Publications by authors named "Anna Elisabetta Salcini"

23 Publications

  • Page 1 of 1

Coordinated maintenance of H3K36/K27 methylation by histone demethylases preserves germ cell identity and immortality.

Cell Rep 2021 Nov;37(8):110050

Biotech Research and Innovation Centre (BRIC), University of Copenhagen, Ole Maaløes vej 5, Copenhagen DK-2200, Denmark. Electronic address:

Germ cells have evolved unique mechanisms to ensure the transmission of genetically and nongenetically encoded information, whose alteration compromises germ cell immortality. Chromatin factors play fundamental roles in these mechanisms. H3K36 and H3K27 methyltransferases shape and propagate a pattern of histone methylation essential for C. elegans germ cell maintenance, but the role of respective histone demethylases remains unexplored. Here, we show that jmjd-5 regulates H3K36me2 and H3K27me3 levels, preserves germline immortality, and protects germ cell identity by controlling gene expression. The transcriptional and biological effects of jmjd-5 loss can be hindered by the removal of H3K27demethylases, indicating that H3K36/K27 demethylases act in a transcriptional framework and promote the balance between H3K36 and H3K27 methylation required for germ cell immortality. Furthermore, we find that in wild-type, but not in jmjd-5 mutants, alterations of H3K36 methylation and transcription occur at high temperature, suggesting a role for jmjd-5 in adaptation to environmental changes.
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http://dx.doi.org/10.1016/j.celrep.2021.110050DOI Listing
November 2021

H3K27 modifiers regulate lifespan in C. elegans in a context-dependent manner.

BMC Biol 2021 03 25;19(1):59. Epub 2021 Mar 25.

Department of Biochemistry, University of Oxford, Oxford, UK.

Background: Evidence of global heterochromatin decay and aberrant gene expression in models of physiological and premature ageing have long supported the "heterochromatin loss theory of ageing", which proposes that ageing is aetiologically linked to, and accompanied by, a progressive, generalised loss of repressive epigenetic signatures. However, the remarkable plasticity of chromatin conformation suggests that the re-establishment of such marks could potentially revert the transcriptomic architecture of animal cells to a "younger" state, promoting longevity and healthspan. To expand our understanding of the ageing process and its connection to chromatin biology, we screened an RNAi library of chromatin-associated factors for increased longevity phenotypes.

Results: We identified the lysine demethylases jmjd-3.2 and utx-1, as well as the lysine methyltransferase mes-2 as regulators of both lifespan and healthspan in C. elegans. Strikingly, we found that both overexpression and loss of function of jmjd-3.2 and utx-1 are all associated with enhanced longevity. Furthermore, we showed that the catalytic activity of UTX-1, but not JMJD-3.2, is critical for lifespan extension in the context of overexpression. In attempting to reconcile the improved longevity associated with both loss and gain of function of utx-1, we investigated the alternative lifespan pathways and tissue specificity of longevity outcomes. We demonstrated that lifespan extension caused by loss of utx-1 function is daf-16 dependent, while overexpression effects are partially independent of daf-16. In addition, lifespan extension was observed when utx-1 was knocked down or overexpressed in neurons and intestine, whereas in the epidermis, only knockdown of utx-1 conferred improved longevity.

Conclusions: We show that the regulation of longevity by chromatin modifiers can be the result of the interaction between distinct factors, such as the level and tissue of expression. Overall, we suggest that the heterochromatin loss model of ageing may be too simplistic an explanation of organismal ageing when molecular and tissue-specific effects are taken into account.
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http://dx.doi.org/10.1186/s12915-021-00984-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7995591PMC
March 2021

Regulators of H3K4 methylation mutated in neurodevelopmental disorders control axon guidance in .

Development 2020 08 7;147(15). Epub 2020 Aug 7.

