Author: "Marc Quinternet" - Searchworks@Jio Institute Digital Library Search Results

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1. The box C/D snoRNP assembly factor Bcd1 interacts with the histone chaperone Rtt106 and controls its transcription dependent activity

Author: Benoît Bragantini, Christophe Charron, Maxime Bourguet, Arnaud Paul, Decebal Tiotiu, Benjamin Rothé, Hélène Marty, Guillaume Terral, Steve Hessmann, Laurence Decourty, Marie-Eve Chagot, Jean-Marc Strub, Séverine Massenet, Edouard Bertrand, Marc Quinternet, Cosmin Saveanu, Sarah Cianférani, Stéphane Labialle, Xavier Manival, and Bruno Charpentier
Subjects: Science
Abstract: Biogenesis of small nucleolar RNAs ribonucleoproteins (snoRNPs) requires dedicated assembly machinery. Here, the authors show that a subset of snoRNP assembly factors interacts, genetically or directly, with factors modulating chromatin architecture, suggesting a link between ribosome formation and chromatin functions.
Published: 2021
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2. The RPAP3-Cterminal domain identifies R2TP-like quaternary chaperones

Author: Chloé Maurizy, Marc Quinternet, Yoann Abel, Céline Verheggen, Paulo E. Santo, Maxime Bourguet, Ana C.F. Paiva, Benoît Bragantini, Marie-Eve Chagot, Marie-Cécile Robert, Claire Abeza, Philippe Fabre, Philippe Fort, Franck Vandermoere, Pedro M.F. Sousa, Jean-Christophe Rain, Bruno Charpentier, Sarah Cianférani, Tiago M. Bandeiras, Bérengère Pradet-Balade, Xavier Manival, and Edouard Bertrand
Subjects: Science
Abstract: R2TP is an HSP90 co-chaperone composed of an RPAP3-PIH1D1 heterodimer, which binds two essential AAA+ ATPases RUVBL1/RUVBL2. Here authors use a structural approach to study RPAP3 and find an RPAP3-like protein (SPAG1) which also forms a co-chaperone complex with PIH1D2 and RUVBL1/2 enriched in testis.
Published: 2018
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3. The spliceosomal phosphopeptide P140 controls the lupus disease by interacting with the HSC70 protein and via a mechanism mediated by gammadelta T cells.

Author: Nicolas Page, Nicolas Schall, Jean-Marc Strub, Marc Quinternet, Olivier Chaloin, Marion Décossas, Manh Thong Cung, Alain Van Dorsselaer, Jean-Paul Briand, and Sylviane Muller
Subjects: Medicine, Science
Abstract: The phosphopeptide P140 issued from the spliceosomal U1-70K snRNP protein is recognized by lupus CD4(+) T cells, transiently abolishes T cell reactivity to other spliceosomal peptides in P140-treated MRL/lpr mice, and ameliorates their clinical features. P140 modulates lupus patients' T cell response ex vivo and is currently included in phase IIb clinical trials. Its underlying mechanism of action remains elusive. Here we show that P140 peptide binds a unique cell-surface receptor, the constitutively-expressed chaperone HSC70 protein, known as a presenting-protein. P140 induces apoptosis of activated MRL/lpr CD4(+) T cells. In P140-treated mice, it increases peripheral blood lymphocyte apoptosis and decreases B cell, activated T cell, and CD4(-)CD8(-)B220(+) T cell counts via a specific mechanism strictly depending on gammadelta T cells. Expression of inflammation-linked genes is rapidly regulated in CD4(+) T cells. This work led us to identify a powerful pathway taken by a newly-designed therapeutic peptide to immunomodulate lupus autoimmunity.
Published: 2009
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4. Deciphering cellular and molecular determinants of human DPCD protein in complex with RUVBL1/RUVBL2 AAA-ATPases

