Rôle de l'interaction entre la protéine virale EBNA1 et le facteur cellulaire RCC1 dans la persistance du génome du virus d'Epstein-Barr

Epstein-Barr virus (EBV) is a ubiquitous herpesvirus associated with several human cancers. In proliferating latently-infected cells, the EBV genome persists as a circular plasmid that is replicated once per cell cycle and partitioned at mitosis. Both of these processes require a single viral protein, Epstein Barr nuclear antigen 1 (EBNA1), which binds to two clusters of cognate binding sites within the origin of plasmid replication (oriP). EBNA1 plays an essential role both in viral episome replication, by recruiting the cellular complex of DNA replication onto the oriP, and in the efficient segregation of the viral episomes, by tethering the viral DNA onto the mitotic chromosomes. Whereas the mechanisms of viral DNA replication have been well documented, the mechanisms involved in tethering EBNA1 to the cellular chromatin are far from being understood. Here we have identified Regulator of Chromosome Condensation 1 (RCC1) as a novel EBNA1 cellular partner. RCC1 is the only known nuclear guanine nucleotide exchange factor (RanGEF) for the small GTPase Ran enzyme. RCC1, associated with chromatin, is involved in the formation of RanGTP gradients critical for nucleo-cytoplasmic transport, mitotic spindle formation, and nuclear envelope reassembly after mitosis. We have used several approaches to demonstrate a direct interaction between these two proteins and to identify the regions. involved Moreover, by using Chromatin ImmunoPrecipitation assay (ChIP) we have shown that RCC1 is enriched in the oriP region of mini viral replicons in a manner dependent on EBNA1. Finally, by using a combination of confocal microscopy and FRET analysis to follow the dynamics of interaction between the two proteins throughout the cell cycle, we have demonstrated that EBNA1 and RCC1 closely associate on the chromosomes during metaphase. Taken together, our data strongly suggest an essential role for RCC1 in tethering EBNA1 - linked to the viral episome - to the metaphasic chromosomes. Our results and those of others lead us to the idea that the interaction between EBNA1 with the cellular chromosomes requires several factors such as direct interactions or cellular proteins and these interactions are complementary and / or redundant.Le virus d’Epstein-Barr (EBV) est un herpesvirus dont la séroprévalence est d’environ 90 % de la population adulte mondiale. EBV est associé à de nombreuses pathologies tumorales. La primo infection conduit à l’établissement du virus sous forme latente dans les lymphocytes B mémoires. Au sein de ces cellules B, le génome viral est sous la forme d’un épisome, un ADN circulaire double brin, et une fraction restreinte de gènes viraux est exprimée. Afin de se maintenir aux cours des divisions cellulaires, le génome viral est répliqué en phase S par la machinerie cellulaire et ségrégé lors de la mitose dans chaque cellules filles. La réplication et la ségrégation du génome viral nécessitent 2 facteurs viraux que sont la protéine virale EBNA1 (Epstein-Barr Nuclear Antigen 1) et la région oriP sur le génome viral. En phase S, EBNA1 interagit directement avec l’oriP et y recrute le complexe de pré-réplication de l’ADN. En mitose, EBNA1 ancre l’épisome à la chromatine ce qui permet une ségrégation efficace. Les mécanismes d’interaction entre EBNA1 et la chromatine reste encore flou. Au cours de notre travail, nous avons identifié la protéine RCC1 comme un partenaire potentiel pour la protéine EBNA1 pouvant être impliqué dans l’ancrage d’EBNA1 à la chromatine. Nous avons validé cette interaction et caractérisé les régions d’interactions pour ces deux protéines. Par ailleurs nous avons démontré que RCC1 est recrutée sur l’oriP en présence d’EBNA1 et que ces deux protéines interagissent en mitose. À la lumière de nos résultats et des données de la littérature, nous proposons que l’interaction d’EBNA1 avec la chromatine est dynamique et implique à la fois des interactions directes (AT-Hook, interaction avec les nucléosomes) mais aussi des facteurs cellulaires (RCC1, EBP2 et HMGB2)

Epstein-Barr virus nuclear antigen 1 interacts with regulator of chromosome condensation 1 dynamically throughout the cell cycle

The Epstein-Barr virus (EBV) nuclear antigen 1 (EBNA1) is a sequence-specific DNA binding protein which plays an essential role in viral episome replication and segregation, by recruiting the cellular complex of DNA replication onto the origin (oriP) and by tethering the viral DNA onto the mitotic chromosomes. Whereas the mechanisms of viral DNA replication are well documented, those involved in tethering EBNA1 to the cellular chromatin are far from being understood. Here, we have identified Regulator of Chromosome Condensation 1 (RCC1) as a novel cellular partner for EBNA1. RCC1 is the major nuclear guanine nucleotide exchange factor (RanGEF) for the small GTPase Ran enzyme. RCC1, associated with chromatin, is involved in the formation of RanGTP gradients critical for nucleo-cytoplasmic transport, mitotic spindle formation, and nuclear envelope reassembly following mitosis. Using several approaches, we have demonstrated a direct interaction between these two proteins and found that the EBNA1 domains responsible for EBNA1 tethering to the mitotic chromosomes are also involved in the interaction with RCC1. The use of an EBNA1 peptide array confirmed the interaction of RCC1 with these regions and also the importance of the N-terminal region of RCC1 in this interaction. Finally, using confocal microscopy and FRET analysis to follow the dynamics of interaction between the two proteins throughout the cell cycle, we have demonstrated that EBNA1 and RCC1 closely associate on the chromosomes during metaphase, suggesting an essential role for the interaction during this phase, perhaps in tethering EBNA1 to mitotic chromosomes