BRIC, University of Copenhagen, Biotech Research and Innovation Centre, Ole Maaloes vej 5, 2200, Copenhagen, Denmark

Post-translational histone modifications regulate chromatin compaction and gene expression to control many aspects of development. Mutations in genes encoding regulators of H3K4 methylation are causally associated with neurodevelopmental disorders characterized by intellectual disability and deficits in motor functions. However, it remains unclear how H3K4 methylation influences nervous system development and contributes to the aetiology of disease. Here, we show that the catalytic activity of , the homologue of the H3K4 methyltransferase KMT2F/G (SETD1A/B) genes, controls embryonic transcription of neuronal genes and is required for establishing proper axon guidance, and for neuronal functions related to locomotion and learning. Moreover, we uncover a striking correlation between components of the H3K4 regulatory machinery mutated in neurodevelopmental disorders and the process of axon guidance in Thus, our study supports an epigenetic-based model for the aetiology of neurodevelopmental disorders, based on an aberrant axon guidance process originating from deregulated H3K4 methylation.
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http://dx.doi.org/10.1242/dev.190637DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7420840PMC
August 2020

JMJD-1.2 controls multiple histone post-translational modifications in germ cells and protects the genome from replication stress.

Sci Rep 2018 02 28;8(1):3765. Epub 2018 Feb 28.

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

Post-translational modifications of histones, constitutive components of chromatin, regulate chromatin compaction and control all DNA-based cellular processes. C. elegans JMJD-1.2, a member of the KDM7 family, is a demethylase active towards several lysine residues on Histone 3 (H3), but its contribution in regulating histone methylation in germ cells has not been fully investigated. Here, we show that jmjd-1.2 is expressed abundantly in the germline where it controls the level of histone 3 lysine 9, lysine 23 and lysine 27 di-methylation (H3K9/K23/K27me2) both in mitotic and meiotic cells. Loss of jmjd-1.2 is not associated with major defects in the germ cells in animals grown under normal conditions or after DNA damage induced by UV or ionizing irradiation. However, jmjd-1.2 mutants are more sensitive to replication stress and the progeny of mutant animals exposed to hydroxyurea show increased embryonic lethality and mutational rate, compared to wild-type. Thus, our results suggest a role for jmjd-1.2 in the maintenance of genome integrity after replication stress and emphasize the relevance of the regulation of histone methylation in genomic stability.
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http://dx.doi.org/10.1038/s41598-018-21914-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5830613PMC
February 2018

Proteomic Characterization of Caenorhabditis elegans Larval Development.

Proteomics 2018 01 27;18(2). Epub 2017 Dec 27.

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

The nematode Caenorhabditis elegans is widely used as a model organism to study cell and developmental biology. Quantitative proteomics of C. elegans is still in its infancy and, so far, most studies have been performed on adult worm samples. Here, we used quantitative mass spectrometry to characterize protein level changes across the four larval developmental stages (L1-L4) of C. elegans. In total, we identified 4130 proteins, and quantified 1541 proteins that were present across all four stages in three biological replicates from independent experiments. Using hierarchical clustering and functional ontological analyses, we identified 21 clusters containing proteins with similar protein profiles across the four stages, and highlighted the most overrepresented biological functions in each of these protein clusters. In addition, we used the dataset to identify putative larval stage-specific proteins in each individual developmental stage, as well as in the early and late developmental stages. In summary, this dataset provides system-wide analysis of protein level changes across the four C. elegans larval developmental stages, which serves as a useful resource for the C. elegans research community. MS data were deposited in ProteomeXchange (http://proteomecentral.proteomexchange.org) via the PRIDE partner repository with the primary accession identifier PXD006676.
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http://dx.doi.org/10.1002/pmic.201700238DOI Listing
January 2018

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

JMJD-1.2/PHF8 controls axon guidance by regulating Hedgehog-like signaling.

Development 2017 03 26;144(5):856-865. Epub 2017 Jan 26.

