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Accueil > Départements > Microbiologie > Jacques OBERTO : Biologie Cellulaire des Archées

Publications de l’équipe

2017


  • D. A. Braun, J. Rao, G. Mollet, D. Schapiro, M. - C. Daugeron, W. Tan, O. Gribouval, O. Boyer, P. Revy, T. Jobst-Schwan, J. M. Schmidt, J. A. Lawson, D. Schanze, S. Ashraf, J. F. P. Ullmann, C. A. Hoogstraten, N. Boddaert, B. Collinet, G. Martin, D. Liger, S. Lovric, M. Furlano, I. C. Guerrera, O. Sanchez-Ferras, J. F. Hu, A. - C. Boschat, S. Sanquer, B. Menten, S. Vergult, N. De Rocker, M. Airik, T. Hermle, S. Shril, E. Widmeier, H. Y. Gee, W. - I. Choi, C. E. Sadowski, W. L. Pabst, J. K. Warejko, A. Daga, T. Basta, V. Matejas, K. Scharmann, S. D. Kienast, B. Behnam, B. Beeson, A. Begtrup, M. Bruce, G. - S. Ch'ng, S. - P. Lin, J. - H. Chang, C. - H. Chen, M. T. Cho, P. M. Gaffney, P. E. Gipson, C. - H. Hsu, J. A. Kari, Y. - Y. Ke, C. Kiraly-Borri, W. - M. Lai, E. Lemyre, R. O. Littlejohn, A. Masri, M. Moghtaderi, K. Nakamura, F. Ozaltin, M. Praet, C. Prasad, A. Prytula, E. R. Roeder, P. Rump, R. E. Schnur, T. Shiihara, M. D. Sinha, N. A. Soliman, K. Soulami, D. A. Sweetser, W. - H. Tsai, J. - D. Tsai, R. Topaloglu, U. Vester, D. H. Viskochil, N. Vatanavicharn, J. L. Waxler, K. J. Wierenga, M. T. F. Wolf, S. - N. Wong, S. A. Leidel, G. Truglio, P. C. Dedon, A. Poduri, S. Mane, R. P. Lifton, M. Bouchard, P. Kannu, D. Chitayat, D. Magen, B. Callewaert, H. van Tilbeurgh, M. Zenker, C. Antignac, et F. Hildebrandt, « Mutations in KEOPS-complex genes cause nephrotic syndrome with primary microcephaly », Nature Genetics, 2017.
    Résumé : Galloway-Mowat syndrome (GAMOS) is an autosomal-recessive disease characterized by the combination of early-onset nephrotic syndrome (SRNS) and microcephaly with brain anomalies. Here we identified recessive mutations in OSGEP, TP53RK, TPRKB, and LAGE3, genes encoding the four subunits of the KEOPS complex, in 37 individuals from 32 families with GAMOS. CRISPR-Cas9 knockout in zebrafish and mice recapitulated the human phenotype of primary microcephaly and resulted in early lethality. Knockdown of OSGEP, TP53RK, or TPRKB inhibited cell proliferation, which human mutations did not rescue. Furthermore, knockdown of these genes impaired protein translation, caused endoplasmic reticulum stress, activated DNA-damage-response signaling, and ultimately induced apoptosis. Knockdown of OSGEP or TP53RK induced defects in the actin cytoskeleton and decreased the migration rate of human podocytes, an established intermediate phenotype of SRNS. We thus identified four new monogenic causes of GAMOS, describe a link between KEOPS function and human disease, and delineate potential pathogenic mechanisms.
    Mots-clés : ARCHEE, B3S, FAAM, MICROBIO.

  • M. Cossu, C. Badel, R. Catchpole, D. Gadelle, E. Marguet, V. Barbe, P. Forterre, et J. Oberto, « Flipping chromosomes in deep-sea archaea », PLoS genetics, vol. 13, nᵒ 6, p. e1006847, 2017.
    Résumé : One of the major mechanisms driving the evolution of all organisms is genomic rearrangement. In hyperthermophilic Archaea of the order Thermococcales, large chromosomal inversions occur so frequently that even closely related genomes are difficult to align. Clearly not resulting from the native homologous recombination machinery, the causative agent of these inversions has remained elusive. We present a model in which genomic inversions are catalyzed by the integrase enzyme encoded by a family of mobile genetic elements. We characterized the integrase from Thermococcus nautili plasmid pTN3 and showed that besides canonical site-specific reactions, it catalyzes low sequence specificity recombination reactions with the same outcome as homologous recombination events on DNA segments as short as 104bp both in vitro and in vivo, in contrast to other known tyrosine recombinases. Through serial culturing, we showed that the integrase-mediated divergence of T. nautili strains occurs at an astonishing rate, with at least four large-scale genomic inversions appearing within 60 generations. Our results and the ubiquitous distribution of pTN3-like integrated elements suggest that a major mechanism of evolution of an entire order of Archaea results from the activity of a selfish mobile genetic element.
    Mots-clés : ARCHEE, MICROBIO.