Hydrothermal activity along the slow-spreading Lucky Strike ridge segment (Mid-Atlantic Ridge) : distribution, heatflux, and geological controls

Author Posting. © The Author(s), 2015. This is the author's version of the work. It is posted here by permission of Elsevier for personal use, not for redistribution. The definitive version was published in Earth and Planetary Science Letters 431 (2015): 1730185, doi:10.1016/j.epsl.2015.09.025.We have reviewed available visual information from the seafloor, and recently acquired microbathymetry for several traverses across the Lucky Strike segment, to evaluate the distribution of hydrothermal activity. We have identified a new on-axis site with diffuse flow, Ewan, and anactive vent structure ~1.2 km from the axis, Capelinhos. These sites are minor relative to the Main field, and our total heatflux estimate for all active sites (200-1200 MW) is only slightly higher than previously published estimates. We also identify fossil sites W of the main Lucky Strike field. A circular feature ~200 m in diameter located on the flanks of a rifted off-axis central volcano, is likely a large and inactive hydrothermal edifice, named Grunnus. We find no indicator of focused hydrothermal activity elsewhere along the segment, suggesting that the enhanced melt supply and the associated melt lenses, required to form central volcanoes, also sustain hydrothermal circulation to form and maintain large and long-lived hydrothermal fields. Hydrothermal discharge to the seafloor occurs along fault traces, suggesting focusing of hydrothermal circulation in the shallow crust along permeable fault zones.This work has been partly financed by ANR (France) Mothseim Project NT05-3 42213 toJE, and by EU-RTN-MOMARNET to MC. The French Ministry of Research financed ship, ROV and AUV time (Graviluck’06, MOMAR’08, Bathyluck’09, MOMARSAT cruises in 2010-2015

Cell therapy of Duchenne muscular dystrophy: preclinical trial in GRMD dogs

Duchenne muscular dystrophy (DMD), a genetic progressive X-linked muscular dystrophy, is the most common genetic disease in humans. Cell therapy based on the use of somatic stem cells is a very promising approach. In a dog myopathy model, we isolated a muscle stem cell (MuStem) with the essential requirements for therapeutic use: high amplification capacity, ability to fuse with muscle fibers, renewal of the satellite cell population, dispersion in the whole body after vascular administration, persistence of long-term effect, and dramatic clinical improvement of treated animals. These preclinical results pave the way for a therapeutic trial in children with Duchenne muscular dystrophy.La dystrophie musculaire de Duchenne (DMD) est une maladie génétique progressive du muscle liée au chromosome X. Elle est la maladie génétique la plus fréquente chez l'homme. La thérapie cellulaire basée sur l'utilisation de cellules souches somatiques est une voie thérapeutique riche d'intérêt. Nous avons isolé, chez un modèle de chien myopathe, une cellule souche musculaire (MuStem) qui présente les qualités indispensables à une utilisation thérapeutique: forte capacité d'amplification, capacité à fusionner avec les fibres musculaires, renouvellement du contingent de cellules satellites, dispersion dans l'organisme après administration vasculaire, persistance de l'effet à long terme, spectaculaire amélioration clinique des animaux traités. Ces résultats précliniques ouvrent la voie à un essai thérapeutique chez l'enfant atteint de dystrophie musculaire de Duchenne

Global value perceptions : the legitimising functions of western representations of democracy

This paper argues that a fundamental antagonism between democracy and nondemocracy organises lay thinking on global issues. We review key findings of a long-standing experimental research programme that examined the "Democracy-as-value" hypothesis across a variety of political and social contexts. This hypothesis contends that democracy is an ideological belief system that provides value to democratic individuals, groups, and institutions and thereby grants legitimacy to their actions. Based on procedural justice theories and social representations theory, we contend that western lay perceivers associate democracy with procedural equality and individual autonomy, whereas nondemocracy is associated with ingroup hierarchy and conformity. We discuss how idealised representations of democracy justify global power arrangements and emphasise the paradoxical justification function of democratic values through which nondemocratic forms of social regulation based on physical force are legitimised with the very democratic norms that call for peaceful resolution of conflicts

Role of the interaction between the viral protein EBNA1 and the cellular factor RCC1 for the persistance of the Epstein-Barr Virus genome