Author: Raphael Dos Santos Morais, Paulo E. Santo, Marie Ley, Cédric Schelcher, Yoann Abel, Laura Plassart, Evolène Deslignière, Marie-Eve Chagot, Marc Quinternet, Ana C.F. Paiva, Steve Hessmann, Nelly Morellet, Pedro M. F. Sousa, Franck Vandermoere, Edouard Bertrand, Bruno Charpentier, Tiago M. Bandeiras, Célia Plisson-Chastang, Céline Verheggen, Sarah Cianférani, Xavier Manival, Ingénierie Moléculaire et Physiopathologie Articulaire (IMoPA), Université de Lorraine (UL)-Centre National de la Recherche Scientifique (CNRS), Instituto de Biologia Experimental e Tecnológica (IBET), Instituto de Tecnologia Química e Biológica António Xavier (ITQB), Universidade Nova de Lisboa = NOVA University Lisbon (NOVA), Laboratoire de Spectrométrie de Masse BioOrganique [Strasbourg] (LSMBO), Département Sciences Analytiques et Interactions Ioniques et Biomoléculaires (DSA-IPHC), Institut Pluridisciplinaire Hubert Curien (IPHC), Université de Strasbourg (UNISTRA)-Institut National de Physique Nucléaire et de Physique des Particules du CNRS (IN2P3)-Centre National de la Recherche Scientifique (CNRS)-Université de Strasbourg (UNISTRA)-Institut National de Physique Nucléaire et de Physique des Particules du CNRS (IN2P3)-Centre National de la Recherche Scientifique (CNRS)-Institut Pluridisciplinaire Hubert Curien (IPHC), Université de Strasbourg (UNISTRA)-Institut National de Physique Nucléaire et de Physique des Particules du CNRS (IN2P3)-Centre National de la Recherche Scientifique (CNRS)-Université de Strasbourg (UNISTRA)-Institut National de Physique Nucléaire et de Physique des Particules du CNRS (IN2P3)-Centre National de la Recherche Scientifique (CNRS), Infrastructure Nationale de Protéomique, FR2048 ProFI, Institut de génétique humaine (IGH), Centre National de la Recherche Scientifique (CNRS)-Université de Montpellier (UM), Unité de biologie moléculaire, cellulaire et du développement (MCD), Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Centre National de la Recherche Scientifique (CNRS)-Centre de Biologie Intégrative (CBI), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Centre National de la Recherche Scientifique (CNRS)-Centre National de la Recherche Scientifique (CNRS), Ingénierie, Biologie et Santé en Lorraine (IBSLor), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lorraine (UL)-Centre National de la Recherche Scientifique (CNRS), Institut de Chimie des Substances Naturelles (ICSN), Institut de Chimie du CNRS (INC)-Université Paris-Saclay-Centre National de la Recherche Scientifique (CNRS), Institut de Génomique Fonctionnelle (IGF), Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Université de Montpellier (UM), Université de Strasbourg (UNISTRA)-Institut National de Physique Nucléaire et de Physique des Particules du CNRS (IN2P3)-Centre National de la Recherche Scientifique (CNRS), Centre de Biologie Intégrative (CBI), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Centre National de la Recherche Scientifique (CNRS), ANR-16-CE11-0032,snoRNPASSEMBLY,Comprendre l'assemblage des complexes macro-moléculaires en utilisant les snoRNP C/D comme modèle(2016), CIANFERANI, Sarah, and Comprendre l'assemblage des complexes macro-moléculaires en utilisant les snoRNP C/D comme modèle - - snoRNPASSEMBLY2016 - ANR-16-CE11-0032 - AAPG2016 - VALID
Subjects: [SDV]Life Sciences [q-bio], [SDV.BC]Life Sciences [q-bio]/Cellular Biology, Saxs, Structural Biology, Em, [CHIM] Chemical Sciences, [SDV.BBM] Life Sciences [q-bio]/Biochemistry, Molecular Biology, Humans, [CHIM]Chemical Sciences, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, Lumier-IP, Molecular Biology, [SDV.BC] Life Sciences [q-bio]/Cellular Biology, XL-MS, Dpcd, DNA Helicases, Proteins, structural MS, Itc, Nmr, [SDV] Life Sciences [q-bio], Ruvbl1, Ruvbl2, Multiprotein Complexes, Silac-IP, ATPases Associated with Diverse Cellular Activities, Carrier Proteins
Abstract: International audience; DPCD is a protein that may play a role in cilia formation and whose absence leads to primary ciliary dyskinesia (PCD), a rare disease caused by impairment of ciliated cells. Except for high-throughput studies that identified DPCD as a possible RUVBL1 (R1) and RUVBL2 (R2) partner, no in-depth cellular, biochemical, and structural investigation involving DPCD have been reported so far. R1 and R2 proteins are ubiquitous highly conserved AAA + family ATPases that assemble and mature a plethora of macromolecular complexes and are pivotal in numerous cellular processes, especially by guaranteeing a co-chaperoning function within R2TP or R2TP-like machineries. In the present study, we identified DPCD as a new R1R2 partner in vivo. We show that DPCD interacts directly with R1 and R2 in vitro and in cells. We characterized the physico-chemical properties of DPCD in solution and built a 3D model of DPCD. In addition, we used a variety of orthogonal biophysical techniques including small-angle X-ray scattering, structural mass spectrometry and electron microscopy to assess the molecular determinants of DPCD interaction with R1R2. Interestingly, DPCD disrupts the dodecameric state of R1R2 complex upon binding and this interaction occurs mainly via the DII domains of R1R2.
Published: 2022
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5. NOPCHAP1 is a PAQosome cofactor that helps loading NOP58 on RUVBL1/2 during box C/D snoRNP biogenesis