  • V. Da Cunha, M. Gaia, D. Gadelle, A. Nasir, et P. Forterre, « Lokiarchaea are close relatives of Euryarchaeota, not bridging the gap between prokaryotes and eukaryotes », PLoS genetics, vol. 13, nᵒ 6, p. e1006810, 2017.
    Résumé : The eocyte hypothesis, in which Eukarya emerged from within Archaea, has been boosted by the description of a new candidate archaeal phylum, "Lokiarchaeota", from metagenomic data. Eukarya branch within Lokiarchaeota in a tree reconstructed from the concatenation of 36 universal proteins. However, individual phylogenies revealed that lokiarchaeal proteins sequences have different evolutionary histories. The individual markers phylogenies revealed at least two subsets of proteins, either supporting the Woese or the Eocyte tree of life. Strikingly, removal of a single protein, the elongation factor EF2, is sufficient to break the Eukaryotes-Lokiarchaea affiliation. Our analysis suggests that the three lokiarchaeal EF2 proteins have a chimeric organization that could be due to contamination and/or homologous recombination with patches of eukaryotic sequences. A robust phylogenetic analysis of RNA polymerases with a new dataset indicates that Lokiarchaeota and related phyla of the Asgard superphylum are sister group to Euryarchaeota, not to Eukarya, and supports the monophyly of Archaea with their rooting in the branch leading to Thaumarchaeota.
    Mots-clés : Archaeal Proteins, ARCHEE, Eukaryota, Euryarchaeota, Evolution, Molecular, MICROBIO, Phylogeny, Prokaryotic Cells.

  • N. Dautin, C. de Sousa-d'Auria, F. Constantinesco-Becker, C. Labarre, J. Oberto, I. L. de la Sierra-Gallay, C. Dietrich, H. Issa, C. Houssin, et N. Bayan, « Mycoloyltransferases: A large and major family of enzymes shaping the cell envelope of Corynebacteriales », Biochimica et Biophysica Acta (BBA) - General Subjects, vol. 1861, nᵒ 1 Pt B, p. 3581-3592, 2017.
    Résumé : Mycobacterium and Corynebacterium are important genera of the Corynebacteriales order, the members of which are characterized by an atypical diderm cell envelope. Indeed the cytoplasmic membrane of these bacteria is surrounded by a thick mycolic acid-arabinogalactan-peptidoglycan (mAGP) covalent polymer. The mycolic acid-containing part of this complex associates with other lipids (mainly trehalose monomycolate (TMM) and trehalose dimycolate (TDM)) to form an outer membrane. The metabolism of mycolates in the cell envelope is governed by esterases called mycoloyltransferases that catalyze the transfer of mycoloyl chains from TMM to another TMM molecule or to other acceptors such as the terminal arabinoses of arabinogalactan or specific polypeptides. In this review we present an overview of this family of Corynebacteriales enzymes, starting with their expression, localization, structure and activity to finally discuss their putative functions in the cell. In addition, we show that Corynebacteriales possess multiple mycoloyltransferases encoding genes in their genome. The reason for this multiplicity is not known, as their function in mycolates biogenesis appear to be only partially redundant. It is thus possible that, in some species living in specific environments, some mycoloyltransferases have evolved to gain some new functions. In any case, the few characterized mycoloyltransferases are very important for the bacterial physiology and are also involved in adaptation in the host where they constitute major secreted antigens. Although not discussed in this review, all these functions make them interesting targets for the discovery of new antibiotics and promising vaccines candidates. This article is part of a Special Issue entitled "Science for Life" Guest Editor: Dr. Austen Angell, Dr. Salvatore Magazù and Dr. Federica Migliardo.
    Mots-clés : Antigen 85, ARCHEE, CORYNE, Esterase, Fibronectin-binding protein, MICROBIO, Mycobacterium, Mycolyltransferases, Mycomembrane.

  • P. Forterre, « Viruses in the 21st Century: From the Curiosity-Driven Discovery of Giant Viruses to New Concepts and Definition of Life », Clinical Infectious Diseases: An Official Publication of the Infectious Diseases Society of America, vol. 65, nᵒ suppl_1, p. S74-S79, 2017.
    Résumé : The curiosity-driven discovery of giant DNA viruses infecting amoebas has triggered an intense debate about the origin, nature, and definition of viruses. This discovery was delayed by the current paradigm confusing viruses with small virions. Several new definitions and concepts have been proposed either to reconcile the unique features of giant viruses with previous paradigms or to propose a completely new vision of the living world. I briefly review here how several other lines of research in virology converged during the last 2 decades with the discovery of giant viruses to change our traditional perception of the viral world. This story emphasizes the power of multidisciplinary curiosity-driven research, from the hospital to the field and the laboratory. Notably, some philosophers have now also joined biologists in their quest to make sense of the abundance and diversity of viruses and related capsidless mobile elements in the biosphere.
    Mots-clés : ARCHEE, giant viruses, Life definition, MICROBIO, phage, Virion, Virocell.