Biotech Research & Innovation Centre (BRIC), University of Copenhagen, 2200, Copenhagen, Denmark

Components of the KDM7 family of histone demethylases are implicated in neuronal development and one member, PHF8, is often found to be mutated in cases of X-linked mental retardation. However, how PHF8 regulates neurodevelopmental processes and contributes to the disease is still largely unknown. Here, we show that the catalytic activity of a PHF8 homolog in , JMJD-1.2, is required non-cell-autonomously for proper axon guidance. Loss of JMJD-1.2 dysregulates transcription of the Hedgehog-related genes and , the overexpression of which is sufficient to induce the axonal defects. Deficiency of either or , or reduced expression of homologs of genes promoting Hedgehog signaling, restores correct axon guidance in mutants. Genetic and overexpression data indicate that Hedgehog-related genes act on axon guidance through actin remodelers. Thus, our study highlights a novel function of in axon guidance that might be relevant for the onset of X-linked mental retardation and provides compelling evidence of a conserved function of the Hedgehog pathway in axon migration.
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http://dx.doi.org/10.1242/dev.142695DOI Listing
March 2017

Impaired removal of H3K4 methylation affects cell fate determination and gene transcription.

Development 2016 10 30;143(20):3751-3762. Epub 2016 Aug 30.

Biotech Research and Innovation Centre (BRIC), University of Copenhagen, Copenhagen DK-2200, Denmark

Methylation of histone 3 lysine 4 (H3K4) is largely associated with promoters and enhancers of actively transcribed genes and is finely regulated during development by the action of histone methyltransferases and demethylases. H3K4me3 demethylases of the KDM5 family have been previously implicated in development, but how the regulation of H3K4me3 level controls developmental processes is not fully established. Here, we show that the H3K4 demethylase RBR-2, the unique member of the KDM5 family in C. elegans, acts cell-autonomously and in a catalytic-dependent manner to control vulva precursor cells fate acquisition, by promoting the LIN-12/Notch pathway. Using genome-wide approaches, we show that RBR-2 reduces the H3K4me3 level at transcription start sites (TSSs) and in regions upstream of the TSSs, and acts both as a transcription repressor and activator. Analysis of the lin-11 genetic locus, a direct RBR-2 target gene required for vulva precursor cell fate acquisition, shows that RBR-2 controls the epigenetic signature of the lin-11 vulva-specific enhancer and lin-11 expression, providing in vivo evidence that RBR-2 can positively regulate transcription and cell fate acquisition by controlling enhancer activity.
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http://dx.doi.org/10.1242/dev.139139DOI Listing
October 2016

Dangerous R loops form in the absence of H3K9 methylation.

Nat Genet 2016 Oct;48(11):1299-1300

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

Methylation of histone H3 on lysine 9 (H3K9) is a hallmark of transcriptionally inactive heterochromatin that is deregulated in pathological conditions. A new study shows that complete loss of H3K9 methylation in Caenorhabditis elegans leads to derepression of repetitive elements and formation of DNA:RNA hybrids (R loops), resulting in increased rates of repeat-specific mutation.
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http://dx.doi.org/10.1038/ng.3705DOI Listing
October 2016

The H3K4me3/2 histone demethylase RBR-2 controls axon guidance by repressing the actin-remodeling gene wsp-1.

Development 2016 Mar 25;143(5):851-63. Epub 2016 Jan 25.