Author: Franck Vandermoere, Tiago M. Bandeiras, Ana C. F. Paiva, Edouard Bertrand, Bruno Charpentier, Xavier Manival, Céline Verheggen, Marie-Eve Chagot, Pedro M. F. Sousa, Marc Quinternet, Marie-Cécile Robert, Philippe Fort, Paulo E. Santo, Jonathan Bizarro, Yoann Abel, Institut de Génétique Moléculaire de Montpellier (IGMM), Centre National de la Recherche Scientifique (CNRS)-Université de Montpellier (UM), Institut de génétique humaine (IGH), Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS), Instituto de Biologia Experimental e Tecnológica (IBET), Universidade Nova de Lisboa = NOVA University Lisbon (NOVA), Ingénierie Moléculaire et Physiopathologie Articulaire (IMoPA), Université de Lorraine (UL)-Centre National de la Recherche Scientifique (CNRS), Ingénierie, Biologie et Santé en Lorraine (IBSLor), Université de Lorraine (UL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Institut de Génomique Fonctionnelle (IGF), Université de Montpellier (UM)-Université Montpellier 1 (UM1)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Montpellier 2 - Sciences et Techniques (UM2)-Centre National de la Recherche Scientifique (CNRS), Centre de recherche en Biologie Cellulaire (CRBM), Université Montpellier 2 - Sciences et Techniques (UM2)-Centre National de la Recherche Scientifique (CNRS)-Université de Montpellier (UM)-Université Montpellier 1 (UM1), CNRS, Université de Montpellier (UM), Université de Lorraine (UL), ANR, Ligue Nationale contre le cancer, Institut National du cancer, ANR-10-INBS-0004,France-BioImaging,Développment d'une infrastructure française distribuée coordonnée(2010), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lorraine (UL)-Centre National de la Recherche Scientifique (CNRS), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS), Centre de recherche en Biologie cellulaire de Montpellier (CRBM), Verheggen, Céline, and Développment d'une infrastructure française distribuée coordonnée - - France-BioImaging2010 - ANR-10-INBS-0004 - INBS - VALID
Subjects: Proteomics, Saccharomyces cerevisiae Proteins, AcademicSubjects/SCI00010, Recombinant Fusion Proteins, [SDV]Life Sciences [q-bio], ATPase, Mutant, Chaperone, Cofactor, Gene Knockout Techniques, 03 medical and health sciences, Adenosine Triphosphate, 0302 clinical medicine, Protein Domains, Genes, Reporter, Ribonucleoproteins, Small Nucleolar, Protein Interaction Mapping, RNA and RNA-protein complexes, Genetics, Humans, RNP biogenesis, HSP90 Heat-Shock Proteins, Small nucleolar RNA, Eye Proteins, 030304 developmental biology, Multi-subunit complex assembly, 0303 health sciences, PAQosome, biology, DNA Helicases, Nuclear Proteins, Box C/D snoRNP, Hsp90, AAA proteins, Cell biology, [SDV] Life Sciences [q-bio], Multiprotein Complexes, Chaperone (protein), biology.protein, ATPases Associated with Diverse Cellular Activities, Carrier Proteins, 030217 neurology & neurosurgery, Biogenesis, HeLa Cells, Molecular Chaperones
Abstract: The PAQosome is a large complex composed of the HSP90/R2TP chaperone and a prefoldin-like module. It promotes the biogenesis of cellular machineries but it is unclear how it discriminates closely related client proteins. Among the main PAQosome clients are C/D snoRNPs and in particular their core protein NOP58. Using NOP58 mutants and proteomic experiments, we identify different assembly intermediates and show that C12ORF45, which we rename NOPCHAP1, acts as a bridge between NOP58 and PAQosome. NOPCHAP1 makes direct physical interactions with the CC-NOP domain of NOP58 and domain II of RUVBL1/2 AAA+ ATPases. Interestingly, NOPCHAP1 interaction with RUVBL1/2 is disrupted upon ATP binding. Moreover, while it robustly binds both yeast and human NOP58, it makes little interactions with NOP56 and PRPF31, two other closely related CC-NOP proteins. Expression of NOP58, but not NOP56 or PRPF31, is decreased in NOPCHAP1 KO cells. We propose that NOPCHAP1 is a client-loading PAQosome cofactor that selects NOP58 to promote box C/D snoRNP assembly.
Published: 2020
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6. Structural Analysis of the Plasmodial Proteins ZNHIT3 and NUFIP1 Provides Insights into the Selectivity of a Conserved Interaction

Author: Marie-Eve Chagot, Alexis Boutilliat, Alexandre Kriznik, Marc Quinternet, GONNET, JULIE, Ingénierie Moléculaire et Physiopathologie Articulaire (IMoPA), Université de Lorraine (UL)-Centre National de la Recherche Scientifique (CNRS), Ingénierie, Biologie et Santé en Lorraine (IBSLor), and Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lorraine (UL)-Centre National de la Recherche Scientifique (CNRS)
Subjects: ZNHIT3, DLS, Plasmodium falciparum, Biophysics, Protozoan Proteins, AlphaFold, Calorimetry, snoRNA, [SDV.BBM.BM] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, Biochemistry, Antimalarials, Druggability, [SDV.BBM.GTP]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Genomics [q-bio.GN], parasitic diseases, Molecular dynamics simulation, Anopheles, Animals, Humans, Protein interaction, Nuclear Proteins, RNA-Binding Proteins, [SDV.BBM.BM]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, Ribophagy, NMR, Malaria, NUFIP1, [SDV.BBM.GTP] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Genomics [q-bio.GN], snoRNP
Abstract: International audience; Malaria is a widespread and lethal disease caused by the Plasmodium parasites that can infect human beings through Anopheles mosquitoes. For that reason, the biology of Plasmodium needs to be studied to develop antimalarial treatments. By determining the three-dimensional structures of macromolecules, structural biology helps to understand the function of proteins and can reveal how interactions occur between biological partners. Here, we studied the ZNHIT3 and NUFIP1 proteins from Plasmodium falciparum, two proteins tightly linked to the ribosome biology. Due to their important functions in post-translational modifications of ribosomal RNAs and in ribophagy, these proteins participate in the survival of cells. In this study, we solved the three-dimensional structure of a thermally stable and species-dependent complex between fragments of these proteins. Our results were compared to the AlphaFold predictions, which motivated the study of the free ZNHIT3 fragment that binds NUFIP1. We showed that the latter fragment multimerized in vitro but also had the inner ability to change its conformation to escape the solvent exposition of key hydrophobic residues involved in the interaction with NUFIP1. Our data could open the gate to selective drug discovery processes involving these two proteins.
Published: 2022
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7. The interaction between RPAP3 and TRBP reveals a possible involvement of the HSP90/R2TP chaperone complex in the regulation of miRNA activity

Author: Yoann Abel, Christophe Charron, Camille Virciglio, Valérie Bourguignon-Igel, Marc Quinternet, Marie-Eve Chagot, Marie-Cécile Robert, Céline Verheggen, Christiane Branlant, Edouard Bertrand, Xavier Manival, Bruno Charpentier, and Mathieu Rederstorff
Subjects: Ribonuclease III, MicroRNAs, Nuclear Receptor Coactivators, Genetics, RNA-Induced Silencing Complex, Gene Silencing, HSP90 Heat-Shock Proteins, Molecular Chaperones
Abstract: MicroRNAs silence mRNAs by guiding the RISC complex. RISC assembly occurs following cleavage of pre-miRNAs by Dicer, assisted by TRBP or PACT, and the transfer of miRNAs to AGO proteins. The R2TP complex is an HSP90 co-chaperone involved in the assembly of ribonucleoprotein particles. Here, we show that the R2TP component RPAP3 binds TRBP but not PACT. The RPAP3-TPR1 domain interacts with the TRBP-dsRBD3, and the 1.5 Å resolution crystal structure of this complex identifies key residues involved in the interaction. Remarkably, binding of TRBP to RPAP3 or Dicer is mutually exclusive. Additionally, we found that AGO(1/2), TRBP and Dicer are all sensitive to HSP90 inhibition, and that TRBP sensitivity is increased in the absence of RPAP3. Finally, RPAP3 seems to impede miRNA activity, raising the possibility that the R2TP chaperone might sequester TRBP to regulate the miRNA pathway.
Published: 2022