2016


  • P. Béguin, N. Charpin, E. V. Koonin, P. Forterre, et M. Krupovic, « Casposon integration shows strong target site preference and recapitulates protospacer integration by CRISPR-Cas systems », Nucleic Acids Research, vol. 44, nᵒ 21, p. 10367-10376, 2016.
    Résumé : Casposons are a recently discovered group of large DNA transposons present in diverse bacterial and archaeal genomes. For integration into the host chromosome, casposons employ an endonuclease that is homologous to the Cas1 protein involved in protospacer integration by the CRISPR-Cas adaptive immune system. Here we describe the site-preference of integration by the Cas1 integrase (casposase) encoded by the casposon of the archaeon Aciduliprofundum boonei Oligonucleotide duplexes derived from the terminal inverted repeats (TIR) of the A. boonei casposon as well as mini-casposons flanked by the TIR inserted preferentially at a site reconstituting the original A. boonei target site. As in the A. boonei genome, the insertion was accompanied by a 15-bp direct target site duplication (TSD). The minimal functional target consisted of the 15-bp TSD segment and the adjacent 18-bp sequence which comprises the 3' end of the tRNA-Pro gene corresponding to the TΨC loop. The functional casposase target site bears clear resemblance to the leader sequence-repeat junction which is the target for protospacer integration catalyzed by the Cas1-Cas2 adaptation module of CRISPR-Cas. These findings reinforce the mechanistic similarities and evolutionary connection between the casposons and the adaptation module of the prokaryotic adaptive immunity systems.
    Mots-clés : ARCHEE, MICROBIO.

  • N. Dahmane, D. Gadelle, S. Delmas, A. Criscuolo, S. Eberhard, N. Desnoues, S. Collin, H. Zhang, Y. Pommier, P. Forterre, et G. Sezonov, « topIb, a phylogenetic hallmark gene of Thaumarchaeota encodes a functional eukaryote-like topoisomerase IB », Nucleic Acids Research, vol. 44, nᵒ 6, p. 2795-2805, 2016.
    Résumé : Type IB DNA topoisomerases can eliminate torsional stresses produced during replication and transcription. These enzymes are found in all eukaryotes and a short version is present in some bacteria and viruses. Among prokaryotes, the long eukaryotic version is only observed in archaea of the phylum Thaumarchaeota. However, the activities and the roles of these topoisomerases have remained an open question. Here, we demonstrate that all available thaumarchaeal genomes contain a topoisomerase IB gene that defines a monophyletic group closely related to the eukaryotic enzymes. We show that the topIB gene is expressed in the model thaumarchaeon Nitrososphaera viennensis and we purified the recombinant enzyme from the uncultivated thaumarchaeon Candidatus Caldiarchaeum subterraneum. This enzyme is active in vitro at high temperature, making it the first thermophilic topoisomerase IB characterized so far. We have compared this archaeal type IB enzyme to its human mitochondrial and nuclear counterparts. The archaeal enzyme relaxes both negatively and positively supercoiled DNA like the eukaryotic enzymes. However, its pattern of DNA cleavage specificity is different and it is resistant to camptothecins (CPTs) and non-CPT Top1 inhibitors, LMP744 and lamellarin D. This newly described thermostable topoisomerases IB should be a promising new model for evolutionary, mechanistic and structural studies.
    Mots-clés : Amino Acid Sequence, Archaea, Archaeal Proteins, ARCHEE, Camptothecin, Cloning, Molecular, Coumarins, DNA Topoisomerases, Type I, DNA, Superhelical, Escherichia coli, gene expression, Heterocyclic Compounds, 4 or More Rings, Hot Temperature, Humans, Isoquinolines, MICROBIO, Mitochondrial Proteins, Models, Molecular, Molecular Sequence Data, Phylogeny, Protein Stability, Protein Structure, Secondary, Protein Structure, Tertiary, Recombinant Proteins, Sequence Alignment, Topoisomerase I Inhibitors.

  • P. Forterre, « To be or not to be alive: How recent discoveries challenge the traditional definitions of viruses and life », Studies in History and Philosophy of Biological and Biomedical Sciences, vol. 59, p. 100-108, 2016.
    Résumé : Three major discoveries have recently profoundly modified our perception of the viral world: molecular ecologists have shown that viral particles are more abundant than cells in natural environments; structural biologists have shown that some viruses from the three domains of life, Bacteria, Eukarya and Archaea, are evolutionarily related, and microbiologists have discovered giant viruses that rival with cells in terms of size and gene content. I discuss here the scientific and philosophical impact of these discoveries on the debates over the definition, nature (living or not), and origin of viruses. I suggest that viruses have often been considered non-living, because they are traditionally assimilated to their virions. However, the term virus describes a biological process and should integrate all aspects of the viral reproduction cycle. It is especially important to focus on the intracellular part of this cycle, the virocell, when viral information is actively expressed and reproduced, allowing the emergence of new viral genes. The virocell concept theoretically removes roadblocks that prevent defining viruses as living organisms. However, defining a "living organism" remains challenging, as indicated by the case of organelles that evolved from intracellular bacteria. To bypass this problem, I suggest considering that all biological entities that actively participate in the process of life are living.
    Mots-clés : ARCHEE, Bacteriophage, Life definition, MICROBIO, Organism, Virocell, Virus.