Biotech Research & Innovation Centre (BRIC), University of Copenhagen, 2200 Copenhagen, Denmark Centre for Epigenetics, University of Copenhagen, 2200 Copenhagen, Denmark

The dynamic regulation of histone modifications is important for modulating transcriptional programs during development. Aberrant H3K4 methylation is associated with neurological disorders, but how the levels and the recognition of this modification affect specific neuronal processes is unclear. Here, we show that RBR-2, the sole homolog of the KDM5 family of H3K4me3/2 demethylases in Caenorhabditis elegans, ensures correct axon guidance by controlling the expression of the actin regulator wsp-1. Loss of rbr-2 results in increased levels of H3K4me3 at the transcriptional start site of wsp-1, with concomitant higher wsp-1 expression responsible for defective axon guidance. In agreement, overexpression of WSP-1 mimics rbr-2 loss, and its depletion restores normal axon guidance in rbr-2 mutants. NURF-1, an H3K4me3-binding protein and member of the chromatin-remodeling complex NURF, is required for promoting aberrant wsp-1 transcription in rbr-2 mutants and its ablation restores wild-type expression of wsp-1 and axon guidance. Thus, our results establish a precise role for epigenetic regulation in neuronal development by demonstrating a functional link between RBR-2 activity, H3K4me3 levels, the NURF complex and the expression of WSP-1.
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http://dx.doi.org/10.1242/dev.132985DOI Listing
March 2016

Dynamic changes of histone H3 marks during Caenorhabditis elegans lifecycle revealed by middle-down proteomics.

Proteomics 2016 Feb 15;16(3):459-64. Epub 2016 Jan 15.

Centre for Epigenetics, Department of Biochemistry and Molecular Biology, University of Southern Denmark, Odense M, Denmark.

We applied a middle-down proteomics strategy for large-scale protein analysis during in vivo development of Caenorhabditis elegans. We characterized PTMs on histone H3 N-terminal tails at eight time points during the C. elegans lifecycle, including embryo, larval stages (L1-L4), dauer, and L1/L4 postdauer. Histones were analyzed by our optimized middle-down protein sequencing platform using high mass accuracy MS/MS. This allows quantification of intact histone tails and detailed characterization of distinct histone tails carrying cooccurring PTMs. We measured temporally distinct combinatorial PTM profiles during C. elegans development. We show that the doubly modified form H3K23me3K27me3, which is rare or nonexistent in mammals, is the most abundant PTM in all stages of C. elegans lifecycle. The abundance of H3K23me3 increased during development and it was mutually exclusive of the active marks H3K18ac, R26me1, and R40me1, suggesting a role for H3K23me3 in silent chromatin. We observed distinct PTM profiles for normal L1 larvae and for L1-postdauer larvae, or L4 and L4 postdauer, suggesting that histone PTMs mediate an epigenetic memory that is transmitted during dauer formation. Collectively, our data describe the dynamics of histone H3 combinatorial code during C. elegans lifecycle and demonstrate the feasibility of using middle-down proteomics to study in vivo development of multicellular organisms. All MS data have been deposited in the ProteomeXchange with identifier PXD002525 (http://proteomecentral.proteomexchange.org/dataset/PXD002525).
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http://dx.doi.org/10.1002/pmic.201500285DOI Listing
February 2016

H3K23me2 is a new heterochromatic mark in Caenorhabditis elegans.

Nucleic Acids Res 2015 Nov 17;43(20):9694-710. Epub 2015 Oct 17.

Biotech Research & Innovation Centre (BRIC), University of Copenhagen, Ole Maaløes Vej 5, 2200 Copenhagen N, Denmark Centre for Epigenetics, University of Copenhagen, Ole Maaløes Vej 5, 2200 Copenhagen N, Denmark