8. The box C/D snoRNP assembly factor Bcd1 interacts with the histone chaperone Rtt106 and controls its transcription dependent activity

Author: Laurence Decourty, Séverine Massenet, Stéphane Labialle, Marie-Eve Chagot, Guillaume Terral, Benoît Bragantini, Marc Quinternet, Maxime Bourguet, Cosmin Saveanu, Edouard Bertrand, Decebal Tiotiu, Christophe Charron, Sarah Cianférani, Bruno Charpentier, Xavier Manival, Jean-Marc Strub, Arnaud Paul, Steve Hessmann, Hélène Marty, Benjamin Rothé, Ingénierie Moléculaire et Physiopathologie Articulaire (IMoPA), Université de Lorraine (UL)-Centre National de la Recherche Scientifique (CNRS), Mayo Clinic [Rochester], Laboratoire de Spectrométrie de Masse BioOrganique [Strasbourg] (LSMBO), Département Sciences Analytiques et Interactions Ioniques et Biomoléculaires (DSA-IPHC), Institut Pluridisciplinaire Hubert Curien (IPHC), Institut National de Physique Nucléaire et de Physique des Particules du CNRS (IN2P3)-Université de Strasbourg (UNISTRA)-Centre National de la Recherche Scientifique (CNRS)-Institut National de Physique Nucléaire et de Physique des Particules du CNRS (IN2P3)-Université de Strasbourg (UNISTRA)-Centre National de la Recherche Scientifique (CNRS)-Institut Pluridisciplinaire Hubert Curien (IPHC), Institut National de Physique Nucléaire et de Physique des Particules du CNRS (IN2P3)-Université de Strasbourg (UNISTRA)-Centre National de la Recherche Scientifique (CNRS)-Institut National de Physique Nucléaire et de Physique des Particules du CNRS (IN2P3)-Université de Strasbourg (UNISTRA)-Centre National de la Recherche Scientifique (CNRS), Ecole Polytechnique Fédérale de Lausanne (EPFL), Génétique des Interactions macromoléculaires, Institut Pasteur [Paris]-Centre National de la Recherche Scientifique (CNRS), Institut de génétique humaine (IGH), Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS), Ingénierie, Biologie et Santé en Lorraine (IBSLor), Université de Lorraine (UL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), This work was supported by the Centre National de la Recherche Scientifique (CNRS), Université de Lorraine (UL), and Université de Strasbourg, the Agence Nationale de la Recherche [ANR-06-BLAN-0208, ANR-11-BSV8-01503, ANR-16-CE11-0032-04, ANR-08-JCJC-0019], the French Proteomic Infrastructure (ProFI, ANR-10-INBS-08-03). The authors thank GIS IBiSA, Région Alsace, Communauté Urbaine de Strasbourg, FEDER, and the IdeX program of the University of Strasbourg for financial support in purchasing HDX-MS and native MS instruments. B.B., B.R., and A.P. are fellows of the French Ministère de l’Enseignement Supérieur et de la Recherche. M.B. was supported by a fellowship from the Région Alsace. G.T. was supported by the Région Alsace and Novalix., ANR-06-BLAN-0208,RNPASSEMBLY,Nufip/Rsa1: a common assembly machine for snoRNPs, telomerase, and selenoprotein mRNPs.(2006), ANR-08-JCJC-0019,GENO-GIM,Obtention d'une carte d'interactions génétiques à l'échelle génomique chez la levure(2008), ANR-10-INBS-0008,ProFI,Infrastructure Française de Protéomique(2010), Université de Strasbourg (UNISTRA)-Université de Haute-Alsace (UHA) Mulhouse - Colmar (Université de Haute-Alsace (UHA))-Institut National de Physique Nucléaire et de Physique des Particules du CNRS (IN2P3)-Centre National de la Recherche Scientifique (CNRS)-Université de Strasbourg (UNISTRA)-Université de Haute-Alsace (UHA) Mulhouse - Colmar (Université de Haute-Alsace (UHA))-Institut National de Physique Nucléaire et de Physique des Particules du CNRS (IN2P3)-Centre National de la Recherche Scientifique (CNRS)-Institut Pluridisciplinaire Hubert Curien (IPHC), Université de Strasbourg (UNISTRA)-Université de Haute-Alsace (UHA) Mulhouse - Colmar (Université de Haute-Alsace (UHA))-Institut National de Physique Nucléaire et de Physique des Particules du CNRS (IN2P3)-Centre National de la Recherche Scientifique (CNRS)-Université de Strasbourg (UNISTRA)-Université de Haute-Alsace (UHA) Mulhouse - Colmar (Université de Haute-Alsace (UHA))-Institut National de Physique Nucléaire et de Physique des Particules du CNRS (IN2P3)-Centre National de la Recherche Scientifique (CNRS), Génétique des Interactions macromoléculaires / Genetics of Macromolecular Interactions, Institut Pasteur [Paris] (IP)-Centre National de la Recherche Scientifique (CNRS), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lorraine (UL)-Centre National de la Recherche Scientifique (CNRS), Charpentier, Bruno, Programme 'blanc' - Nufip/Rsa1: a common assembly machine for snoRNPs, telomerase, and selenoprotein mRNPs. - - RNPASSEMBLY2006 - ANR-06-BLAN-0208 - BLANC - VALID, Jeunes chercheuses et jeunes chercheurs - Obtention d'une carte d'interactions génétiques à l'échelle génomique chez la levure - - GENO-GIM2008 - ANR-08-JCJC-0019 - JCJC - VALID, Infrastructure Française de Protéomique - - ProFI2010 - ANR-10-INBS-0008 - INBS - VALID, Université de Strasbourg (UNISTRA)-Institut National de Physique Nucléaire et de Physique des Particules du CNRS (IN2P3)-Centre National de la Recherche Scientifique (CNRS)-Université de Strasbourg (UNISTRA)-Institut National de Physique Nucléaire et de Physique des Particules du CNRS (IN2P3)-Centre National de la Recherche Scientifique (CNRS)-Institut Pluridisciplinaire Hubert Curien (IPHC), and Université de Strasbourg (UNISTRA)-Institut National de Physique Nucléaire et de Physique des Particules du CNRS (IN2P3)-Centre National de la Recherche Scientifique (CNRS)-Université de Strasbourg (UNISTRA)-Institut National de Physique Nucléaire et de Physique des Particules du CNRS (IN2P3)-Centre National de la Recherche Scientifique (CNRS)
Subjects: 0301 basic medicine, Transcription, Genetic, hal-03181046, General Physics and Astronomy, RNA polymerase II, [SDV.BBM.BM] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, Crystallography, X-Ray, Histones, Transcription (biology), Ribonucleoproteins, Small Nucleolar, Multidisciplinary, biology, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], Chemistry, RNA-Binding Proteins, Chromatin, Cell biology, Histone, Box C/D snoRNP assembly, [SDV.BBM.GTP] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Genomics [q-bio.GN], RNA Polymerase II, Structural biology, Transcription, Ribosomal Proteins, Transcriptional Activation, Saccharomyces cerevisiae Proteins, [SDV.BBM.BS] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], Science, Chromatin remodelling, Saccharomyces cerevisiae, Sciences du Vivant [q-bio]/Biochimie, Biologie Moléculaire, General Biochemistry, Genetics and Molecular Biology, Article, 03 medical and health sciences, Histone H3, [SDV.BBM.GTP]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Genomics [q-bio.GN], Nuclear Magnetic Resonance, Biomolecular, R2TP complex, Cell Proliferation, 030102 biochemistry & molecular biology, Small RNAs, [SDV.BBM.BM]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, General Chemistry, Chromatin Assembly and Disassembly, 030104 developmental biology, Chaperone (protein), biology.protein, Ribosomes, Molecular Chaperones
Abstract: Biogenesis of eukaryotic box C/D small nucleolar ribonucleoproteins initiates co-transcriptionally and requires the action of the assembly machinery including the Hsp90/R2TP complex, the Rsa1p:Hit1p heterodimer and the Bcd1 protein. We present genetic interactions between the Rsa1p-encoding gene and genes involved in chromatin organization including RTT106 that codes for the H3-H4 histone chaperone Rtt106p controlling H3K56ac deposition. We show that Bcd1p binds Rtt106p and controls its transcription-dependent recruitment by reducing its association with RNA polymerase II, modulating H3K56ac levels at gene body. We reveal the 3D structures of the free and Rtt106p-bound forms of Bcd1p using nuclear magnetic resonance and X-ray crystallography. The interaction is also studied by a combination of biophysical and proteomic techniques. Bcd1p interacts with a region that is distinct from the interaction interface between the histone chaperone and histone H3. Our results are evidence for a protein interaction interface for Rtt106p that controls its transcription-associated activity., Biogenesis of small nucleolar RNAs ribonucleoproteins (snoRNPs) requires dedicated assembly machinery. Here, the authors show that a subset of snoRNP assembly factors interacts, genetically or directly, with factors modulating chromatin architecture, suggesting a link between ribosome formation and chromatin functions.
Published: 2021
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9. Optimizing the First TPR Domain of the Human SPAG1 Protein Provides Insight into the HSP70 and HSP90 Binding Properties