  • P. Forterre et M. Gaïa, « Giant viruses and the origin of modern eukaryotes », Current Opinion in Microbiology, vol. 31, p. 44-49, 2016.
    Résumé : Several authors have suggested that viruses from the NucleoCytoplasmic Large DNA Viruses group (NCLDV) have played an important role in the origin of modern eukaryotes. Notably, the viral eukaryogenesis theory posits that the nucleus originated from an ancient NCLDV-related virus. Focusing on the viral factory instead of the virion adds credit to this hypothesis, but also suggests alternative scenarios. Beside a role in the emergence of the nucleus, ancient NCLDV may have provided new genes and/or chromosomes to the proto-eukaryotic lineage. Phylogenetic analyses suggest that NCLDV informational proteins, related to those of Archaea and Eukarya, were either recruited by ancient NCLDV from proto-eukaryotes and/or transferred to proto-eukaryotes, in agreement with the antiquity of NCLDV and their possible role in eukaryogenesis.
    Mots-clés : ARCHEE, MICROBIO.

  • M. Krupovic, S. Shmakov, K. S. Makarova, P. Forterre, et E. V. Koonin, « Recent Mobility of Casposons, Self-Synthesizing Transposons at the Origin of the CRISPR-Cas Immunity », Genome Biology and Evolution, vol. 8, nᵒ 2, p. 375-386, 2016.
    Résumé : Casposons are a superfamily of putative self-synthesizing transposable elements that are predicted to employ a homolog of Cas1 protein as a recombinase and could have contributed to the origin of the CRISPR-Cas adaptive immunity systems in archaea and bacteria. Casposons remain uncharacterized experimentally, except for the recent demonstration of the integrase activity of the Cas1 homolog, and given their relative rarity in archaea and bacteria, original comparative genomic analysis has not provided direct indications of their mobility. Here, we report evidence of casposon mobility obtained by comparison of the genomes of 62 strains of the archaeon Methanosarcina mazei. In these genomes, casposons are variably inserted in three distinct sites indicative of multiple, recent gains, and losses. Some casposons are inserted into other mobile genetic elements that might provide vehicles for horizontal transfer of the casposons. Additionally, many M. mazei genomes contain previously undetected solo terminal inverted repeats that apparently are derived from casposons and could resemble intermediates in CRISPR evolution. We further demonstrate the sequence specificity of casposon insertion and note clear parallels with the adaptation mechanism of CRISPR-Cas. Finally, besides identifying additional representatives in each of the three originally defined families, we describe a new, fourth, family of casposons.
    Mots-clés : Archaeal Proteins, ARCHEE, Base Sequence, casposons, Clustered Regularly Interspaced Short Palindromic Repeats, CRISPR-Cas, DNA Transposable Elements, Endodeoxyribonucleases, Gene Transfer, Horizontal, Genome, Archaeal, Methanosarcina barkeri, MICROBIO, mobile genetic elements, Molecular Sequence Data, Phylogeny, self-synthesizing transposons, transposition.

  • E. R. J. Quemin, P. Chlanda, M. Sachse, P. Forterre, D. Prangishvili, et M. Krupovic, « Eukaryotic-Like Virus Budding in Archaea », mBio, vol. 7, nᵒ 5, 2016.
    Résumé : Similar to many eukaryotic viruses (and unlike bacteriophages), viruses infecting archaea are often encased in lipid-containing envelopes. However, the mechanisms of their morphogenesis and egress remain unexplored. Here, we used dual-axis electron tomography (ET) to characterize the morphogenesis of Sulfolobus spindle-shaped virus 1 (SSV1), the prototype of the family Fuselloviridae and representative of the most abundant archaea-specific group of viruses. Our results show that SSV1 assembly and egress are concomitant and occur at the cellular cytoplasmic membrane via a process highly reminiscent of the budding of enveloped viruses that infect eukaryotes. The viral nucleoprotein complexes are extruded in the form of previously unknown rod-shaped intermediate structures which have an envelope continuous with the host membrane. Further maturation into characteristic spindle-shaped virions takes place while virions remain attached to the cell surface. Our data also revealed the formation of constricted ring-like structures which resemble the budding necks observed prior to the ESCRT machinery-mediated membrane scission during egress of various enveloped viruses of eukaryotes. Collectively, we provide evidence that archaeal spindle-shaped viruses contain a lipid envelope acquired upon budding of the viral nucleoprotein complex through the host cytoplasmic membrane. The proposed model bears a clear resemblance to the egress strategy employed by enveloped eukaryotic viruses and raises important questions as to how the archaeal single-layered membrane composed of tetraether lipids can undergo scission. IMPORTANCE: The replication of enveloped viruses has been extensively studied in eukaryotes but has remained unexplored for enveloped viruses infecting Archaea Here, we provide a sequential view on the assembly and egress of SSV1, a prototypic archaeal virus. The observed process is highly similar to the budding of eukaryotic enveloped viruses, including human immunodeficiency virus, influenza virus, and Ebola virus. The present study is the first to characterize such a phenomenon in archeal cells, showing that membrane budding is not an exclusive feature of eukaryotic viruses. Our results provide significant insights into the biogenesis and architecture of unique, spindle-shaped virions that infect archaea. Furthermore, our findings open doors for future inquiries into (i) the evolution of the virus budding process, (ii) mechanistic details of virus-mediated membrane scission in Archaea, and (iii) elucidation of virus- and host-encoded molecular players responsible for archaeal membrane and surface remodeling.
    Mots-clés : ARCHEE, MICROBIO.