Genome-wide analyses in Caenorhabditis elegans show that post-translational modifications (PTMs) of histones are evolutionary conserved and distributed along functionally distinct genomic domains. However, a global profile of PTMs and their co-occurrence on the same histone tail has not been described in this organism. We used mass spectrometry based middle-down proteomics to analyze histone H3 N-terminal tails from C. elegans embryos for the presence, the relative abundance and the potential cross-talk of co-existing PTMs. This analysis highlighted that the lysine 23 of histone H3 (H3K23) is extensively modified by methylation and that tri-methylated H3K9 (H3K9me3) is exclusively detected on histone tails with di-methylated H3K23 (H3K23me2). Chromatin immunoprecipitation approaches revealed a positive correlation between H3K23me2 and repressive marks. By immunofluorescence analyses, H3K23me2 appears differentially regulated in germ and somatic cells, in part by the action of the histone demethylase JMJD-1.2. H3K23me2 is enriched in heterochromatic regions, localizing in H3K9me3 and heterochromatin protein like-1 (HPL-1)-positive foci. Biochemical analyses indicated that HPL-1 binds to H3K23me2 and interacts with a conserved CoREST repressive complex. Thus, our study suggests that H3K23me2 defines repressive domains and contributes to organizing the genome in distinct heterochromatic regions during embryogenesis.
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http://dx.doi.org/10.1093/nar/gkv1063DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4787770PMC
November 2015

EZH2 protects glioma stem cells from radiation-induced cell death in a MELK/FOXM1-dependent manner.

Stem Cell Reports 2015 Feb 15;4(2):226-38. Epub 2015 Jan 15.

Department of Neurological Surgery, The James Comprehensive Cancer Center, The Ohio State University, Columbus, OH 43210, USA. Electronic address:

Glioblastoma (GBM)-derived tumorigenic stem-like cells (GSCs) may play a key role in therapy resistance. Previously, we reported that the mitotic kinase MELK binds and phosphorylates the oncogenic transcription factor FOXM1 in GSCs. Here, we demonstrate that the catalytic subunit of Polycomb repressive complex 2, EZH2, is targeted by the MELK-FOXM1 complex, which in turn promotes resistance to radiation in GSCs. Clinically, EZH2 and MELK are coexpressed in GBM and significantly induced in postirradiation recurrent tumors whose expression is inversely correlated with patient prognosis. Through a gain-and loss-of-function study, we show that MELK or FOXM1 contributes to GSC radioresistance by regulation of EZH2. We further demonstrate that the MELK-EZH2 axis is evolutionarily conserved in Caenorhabditis elegans. Collectively, these data suggest that the MELK-FOXM1-EZH2 signaling axis is essential for GSC radioresistance and therefore raise the possibility that MELK-FOXM1-driven EZH2 signaling can serve as a therapeutic target in irradiation-resistant GBM tumors.
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http://dx.doi.org/10.1016/j.stemcr.2014.12.006DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4325196PMC
February 2015

Catalytic-independent roles of UTX-1 in C. elegans development.

Worm 2013 Apr;2(2):e22188

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

We recently analyzed the functional roles of UTX-1 during development. utx-1 is an essential gene required for the correct embryonic and post-embryonic development of C. elegans, and it displays an H3K27me3 demethylase activity. Rescue experiments demonstrated that the enzymatic activity of UTX-1 is not relevant for its role in development. The phenotypes associated with loss of UTX-1 might, instead, be a result of compromised functions of an UTX-1-containing complex. Here we discuss the possible mechanisms by which UTX-1 contributes to normal development.
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http://dx.doi.org/10.4161/worm.22188DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3704440PMC
April 2013

A snapshot of the physical and functional wiring of the Eps15 homology domain network in the nematode.

PLoS One 2013 12;8(2):e56383. Epub 2013 Feb 12.

IFOM, Fondazione Istituto FIRC di Oncologia Molecolare, Milan, Italy.

Protein interaction modules coordinate the connections within and the activity of intracellular signaling networks. The Eps15 Homology (EH) module, a protein-protein interaction domain that is a key feature of the EH-network, was originally identified in a few proteins involved in endocytosis and vesicle trafficking, and has subsequently also been implicated in actin reorganization, nuclear shuttling, and DNA repair. Here we report an extensive characterization of the physical connections and of the functional wirings of the EH-network in the nematode. Our data show that one of the major physiological roles of the EH-network is in neurotransmission. In addition, we found that the proteins of the network intersect, and possibly coordinate, a number of "territories" of cellular activity including endocytosis/recycling/vesicle transport, actin dynamics, general metabolism and signal transduction, ubiquitination/degradation of proteins, DNA replication/repair, and miRNA biogenesis and processing.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0056383PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3570524PMC
August 2013

The C. elegans H3K27 demethylase UTX-1 is essential for normal development, independent of its enzymatic activity.