Author: Marie-Eve Chagot, Marc Quinternet, Sana Dermouche, Xavier Manival, Ingénierie Moléculaire et Physiopathologie Articulaire (IMoPA), Université de Lorraine (UL)-Centre National de la Recherche Scientifique (CNRS), Ingénierie, Biologie et Santé en Lorraine (IBSLor), and Université de Lorraine (UL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)
Subjects: Protein domain, Druggability, Computational biology, Biochemistry, Ribosome, Domain (software engineering), 03 medical and health sciences, Protein sequencing, Protein Domains, GTP-Binding Proteins, Ciliogenesis, [SDV.BBM.GTP]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Genomics [q-bio.GN], Humans, HSP70 Heat-Shock Proteins, HSP90 Heat-Shock Proteins, ComputingMilieux_MISCELLANEOUS, 0303 health sciences, biology, Chemistry, 030302 biochemistry & molecular biology, DNA Helicases, [SDV.BBM.BM]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, Hsp90, Tetratricopeptide, Antigens, Surface, biology.protein, ATPases Associated with Diverse Cellular Activities, Carrier Proteins, Protein Binding
Abstract: Tetratricopeptide repeat domains, or TPR domains, are protein domains that mediate protein:protein interaction. As they allow contacts between proteins, they are of particular interest in transient steps of the assembly process of macromolecular complexes, such as the ribosome or the dynein arms. In this study, we focused on the first TPR domain of the human SPAG1 protein. SPAG1 is a multidomain protein that is important for ciliogenesis whose known mutations are linked to primary ciliary dyskinesia syndrome. It can interact with the chaperones RUVBL1/2, HSP70, and HSP90. Using protein sequence optimization in combination with structural and biophysical approaches, we analyzed, with atomistic precision, how the C-terminal tails of HSPs bind a variant form of SPAG1-TPR1 that mimics the wild-type domain. We discuss our results with regard to other complex three-dimensional structures with the aim of highlighting the motifs in the TPR sequences that could drive the positioning of the HSP peptides. These data could be important for the druggability of TPR regulators.
Published: 2021
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10. Constitutive and variable 2’-O-methylation (Nm) in human ribosomal RNA