  • G. Richarme, E. Marguet, P. Forterre, S. Ishino, et Y. Ishino, « DJ-1 family Maillard deglycases prevent acrylamide formation », Biochemical and Biophysical Research Communications, vol. 478, nᵒ 3, p. 1111-1116, 2016.
    Résumé : The presence of acrylamide in food is a worldwide concern because it is carcinogenic, reprotoxic and neurotoxic. Acrylamide is generated in the Maillard reaction via condensation of reducing sugars and glyoxals arising from their decomposition, with asparagine, the amino acid forming the backbone of the acrylamide molecule. We reported recently the discovery of the Maillard deglycases (DJ-1/Park7 and its prokaryotic homologs) which degrade Maillard adducts formed between glyoxals and lysine or arginine amino groups, and prevent glycation damage in proteins. Here, we show that these deglycases prevent acrylamide formation, likely by degrading asparagine/glyoxal Maillard adducts. We also report the discovery of a deglycase from the hyperthermophilic archaea Pyrococcus furiosus, which prevents acrylamide formation at 100 °C. Thus, Maillard deglycases constitute a unique enzymatic method to prevent acrylamide formation in food without depleting the components (asparagine and sugars) responsible for its formation.
    Mots-clés : Acrylamide, Advanced glycation endproducts, AGEs, ARCHEE, Asparagine, cancer, Food polluant, Fructose, Glucose, Glycation, Glyoxal, Maillard reaction, Methylglyoxal, MICROBIO.

  • S. Roux, F. Enault, V. Ravet, J. Colombet, Y. Bettarel, J. - C. Auguet, T. Bouvier, S. Lucas-Staat, A. Vellet, D. Prangishvili, P. Forterre, D. Debroas, et T. Sime-Ngando, « Analysis of metagenomic data reveals common features of halophilic viral communities across continents », Environmental Microbiology, vol. 18, nᵒ 3, p. 889-903, 2016.
    Résumé : Microbial communities from hypersaline ponds, dominated by halophilic archaea, are considered specific of such extreme conditions. The associated viral communities have accordingly been shown to display specific features, such as similar morphologies among different sites. However, little is known about the genetic diversity of these halophilic viral communities across the Earth. Here, we studied viral communities in hypersaline ponds sampled on the coast of Senegal (8-36% of salinity) using metagenomics approach, and compared them with hypersaline viromes from Australia and Spain. The specificity of hyperhalophilic viruses could first be demonstrated at a community scale, salinity being a strong discriminating factor between communities. For the major viral group detected in all samples (Caudovirales), only a limited number of halophilic Caudovirales clades were highlighted. These clades gather viruses from different continents and display consistent genetic composition, indicating that they represent related lineages with a worldwide distribution. Non-tailed hyperhalophilic viruses display a greater rate of gene transfer and recombination, with uncharacterized genes conserved across different kind of viruses and plasmids. Thus, hypersaline viral communities around the world appear to form a genetically consistent community that are likely to harbour new genes coding for enzymes specifically adapted to these environments.
    Mots-clés : ARCHEE, Australia, Caudovirales, Chromosome Mapping, Genetic Variation, Genome, Viral, metagenomics, MICROBIO, Ponds, Salinity, Senegal, Spain.

2015


  • P. Béguin, S. Gill, N. Charpin, et P. Forterre, « Synergistic template-free synthesis of dsDNA by Thermococcus nautili primase PolpTN2, DNA polymerase PolB, and pTN2 helicase », Extremophiles: Life Under Extreme Conditions, vol. 19, nᵒ 1, p. 69-76, 2015.
    Résumé : A combination of three enzymes from the hyperthermophilic archaeon Thermococcus nautili, DNA primase PolpTN2, DNA polymerase PolB, and pTN2 DNA helicase, was found to synthesize up to 300-400 ng/µl dsDNA from deoxynucleotide triphosphates in less than 30 min in the absence of added template DNA and oligonucleotide primer. The reaction did not occur below 64 °C. No synthesis was observed if PolpTN2 or PolB were left out; helicase was not essential but accelerated the reaction. The DNA synthesized consisted of highly reiterated palindromic sequences reaching up to more that 10 kb. Sequence analysis of three independent reaction products synthesized at different temperatures showed that the palindromes shared a common pentanucleotide core, suggesting that random nucleic acid fragments were not responsible for priming the reaction. When enzymes were added sequentially, preincubation with primase plus helicase followed by PolB led to a shorter delay before the onset of the reaction as compared to preincubation with PolB plus helicase followed by primase. This suggests that the primase generates seeds that are subsequently amplified and elongated in synergy with PolB by a mechanism involving hairpin formation and slippage synthesis.
    Mots-clés : ARCHEE, Base Sequence, Cloning, Molecular, DNA, DNA Helicases, DNA Polymerase II, DNA Primase, DNA Primers, Electrophoresis, Agar Gel, MICROBIO, Micrococcal Nuclease, Molecular Sequence Data, Nucleic Acids, Temperature, Thermococcus.