PLoS Genet 2012 3;8(5):e1002647. Epub 2012 May 3.

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

Epigenetic modifications influence gene expression and provide a unique mechanism for fine-tuning cellular differentiation and development in multicellular organisms. Here we report on the biological functions of UTX-1, the Caenorhabditis elegans homologue of mammalian UTX, a histone demethylase specific for H3K27me2/3. We demonstrate that utx-1 is an essential gene that is required for correct embryonic and postembryonic development. Consistent with its homology to UTX, UTX-1 regulates global levels of H3K27me2/3 in C. elegans. Surprisingly, we found that the catalytic activity is not required for the developmental function of this protein. Biochemical analysis identified UTX-1 as a component of a complex that includes SET-16(MLL), and genetic analysis indicates that the defects associated with loss of UTX-1 are likely mediated by compromised SET-16/UTX-1 complex activity. Taken together, these results demonstrate that UTX-1 is required for many aspects of nematode development; but, unexpectedly, this function is independent of its enzymatic activity.
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http://dx.doi.org/10.1371/journal.pgen.1002647DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3342935PMC
September 2012

PI3KC2α, a class II PI3K, is required for dynamin-independent internalization pathways.

J Cell Sci 2010 Dec 16;123(Pt 24):4240-50. Epub 2010 Nov 16.

Biotech Research and Innovation Centre, BRIC, University of Copenhagen, Ole Maaløes Vej 5, DK2200 Copenhagen, Denmark.

Increasing evidence indicates that cellular uptake of several molecules can occur independently of functional dynamin, but the molecular players that regulate dynamin-independent endocytosis and the subsequent trafficking steps are still largely unknown. A survival-based short-hairpin (sh) RNA screen using a cell line expressing a diphtheria toxin receptor (DTR, officially known as HBEGF) anchored to GPI (DTR-GPI), which internalizes diphtheria toxin (DT, officially known as DTX) in a dynamin-independent manner, identified PI3KC2α, a class II phosphoinositide 3-kinase (PI3K), as a specific regulator of dynamin-independent DT internalization. We found that the internalization of several proteins that enter the cell through dynamin-independent pathways led to a relocalization of PI3KC2α to cargo-positive vesicles. Furthermore, downregulation of PI3KC2α impaired internalization of CD59 as well as fluid-phase endocytosis. Our data suggest a general role for PI3KC2α in regulating physiologically relevant dynamin-independent internalization pathways by recruiting early endosome antigen 1 (EEA1) to vesicular compartments, a step required for the intracellular trafficking of vesicles generated by dynamin-independent endocytic pathways.
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http://dx.doi.org/10.1242/jcs.071712DOI Listing
December 2010

A functional link between the histone demethylase PHF8 and the transcription factor ZNF711 in X-linked mental retardation.

Mol Cell 2010 Apr 25;38(2):165-78. Epub 2010 Mar 25.

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

X-linked mental retardation (XLMR) is an inherited disorder that mostly affects males and is caused by mutations in genes located on the X chromosome. Here, we show that the XLMR protein PHF8 and a C. elegans homolog F29B9.2 catalyze demethylation of di- and monomethylated lysine 9 of histone H3 (H3K9me2/me1). The PHD domain of PHF8 binds to H3K4me3 and colocalizes with H3K4me3 at transcription initiation sites. Furthermore, PHF8 interacts with another XMLR protein, ZNF711, which binds to a subset of PHF8 target genes, including the XLMR gene JARID1C. Of interest, the C. elegans PHF8 homolog is highly expressed in neurons, and mutant animals show impaired locomotion. Taken together, our results functionally link the XLMR gene PHF8 to two other XLMR genes, ZNF711 and JARID1C, indicating that MR genes may be functionally linked in pathways, causing the complex phenotypes observed in patients developing MR.
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http://dx.doi.org/10.1016/j.molcel.2010.03.002DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2989439PMC
April 2010

Caenorhabditis elegans intersectin: a synaptic protein regulating neurotransmission.