Author: Marc Quinternet, Yuri Motorin, Wassim Rhalloussi, Virginie Marchand, Ingénierie Moléculaire et Physiopathologie Articulaire (IMoPA), Université de Lorraine (UL)-Centre National de la Recherche Scientifique (CNRS), Ingénierie, Biologie et Santé en Lorraine (IBSLor), and Université de Lorraine (UL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)
Subjects: Deep sequencing, variable and constitutive, Biology, Methylation, human rRNA, 03 medical and health sciences, 0302 clinical medicine, epitranscriptome, [SDV.BBM.GTP]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Genomics [q-bio.GN], medicine, Humans, RNA Processing, Post-Transcriptional, 2ʹ-O-ribose methylation, Molecular Biology, ComputingMilieux_MISCELLANEOUS, 030304 developmental biology, Genetics, 0303 health sciences, 2'-O-methylation, Cancer, [SDV.BBM.BM]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, regulation, Cell Biology, Ribosomal RNA, medicine.disease, RNA, Ribosomal, 030220 oncology & carcinogenesis, Protein Biosynthesis, Ribosomes, Research Article, Research Paper
Abstract: Epitranscriptomic modifications of stable RNAs are dynamically regulated and specific profiles of 2ʹ-O-methylation in rRNA have been associated with distinct cancer types. However, these observations pointed out the existence of at least two distinct groups: a rather large group with constitutive rRNA Nm residues exhibiting a stable level of methylation and a more restricted set of variable modifications, giving rise to the concept of ‘specialized ribosomes’. These heterogeneous ribosomes can modulate their translational properties and be key regulatory players, depending on the physiological state of the cell. However, these conclusions were drawn from a limited set of explored human cell lines or tissues, mostly related to cancer cells of the same type. Here, we report a comprehensive analysis of human rRNA Nm modification variability observed for >15 human cell lines grown in different media and conditions. Our data demonstrate that human Nm sites can be classified into four groups, depending on their observed variability. About ⅓ of rRNA 2ʹ-O-methylations are almost invariably modified at the same level in all tested samples (stable modifications), the second group of relatively invariant modifications (another ½ of the total) showing a slightly higher variance (low variable group) and two variable groups, showing an important heterogeneity. Mapping of these four classes on the human ribosome 3D structure shows that stably modified positions are preferentially located in the important ribosome functional sites, while variable and highly variable residues are mostly distributed to the ribosome periphery. Possible relationships of such stable and variable modifications to the ribosome functions are discussed.
Published: 2021
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11. Application of NMR Spectroscopy to Determine the 3D Structure of Small Non-Coding RNAs

Author: Isabelle Lebars, Marc Quinternet, Marie-Eve Chagot, Xavier Manival, Ingénierie Moléculaire et Physiopathologie Articulaire (IMoPA), Université de Lorraine (UL)-Centre National de la Recherche Scientifique (CNRS), Ingénierie, Biologie et Santé en Lorraine (IBSLor), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lorraine (UL)-Centre National de la Recherche Scientifique (CNRS), Architecture et Réactivité de l'ARN (ARN), Institut de biologie moléculaire et cellulaire (IBMC), Université de Strasbourg (UNISTRA)-Centre National de la Recherche Scientifique (CNRS)-Université de Strasbourg (UNISTRA)-Centre National de la Recherche Scientifique (CNRS)-Centre National de la Recherche Scientifique (CNRS), Mathieu Rederstorff, univOAK, Archive ouverte, and Université de Lorraine (UL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)
Subjects: 0301 basic medicine, [SDV.BBM.BS] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], Saccharomyces cerevisiae, Computational biology, 010402 general chemistry, 3D structure, 01 natural sciences, 03 medical and health sciences, Small nucleolar RNA, Nuclear Magnetic Resonance, Biomolecular, Regulation of gene expression, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], Chemistry, RNA, RNA, Fungal, Nuclear magnetic resonance spectroscopy, dynamics, Non-coding RNA, Small molecule, NMR, 0104 chemical sciences, 030104 developmental biology, Nucleic acid, Nucleic Acid Conformation, RNA, Small Untranslated
Abstract: Many RNA architectures were discovered to be involved in a wide range of essential biological processes in all organisms from carrying genetic information to gene expression regulation. The remarkable ability of RNAs to adopt various architectures depending on their environment enables the achievement of their myriads of biological functions. Nuclear Magnetic Resonance (NMR) is a powerful technique to investigate both their structure and dynamics. NMR is also a key tool for studying interactions between RNAs and their numerous partners such as small molecules, ions, proteins, or other nucleic acids. In this chapter, to illustrate the use of NMR for 3D structure determination of small noncoding RNA, we describe detailed methods that we used for the yeast C/D box small nucleolar RNA U14 from sample preparation to 3D structure calculation.
Published: 2021
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12. NMR assignment and solution structure of the external DII domain of the yeast Rvb2 protein