  • D. Bréchemier-Baey, L. Domínguez-Ramírez, J. Oberto, et J. Plumbridge, « Operator recognition by the ROK transcription factor family members, NagC and Mlc », Nucleic Acids Research, vol. 43, nᵒ 1, p. 361-372, 2015.
    Résumé : NagC and Mlc, paralogous members of the ROK family of proteins with almost identical helix-turn-helix DNA binding motifs, specifically regulate genes for transport and utilization of N-acetylglucosamine and glucose. We previously showed that two amino acids in a linker region outside the canonical helix-turn-helix motif are responsible for Mlc site specificity. In this work we identify four amino acids in the linker, which are required for recognition of NagC targets. These amino acids allow Mlc and NagC to distinguish between a C/G and an A/T bp at positions ±11 of the operators. One linker position, glycine in NagC and arginine in Mlc, corresponds to the major specificity determinant for the two proteins. In certain contexts it is possible to switch repression from Mlc-style to NagC-style, by interchanging this glycine and arginine. Secondary determinants are supplied by other linker positions or the helix-turn-helix motif. A wide genomic survey of unique ROK proteins shows that glycine- and arginine-rich sequences are present in the linkers of nearly all ROK family repressors. Conserved short sequence motifs, within the branches of the ROK evolutionary tree, suggest that these sequences could also be involved in operator recognition in other ROK family members.
    Mots-clés : Amino Acid Motifs, ARCHEE, Binding Sites, Escherichia coli Proteins, Gene Expression Regulation, Bacterial, MICROBIO, Mutation, Operator Regions, Genetic, Protein Binding, Repressor Proteins.

  • M. Cossu, V. Da Cunha, C. Toffano-Nioche, P. Forterre, et J. Oberto, « Comparative genomics reveals conserved positioning of essential genomic clusters in highly rearranged Thermococcales chromosomes », Biochimie, vol. 118, p. 313-321, 2015.
    Résumé : The genomes of the 21 completely sequenced Thermococcales display a characteristic high level of rearrangements. As a result, the prediction of their origin and termination of replication on the sole basis of chromosomal DNA composition or skew is inoperative. Using a different approach based on biologically relevant sequences, we were able to determine oriC position in all 21 genomes. The position of dif, the site where chromosome dimers are resolved before DNA segregation could be predicted in 19 genomes. Computation of the core genome uncovered a number of essential gene clusters with a remarkably stable chromosomal position across species, in sharp contrast with the scrambled nature of their genomes. The active chromosomal reorganization of numerous genes acquired by horizontal transfer, mainly from mobile elements, could explain this phenomenon.
    Mots-clés : Archaea, ARCHEE, Base Sequence, Bioinformatics, Chromosomal landmarks, Chromosomes, Comparative Genomic Hybridization, DBG, Evolution, Molecular, Gene Rearrangement, Genes, Archaeal, Genome, Genome evolution, MICROBIO, Mobile elements, Molecular Sequence Data, SSFA, Thermococcales.

  • P. Forterre, « The universal tree of life: an update », Frontiers in Microbiology, vol. 6, p. 717, 2015.
    Résumé : Biologists used to draw schematic "universal" trees of life as metaphors illustrating the history of life. It is indeed a priori possible to construct an organismal tree connecting the three major domains of ribosome encoding organisms: Archaea, Bacteria and Eukarya, since they originated by cell division from LUCA. Several universal trees based on ribosomal RNA sequence comparisons proposed at the end of the last century are still widely used, although some of their main features have been challenged by subsequent analyses. Several authors have proposed to replace the traditional universal tree with a ring of life, whereas others have proposed more recently to include viruses as new domains. These proposals are misleading, suggesting that endosymbiosis can modify the shape of a tree or that viruses originated from the last universal common ancestor (LUCA). I propose here an updated version of Woese's universal tree that includes several rootings for each domain and internal branching within domains that are supported by recent phylogenomic analyses of domain specific proteins. The tree is rooted between Bacteria and Arkarya, a new name proposed for the clade grouping Archaea and Eukarya. A consensus version, in which each of the three domains is unrooted, and a version in which eukaryotes emerged within archaea are also presented. This last scenario assumes the transformation of a modern domain into another, a controversial evolutionary pathway. Viruses are not indicated in these trees but are intrinsically present because they infect the tree from its roots to its leaves. Finally, I present a detailed tree of the domain Archaea, proposing the sub-phylum neo-Euryarchaeota for the monophyletic group of euryarchaeota containing DNA gyrase. These trees, that will be easily updated as new data become available, could be useful to discuss controversial scenarios regarding early life evolution.
    Mots-clés : Archaea, ARCHEE, bacteria, eukarya, evolution, LUCA, MICROBIO, universal tree.