Mol Biol Cell 2007 Dec 17;18(12):5091-9. Epub 2007 Oct 17.

Biotech Research and Innovation Centre, DK-2200 Copenhagen, Denmark.

Intersectin is a multifunctional protein that interacts with components of the endocytic and exocytic pathways, and it is also involved in the control of actin dynamics. Drosophila intersectin is required for viability, synaptic development, and synaptic vesicle recycling. Here, we report the characterization of intersectin function in Caenorhabditis elegans. Nematode intersectin (ITSN-1) is expressed in the nervous system, and it is enriched in presynaptic regions. The C. elegans intersectin gene (itsn-1) is nonessential for viability. In addition, itsn-1-null worms do not display any evident phenotype, under physiological conditions. However, they display aldicarb-hypersensitivity, compatible with a negative regulatory role of ITSN-1 on neurotransmission. ITSN-1 physically interacts with dynamin and EHS-1, two proteins involved in synaptic vesicle recycling. We have previously shown that EHS-1 is a positive modulator of synaptic vesicle recycling in the nematode, likely through modulation of dynamin or dynamin-controlled pathways. Here, we show that ITSN-1 and EHS-1 have opposite effects on aldicarb sensitivity, and on dynamin-dependent phenotypes. Thus, the sum of our results identifies dynamin, or a dynamin-controlled pathway, as a potential target for the negative regulatory role of ITSN-1.
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http://dx.doi.org/10.1091/mbc.e07-05-0460DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2096573PMC
December 2007

UTX and JMJD3 are histone H3K27 demethylases involved in HOX gene regulation and development.

Nature 2007 Oct 22;449(7163):731-4. Epub 2007 Aug 22.

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

The trithorax and the polycomb group proteins are chromatin modifiers, which play a key role in the epigenetic regulation of development, differentiation and maintenance of cell fates. The polycomb repressive complex 2 (PRC2) mediates transcriptional repression by catalysing the di- and tri-methylation of Lys 27 on histone H3 (H3K27me2/me3). Owing to the essential role of the PRC2 complex in repressing a large number of genes involved in somatic processes, the H3K27me3 mark is associated with the unique epigenetic state of stem cells. The rapid decrease of the H3K27me3 mark during specific stages of embryogenesis and stem-cell differentiation indicates that histone demethylases specific for H3K27me3 may exist. Here we show that the human JmjC-domain-containing proteins UTX and JMJD3 demethylate tri-methylated Lys 27 on histone H3. Furthermore, we demonstrate that ectopic expression of JMJD3 leads to a strong decrease of H3K27me3 levels and causes delocalization of polycomb proteins in vivo. Consistent with the strong decrease in H3K27me3 levels associated with HOX genes during differentiation, we show that UTX directly binds to the HOXB1 locus and is required for its activation. Finally mutation of F18E9.5, a Caenorhabditis elegans JMJD3 orthologue, or inhibition of its expression, results in abnormal gonad development. Taken together, these results suggest that H3K27me3 demethylation regulated by UTX/JMJD3 proteins is essential for proper development. Moreover, the recent demonstration that UTX associates with the H3K4me3 histone methyltransferase MLL2 (ref. 8) supports a model in which the coordinated removal of repressive marks, polycomb group displacement, and deposition of activating marks are important for the stringent regulation of transcription during cellular differentiation.
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http://dx.doi.org/10.1038/nature06145DOI Listing
October 2007

RBP2 belongs to a family of demethylases, specific for tri-and dimethylated lysine 4 on histone 3.