Author: Bruno Charpentier, Xavier Manival, Marc Quinternet, Clément Rouillon, Benoit Bragantini, Charpentier, Bruno, Ingénierie Moléculaire et Physiopathologie Articulaire (IMoPA), Université de Lorraine (UL)-Centre National de la Recherche Scientifique (CNRS), Ingénierie, Biologie et Santé en Lorraine (IBSLor), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lorraine (UL)-Centre National de la Recherche Scientifique (CNRS), and Université de Lorraine (UL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)
Subjects: 0301 basic medicine, Saccharomyces cerevisiae Proteins, [SDV.BBM.BS] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], ATPase, Saccharomyces cerevisiae, Biochemistry, 03 medical and health sciences, Protein Domains, Structural Biology, Hydrolase, Nuclear Magnetic Resonance, Biomolecular, ComputingMilieux_MISCELLANEOUS, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], biology, Chemistry, DNA Helicases, SUPERFAMILY, Solution structure, Yeast, AAA proteins, 3. Good health, Solutions, 030104 developmental biology, Chaperone (protein), biology.protein, Biophysics
Abstract: We report the nearly complete 1H, 15N and 13C resonance assignment and the solution structure of the external DII domain of the yeast Rvb2 protein, a member of the AAA+ATPase superfamily.
Published: 2018
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13. Box C/D snoRNPs: solid-state NMR fingerprint of an early-stage 50 kDa assembly intermediate

Author: Marie-Eve Chagot, Carole Gardiennet, Marc Quinternet, Xavier Manival, Clémence Jacquemin, Ingénierie Moléculaire et Physiopathologie Articulaire (IMoPA), Université de Lorraine (UL)-Centre National de la Recherche Scientifique (CNRS), Ingénierie, Biologie et Santé en Lorraine (IBSLor), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lorraine (UL)-Centre National de la Recherche Scientifique (CNRS), Centre International de Recherche en Infectiologie (CIRI), École normale supérieure de Lyon (ENS de Lyon)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Université Jean Monnet - Saint-Étienne (UJM)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Biologie cellulaire des infections virales – Cell biology of viral infection, Université de Lyon-Université de Lyon-Université Jean Monnet - Saint-Étienne (UJM)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-École normale supérieure de Lyon (ENS de Lyon)-Université Claude Bernard Lyon 1 (UCBL), Cristallographie, Résonance Magnétique et Modélisations (CRM2), Centre National de la Recherche Scientifique (CNRS)-Université de Lorraine (UL), Université de Lorraine (UL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Centre International de Recherche en Infectiologie - UMR (CIRI), École normale supérieure - Lyon (ENS Lyon)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), and Université de Lyon-Université de Lyon-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-École normale supérieure - Lyon (ENS Lyon)-Université Claude Bernard Lyon 1 (UCBL)
Subjects: Snu13p, box C/D snoRNP, Saccharomyces cerevisiae Proteins, 030303 biophysics, Saccharomyces cerevisiae, Ribosome biogenesis, Context (language use), Biochemistry, Protein Structure, Secondary, Protein–protein interaction, protein-protein interaction, 03 medical and health sciences, Structural Biology, Ribonucleoproteins, Small Nucleolar, [SDV.BBM.GTP]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Genomics [q-bio.GN], Guide RNA, Small nucleolar RNA, Carbon-13 Magnetic Resonance Spectroscopy, Protein secondary structure, Nuclear Magnetic Resonance, Biomolecular, 030304 developmental biology, 0303 health sciences, biology, RNA, [SDV.BBM.BM]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, biology.organism_classification, Molecular Weight, Biophysics, solid-state NMR
Abstract: International audience; Many cellular functions rely on stable protein-only or protein-RNA complexes. Deciphering their assembly mechanism is a key question in cell biology. We here focus on box C/D small nucleolar ribonucleoproteins involved in ribosome biogenesis. The mature particles contain four core proteins and a guide RNA. Despite their relatively simple composition, these particles don't self-assemble in eukaryote and the production of a native and functional particle requires a large number of transient other proteins, called assembly factors. We present here 13C and 15N solid-state NMR assignment of yeast 126-residue core protein Snu13 in the context of its 50 kDa pre-complex with assembly factors Rsa1p:Hit1p. In this sample, only one third of the protein is labelled, leading to a low sensitivity. We could nevertheless obtain assignment data for 91% of the residues. Secondary structure derived from our assignments shows that Snu13p overall structure is maintained in the context of the complex. Chemical shift perturbations are analysed to evaluate Snu13p conformational changes and interaction interface upon binding to its partner proteins. While indirect perturbations are observed in the hydrophobic core, we find other good candidate residues belonging to the interaction interface. We describe the role of some Snu13p N-terminal and C-terminal residues, not identified in previous structural studies. These preliminary results will serve as a basis for future interaction studies, especially by adding RNA, to decipher box C/D snoRNP particles assembly pathway.
Published: 2019
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14. Binding properties of the quaternary assembly protein SPAG1