  • S. Gélis-Jeanvoine, S. Lory, J. Oberto, et N. Buddelmeijer, « Residues located on membrane-embedded flexible loops are essential for the second step of the apolipoprotein N-acyltransferase reaction », Molecular Microbiology, vol. 95, nᵒ 4, p. 692-705, 2015.
    Résumé : Apolipoprotein N-acyltransferase (Lnt) is an essential membrane-bound enzyme that catalyzes the third and last step in the post-translational modification of bacterial lipoproteins. In order to identify essential residues implicated in substrate recognition and/or binding we screened for non-functional variants of Lnt obtained by error-prone polymerase chain reaction in a complementation assay using a lnt depletion strain. Mutations included amino acid substitutions in the active site and of residues located on flexible loops in the catalytic periplasmic domain. All, but one mutation, led to the formation of the thioester acyl-enzyme intermediate and to the accumulation of apo-Lpp, suggesting that these residues are involved in the second step of the reaction. A large cytoplasmic loop contains a highly conserved region and two hydrophobic segments. Accessibility analysis to alkylating reagents of substituted cysteine residues introduced in this region demonstrated that the hydrophobic segments do not completely span the membrane. Two residues in the highly conserved cytoplasmic region were shown to be essential for Lnt function. Together, our data suggest that amino acids located on flexible cytoplasmic and periplasmic loops, predicted to be membrane embedded, are required for efficient N-acylation of lipoproteins.
    Mots-clés : Acylation, Acyltransferases, Amino Acid Substitution, Apolipoproteins, ARCHEE, Catalytic Domain, Cysteine, Escherichia coli, Escherichia coli Proteins, Genetic Complementation Test, Lipoproteins, MICROBIO, Models, Molecular, Mutation, Phospholipids, Polymerase Chain Reaction, Protein Processing, Post-Translational.

  • M. Krupovic et P. Forterre, « Single-stranded DNA viruses employ a variety of mechanisms for integration into host genomes », Annals of the New York Academy of Sciences, vol. 1341, p. 41-53, 2015.
    Résumé : Single-stranded DNA (ssDNA) viruses are widespread in the environment and include economically, medically, and ecologically important pathogens. Recently, it has been discovered that ssDNA virus genomes are also prevalent in the chromosomes of their bacterial, archaeal, and eukaryotic hosts. Sequences originating from viruses of the families Parvoviridae, Circoviridae, and Geminiviridae are particularly widespread in the genomes of eukaryotes, where they are often fossilized as endogenous viral elements. ssDNA viruses have evolved diverse mechanisms to invade cellular genomes, and these principally vary between viruses infecting bacteria/archaea and eukaryotes. Filamentous bacteriophages (Inoviridae) use at least three major mechanisms of integration. Some of these phages encode integrases of serine or tyrosine recombinase superfamilies, while others utilize DDE transposases of the IS3, IS30, or IS110/IS492 families, whereas some inoviruses, and possibly certain members of the Microviridae, hijack the host XerCD recombination machinery. By contrast, eukaryotic viruses for integration rely on the endonuclease activity of their rolling-circle replication-initiation proteins, mimicking the mechanisms used by some bacterial transposons. Certain bacterial and eukaryotic ssDNA viruses have embraced a transposon-like means of propagation, with occasionally dramatic effects on host genome evolution. Here, we review the diversity of experimentally verified and hypothetical mechanisms of genome integration employed by ssDNA viruses, and consider the evolutionary implications of these processes, particularly in the emergence of novel virus groups.
    Mots-clés : Archaea, ARCHEE, bacteria, DNA Viruses, DNA, Single-Stranded, Eukaryotic Cells, Evolution, Molecular, Genome, Viral, Host-Pathogen Interactions, Integrases, MICROBIO, Models, Genetic, ssDNA viruses, transposition, transposons, virus evolution, Virus Integration.

  • J. Lossouarn, S. Dupont, A. Gorlas, C. Mercier, N. Bienvenu, E. Marguet, P. Forterre, et C. Geslin, « An abyssal mobilome: viruses, plasmids and vesicles from deep-sea hydrothermal vents », Research in Microbiology, vol. 166, nᵒ 10, p. 742-752, 2015.
    Résumé : Mobile genetic elements (MGEs) such as viruses, plasmids, vesicles, gene transfer agents (GTAs), transposons and transpovirions, which collectively represent the mobilome, interact with cellular organisms from all three domains of life, including those thriving in the most extreme environments. While efforts have been made to better understand deep-sea vent microbial ecology, our knowledge of the mobilome associated with prokaryotes inhabiting deep-sea hydrothermal vents remains limited. Here we focus on the abyssal mobilome by reviewing accumulating data on viruses, plasmids and vesicles associated with thermophilic and hyperthermophilic Bacteria and Archaea present in deep-sea hydrothermal vents.
    Mots-clés : (Hyper-)thermophiles, Archaea, ARCHEE, bacteria, Deep-sea hydrothermal vent, Hydrothermal Vents, Interspersed Repetitive Sequences, MICROBIO, Mobilome, Phylogeny, Plasmids, Seawater, Viruses.

