets of hypoxia-inducible factor HIF.

J Biol Chem 2008 Dec 4;283(52):36542-52. Epub 2008 Nov 4.

Biotech Research and Innovation Centre, University of Copenhagen, Copenhagen, Denmark.

Posttranslational histone modifications serve to store epigenetic information and control both nucleosome assembly and recruitment of non-histone proteins. Histone methylation occurs on arginine and lysine residues and is involved in the regulation of gene transcription. A dynamic control of these modifications is exerted by histone methyltransferases and the recently discovered histone demethylases. Here we show that the hypoxia-inducible factor HIF-1alpha binds to specific recognition sites in the genes encoding the jumonji family histone demethylases JMJD1A and JMJD2B and induces their expression. Accordingly, hypoxic cells express elevated levels of JMJD1A and JMJD2B mRNA and protein. Furthermore, we find increased expression of JMJD1A and JMJD2B in renal cancer cells that have lost the von Hippel Lindau tumor suppressor protein VHL and therefore display a deregulated expression of hypoxia-inducible factor. Studies on ectopically expressed JMJD1A and JMJD2B indicate that both proteins retain their histone lysine demethylase activity in hypoxia and thereby might impact the hypoxic gene expression program.
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http://dx.doi.org/10.1074/jbc.M804578200DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2662309PMC
December 2008
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