Cell 2007 Mar 22;128(6):1063-76. Epub 2007 Feb 22.

Centre for Epigenetics, University of Copenhagen, Copenhagen, Denmark.

Methylation of histones has been regarded as a stable modification defining the epigenetic program of the cell, which regulates chromatin structure and transcription. However, the recent discovery of histone demethylases has challenged the stable nature of histone methylation. Here we demonstrate that the JARID1 proteins RBP2, PLU1, and SMCX are histone demethylases specific for di- and trimethylated histone 3 lysine 4 (H3K4). Consistent with a role for the JARID1 Drosophila homolog Lid in regulating expression of homeotic genes during development, we show that RBP2 is displaced from Hox genes during embryonic stem (ES) cell differentiation correlating with an increase of their H3K4me3 levels and expression. Furthermore, we show that mutation or RNAi depletion of the C. elegans JARID1 homolog rbr-2 leads to increased levels of H3K4me3 during larval development and defects in vulva formation. Taken together, these results suggest that H3K4me3/me2 demethylation regulated by the JARID1 family plays an important role during development.
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http://dx.doi.org/10.1016/j.cell.2007.02.003DOI Listing
March 2007

TTP specifically regulates the internalization of the transferrin receptor.

Cell 2005 Dec;123(5):875-88

IFOM, Istituto FIRC di Oncologia Molecolare, Via Adamello 16, 20139 Milan, Italy.

Different plasma membrane receptors are internalized through saturable/noncompetitive pathways, suggesting cargo-specific regulation. Here, we report that TTP (SH3BP4), a SH3-containing protein, specifically regulates the internalization of the transferrin receptor (TfR). TTP interacts with endocytic proteins, including clathrin, dynamin, and the TfR, and localizes selectively to TfR-containing coated-pits (CCP) and -vesicles (CCV). Overexpression of TTP specifically inhibits TfR internalization, and causes the formation of morphologically aberrant CCP, which are probably fission impaired. This effect is mediated by the SH3 of TTP, which can bind to dynamin, and it is rescued by overexpression of dynamin. Functional ablation of TTP causes a reduction in TfR internalization, and reduced cargo loading and size of TfR-CCV. Tyrosine phosphorylation of either TTP or dynamin prevents their interaction, pointing to a possible mechanism of exclusion of TTP from some CCP. Thus, TTP might represent one of the long sought for molecules that allow cargo-specific control of clathrin endocytosis.
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http://dx.doi.org/10.1016/j.cell.2005.10.021DOI Listing
December 2005

EH and UIM: endocytosis and more.

Sci STKE 2003 Dec 16;2003(213):re17. Epub 2003 Dec 16.

Istituto FIRC di Oncologia Molecolare, Via Adamello 16, 20139 Milan, Italy.

Exogenously and endogenously originated signals are propagated within the cell by functional and physical networks of proteins, leading to numerous biological outcomes. Many protein-protein interactions take place between binding domains and short peptide motifs. Frequently, these interactions are inducible by upstream signaling events, in which case one of the two binding surfaces may be created by a posttranslational modification. Here, we discuss two protein networks. One, the EH-network, is based on the Eps15 homology (EH) domain, which binds to peptides containing the sequence Asp-Pro-Phe (NPF). The other, which we define as the monoubiquitin (mUb) network, relies on monoubiquitination, which is emerging as an important posttranslational modification that regulates protein function. Both networks were initially implicated in the control of plasma membrane receptor endocytosis and in the regulation of intracellular trafficking routes. The ramifications of these two networks, however, appear to extend into many other aspects of cell physiology as well, such as transcriptional regulation, actin cytoskeleton remodeling, and DNA repair. The focus of this review is to integrate available knowledge of the EH- and mUb networks with predictions of genetic and physical interactions stemming from functional genomics approaches.
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http://dx.doi.org/10.1126/stke.2132003re17DOI Listing
December 2003
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