  • L. Mora, K. Moncoq, P. England, J. Oberto, et M. de Zamaroczy, « The Stable Interaction Between Signal Peptidase LepB of Escherichia coli and Nuclease Bacteriocins Promotes Toxin Entry into the Cytoplasm », The Journal of Biological Chemistry, vol. 290, nᵒ 52, p. 30783-30796, 2015.
    Résumé : LepB is a key membrane component of the cellular secretion machinery, which releases secreted proteins into the periplasm by cleaving the inner membrane-bound leader. We showed that LepB is also an essential component of the machinery hijacked by the tRNase colicin D for its import. Here we demonstrate that this non-catalytic activity of LepB is to promote the association of the central domain of colicin D with the inner membrane before the FtsH-dependent proteolytic processing and translocation of the toxic tRNase domain into the cytoplasm. The novel structural role of LepB results in a stable interaction with colicin D, with a stoichiometry of 1:1 and a nanomolar Kd determined in vitro. LepB provides a chaperone-like function for the penetration of several nuclease-type bacteriocins into target cells. The colicin-LepB interaction is shown to require only a short peptide sequence within the central domain of these bacteriocins and to involve residues present in the short C-terminal Box E of LepB. Genomic screening identified the conserved LepB binding motif in colicin-like ORFs from 13 additional bacterial species. These findings establish a new paradigm for the functional adaptability of an essential inner-membrane enzyme.
    Mots-clés : Amino Acid Motifs, Amino Acid Sequence, ARCHEE, Bacterial Toxins, bacteriocin, Bacteriocins, Biological Transport, colicin, Cytoplasm, Deoxyribonucleases, endonuclease, Escherichia coli, Escherichia coli Proteins, LepB, membrane, Membrane Proteins, MICROBIO, Microbiology, Molecular Sequence Data, protease, Protein Binding, protein import, Protein Structure, Tertiary, Ribonucleases, Sequence Alignment, Serine Endopeptidases, toxicity.

  • D. Prangishvili, S. Gribaldo, B. Franzetti, et P. Forterre, « Special section on Molecular biology of Archaea », Biochimie, vol. 118, p. 253, 2015.
    Mots-clés : Archaea, ARCHEE, MICROBIO, Phylogeny.

  • E. R. J. Quemin, M. K. Pietilä, H. M. Oksanen, P. Forterre, W. I. C. Rijpstra, S. Schouten, D. H. Bamford, D. Prangishvili, et M. Krupovic, « Sulfolobus Spindle-Shaped Virus 1 Contains Glycosylated Capsid Proteins, a Cellular Chromatin Protein, and Host-Derived Lipids », Journal of Virology, vol. 89, nᵒ 22, p. 11681-11691, 2015.
    Résumé : Geothermal and hypersaline environments are rich in virus-like particles, among which spindle-shaped morphotypes dominate. Currently, viruses with spindle- or lemon-shaped virions are exclusive to Archaea and belong to two distinct viral families. The larger of the two families, the Fuselloviridae, comprises tail-less, spindle-shaped viruses, which infect hosts from phylogenetically distant archaeal lineages. Sulfolobus spindle-shaped virus 1 (SSV1) is the best known member of the family and was one of the first hyperthermophilic archaeal viruses to be isolated. SSV1 is an attractive model for understanding virus-host interactions in Archaea; however, the constituents and architecture of SSV1 particles remain only partially characterized. Here, we have conducted an extensive biochemical characterization of highly purified SSV1 virions and identified four virus-encoded structural proteins, VP1 to VP4, as well as one DNA-binding protein of cellular origin. The virion proteins VP1, VP3, and VP4 undergo posttranslational modification by glycosylation, seemingly at multiple sites. VP1 is also proteolytically processed. In addition to the viral DNA-binding protein VP2, we show that viral particles contain the Sulfolobus solfataricus chromatin protein Sso7d. Finally, we provide evidence indicating that SSV1 virions contain glycerol dibiphytanyl glycerol tetraether (GDGT) lipids, resolving a long-standing debate on the presence of lipids within SSV1 virions. A comparison of the contents of lipids isolated from the virus and its host cell suggests that GDGTs are acquired by the virus in a selective manner from the host cytoplasmic membrane, likely during progeny egress. IMPORTANCE: Although spindle-shaped viruses represent one of the most prominent viral groups in Archaea, structural data on their virion constituents and architecture still are scarce. The comprehensive biochemical characterization of the hyperthermophilic virus SSV1 presented here brings novel and significant insights into the organization and architecture of spindle-shaped virions. The obtained data permit the comparison between spindle-shaped viruses residing in widely different ecological niches, improving our understanding of the adaptation of viruses with unusual morphotypes to extreme environmental conditions.
    Mots-clés : Amino Acid Sequence, Archaeal Proteins, ARCHEE, Capsid Proteins, DNA-Binding Proteins, Fuselloviridae, Genome, Viral, Glycosylation, Haloarcula, Host-Pathogen Interactions, Hydrophobic and Hydrophilic Interactions, Membrane Lipids, MICROBIO, Molecular Sequence Data, Sulfolobus solfataricus, Viral Proteins, Virus Assembly.

  • N. Soler, M. Krupovic, E. Marguet, et P. Forterre, « Membrane vesicles in natural environments: a major challenge in viral ecology », The ISME journal, vol. 9, nᵒ 4, p. 793-796, 2015.
    Mots-clés : ARCHEE, Carbon, Cell Membrane, DNA, Bacterial, MICROBIO, Prochlorococcus, Seawater.
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Bibliographie

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