Le virus d’Epstein-Barr (EBV) est un herpesvirus dont la séroprévalence est d’environ 90 % de la population adulte mondiale. EBV est associé à de nombreuses pathologies tumorales. La primo infection conduit à l’établissement du virus sous forme latente dans les lymphocytes B mémoires. Au sein de ces cellules B, le génome viral est sous la forme d’un épisome, un ADN circulaire double brin, et une fraction restreinte de gènes viraux est exprimée. Afin de se maintenir aux cours des divisions cellulaires, le génome viral est répliqué en phase S par la machinerie cellulaire et ségrégé lors de la mitose dans chaque cellules filles. La réplication et la ségrégation du génome viral nécessitent 2 facteurs viraux que sont la protéine virale EBNA1 (Epstein-Barr Nuclear Antigen 1) et la région oriP sur le génome viral. En phase S, EBNA1 interagit directement avec l’oriP et y recrute le complexe de pré-réplication de l’ADN. En mitose, EBNA1 ancre l’épisome à la chromatine ce qui permet une ségrégation efficace. Les mécanismes d’interaction entre EBNA1 et la chromatine reste encore flou. Au cours de notre travail, nous avons identifié la protéine RCC1 comme un partenaire potentiel pour la protéine EBNA1 pouvant être impliqué dans l’ancrage d’EBNA1 à la chromatine. Nous avons validé cette interaction et caractérisé les régions d’interactions pour ces deux protéines. Par ailleurs nous avons démontré que RCC1 est recrutée sur l’oriP en présence d’EBNA1 et que ces deux protéines interagissent en mitose. À la lumière de nos résultats et des données de la littérature, nous proposons que l’interaction d’EBNA1 avec la chromatine est dynamique et implique à la fois des interactions directes (AT-Hook, interaction avec les nucléosomes) mais aussi des facteurs cellulaires (RCC1, EBP2 et HMGB2).Epstein-Barr virus (EBV) is a ubiquitous herpesvirus associated with several human cancers. In proliferating latently-infected cells, the EBV genome persists as a circular plasmid that is replicated once per cell cycle and partitioned at mitosis. Both of these processes require a single viral protein, Epstein Barr nuclear antigen 1 (EBNA1), which binds to two clusters of cognate binding sites within the origin of plasmid replication (oriP). EBNA1 plays an essential role both in viral episome replication, by recruiting the cellular complex of DNA replication onto the oriP, and in the efficient segregation of the viral episomes, by tethering the viral DNA onto the mitotic chromosomes. Whereas the mechanisms of viral DNA replication have been well documented, the mechanisms involved in tethering EBNA1 to the cellular chromatin are far from being understood. Here we have identified Regulator of Chromosome Condensation 1 (RCC1) as a novel EBNA1 cellular partner. RCC1 is the only known nuclear guanine nucleotide exchange factor (RanGEF) for the small GTPase Ran enzyme. RCC1, associated with chromatin, is involved in the formation of RanGTP gradients critical for nucleo-cytoplasmic transport, mitotic spindle formation, and nuclear envelope reassembly after mitosis. We have used several approaches to demonstrate a direct interaction between these two proteins and to identify the regions. involved Moreover, by using Chromatin ImmunoPrecipitation assay (ChIP) we have shown that RCC1 is enriched in the oriP region of mini viral replicons in a manner dependent on EBNA1. Finally, by using a combination of confocal microscopy and FRET analysis to follow the dynamics of interaction between the two proteins throughout the cell cycle, we have demonstrated that EBNA1 and RCC1 closely associate on the chromosomes during metaphase. Taken together, our data strongly suggest an essential role for RCC1 in tethering EBNA1 - linked to the viral episome - to the metaphasic chromosomes. Our results and those of others lead us to the idea that the interaction between EBNA1 with the cellular chromosomes requires several factors such as direct interactions or cellular proteins and these interactions are complementary and / or redundant

Extracellular vesicles during Herpes Simplex Virus type 1 infection: an inquire

AbstractExtracellular vesicles are defined as a heterogeneous group of vesicles that are released by prokaryotic to highereukaryotic cells and by plant cells in an evolutionary conserved manner. The significance of these vesicles lies intheir capacity to transfer selected cargo composed of proteins, lipids and nucleic acids to both recipient and parentcells and to influence various physiological and pathological functions. Microorganisms such as parasites, fungi andprotozoa and even single cell organisms such as bacteria generate extracellular vesicles. In addition, several viruseshave evolved strategies to hijack the extracellular vesicles for egress or to alter the surrounding environment. Thethesis of this article is that: a) during HSV-1 infection vesicles are delivered from infected to uninfected cells thatinfluence the infection; b) the cargo of these vesicles consists of viral and host transcripts (mRNAs, miRNAs andnon-coding RNAs) and proteins including innate immune components, such as STING; and c) the viral vesicles carrythe tetraspanins CD9, CD63 and CD81, which are considered as markers of exosomes. Therefore, we assume thatthe STING-carrying vesicles, produced during HSV-1 infection, are reminiscent to exosomes. The presumed functionsof the exosomes released from HSV-1 infected cells include priming the recipient cells and accelerating antiviralresponses to control the dissemination of the virus. This may be one strategy used by the virus to prevent theelimination by the host and establish persistent infection. In conclusion, the modification of the cargo of exosomesappears to be part of the strategy that HSV-1 has evolved to establish lifelong persistent infections into the humanbody to ensure successful dissemination between individuals

Foreword

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