Author: Marc Quinternet, Marie-Eve Chagot, Christophe Chipot, Dorian Lefebvre, François Dehez, Raphael Dos Santos Morais, Sana Dermouche, Xavier Manival, Ingénierie Moléculaire et Physiopathologie Articulaire (IMoPA), Université de Lorraine (UL)-Centre National de la Recherche Scientifique (CNRS), Ingénierie, Biologie et Santé en Lorraine (IBSLor), Université de Lorraine (UL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Laboratoire de Physique et Chimie Théoriques (LPCT), Institut de Chimie du CNRS (INC)-Université de Lorraine (UL)-Centre National de la Recherche Scientifique (CNRS), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lorraine (UL)-Centre National de la Recherche Scientifique (CNRS), and GONNET, JULIE
Subjects: Spliceosome, GTPase, Molecular Dynamics Simulation, [SDV.BBM.BM] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, Biochemistry, Ribosome, 03 medical and health sciences, GTP-Binding Proteins, Heat shock protein, [SDV.BBM.GTP]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Genomics [q-bio.GN], Humans, HSP70 Heat-Shock Proteins, Protein Interaction Domains and Motifs, HSP90 Heat-Shock Proteins, Molecular Biology, Nuclear Magnetic Resonance, Biomolecular, ComputingMilieux_MISCELLANEOUS, 030304 developmental biology, TRPC Cation Channels, 0303 health sciences, biology, Chemistry, 030302 biochemistry & molecular biology, [SDV.BBM.BM]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, Cell Biology, Hsp90, Recombinant Proteins, Molecular Docking Simulation, Proteasome, Docking (molecular), Chaperone (protein), Antigens, Surface, biology.protein, Biophysics, [SDV.BBM.GTP] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Genomics [q-bio.GN], Guanosine Triphosphate, Apoptosis Regulatory Proteins, Carrier Proteins, Protein Binding
Abstract: In cells, many constituents are able to assemble resulting in large macromolecular machineries possessing very specific biological and physiological functions, e.g. ribosome, spliceosome and proteasome. Assembly of such entities is commonly mediated by transient protein factors. SPAG1 is a multidomain protein, known to participate in the assembly of both the inner and outer dynein arms. These arms are required for the function of sensitive and motile cells. Together with RUVBL1, RUVBL2 and PIH1D2, SPAG1 is a key element of R2SP, a protein complex assisting the quaternary assembly of specific protein clients in a tissue-specific manner and associating with heat shock proteins (HSPs) and regulators. In this study, we have investigated the role of TPR domains of SPAG1 in the recruitment of HSP chaperones by combining biochemical assays, ITC, NMR spectroscopy and molecular dynamics (MD) simulations. First, we propose that only two, out of the three TPR domains, are able to recruit the protein chaperones HSP70 and HSP90. We then focused on one of these TPR domains and elucidated its 3D structure using NMR spectroscopy. Relying on an NMR-driven docking approach and MD simulations, we deciphered its binding interface with the C-terminal tails of both HSP70 and HSP90. Finally, we addressed the biological function of SPAG1 and specifically demonstrated that a SPAG1 sub-fragment, containing a putative P-loop motif, cannot efficiently bind and hydrolyze GTP in vitro. Our data challenge the interpretation of SPAG1 possessing GTPase activity. We propose instead that SPAG1 regulates nucleotide hydrolysis activity of the HSP and RUVBL1/2 partners.
Published: 2019
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15. The yeast C/D box snoRNA U14 adopts a 'weak' K-turn like conformation recognized by the Snu13 core protein in solution

Author: Bruno Charpentier, Xavier Manival, Marie-Eve Chagot, Jérôme Coutant, Isabelle Lebars, Marc Quinternet, Benjamin Rothé, Ingénierie Moléculaire et Physiopathologie Articulaire (IMoPA), Université de Lorraine (UL)-Centre National de la Recherche Scientifique (CNRS), Ingénierie, Biologie et Santé en Lorraine (IBSLor), Université de Lorraine (UL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Bruker, Architecture et Réactivité de l'ARN (ARN), Institut de biologie moléculaire et cellulaire (IBMC), Université de Strasbourg (UNISTRA)-Centre National de la Recherche Scientifique (CNRS)-Université de Strasbourg (UNISTRA)-Centre National de la Recherche Scientifique (CNRS)-Centre National de la Recherche Scientifique (CNRS), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lorraine (UL)-Centre National de la Recherche Scientifique (CNRS), and GONNET, JULIE
Subjects: Ribosomal Proteins, 0301 basic medicine, RNA Folding, Saccharomyces cerevisiae Proteins, base-pairs, Base pair, Stereochemistry, RNase P, small nucleolar rna, b/c motif, Saccharomyces cerevisiae, [SDV.BBM.BM] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, Biochemistry, Ribosome, nmr, 03 medical and health sciences, dipolar couplings, snorna u14, [SDV.BBM.GTP]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Genomics [q-bio.GN], nmr-spectroscopy, RNA, Small Nucleolar, Small nucleolar RNA, Structural motif, ComputingMilieux_MISCELLANEOUS, Ribonucleoprotein, 15.5k protein, Binding Sites, 030102 biochemistry & molecular biology, Chemistry, RNA, [SDV.BBM.BM]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, General Medicine, Ribonucleoproteins, Small Nuclear, pre-ribosomal-rna, 030104 developmental biology, messenger-rna, k-turn, snu13 protein, Nucleic Acid Conformation, [SDV.BBM.GTP] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Genomics [q-bio.GN], triple-resonance experiment, Sequence motif, Protein Binding, structural features
Abstract: Non-coding RNAs associate with proteins to form ribonucleoproteins (RNPs), such as ribosome, box C/D snoRNPs, H/ACA snoRNPs, ribonuclease P, telomerase and spliceosome to ensure cell viability. The assembly of these RNA-protein complexes relies on the ability of the RNA to adopt the correct bound conformation. K-turn motifs represent ubiquitous binding platform for proteins found in several cellular environment. This structural motif has an internal three-nucleotide bulge flanked on its 3' side by a G.A/A.G tandem pairs followed by one or two non-Watson-Crick pairs, and on its 5' side by a classical RNA helix. This peculiar arrangement induces a strong curvature of the phosphodiester backbone, which makes it conducive to multiple tertiary interactions. SNU13/Snu13p (Human/Yeast) binds specifically the U14 C/D box snoRNA K-turn sequence motif. This event is the prerequisite to promote the assembly of the RNP, which contains NOP58/Nop58 and NOP56/Nop56 core proteins and the 2'-O-methyl-transferase, Fibrillarin/Noplp. The U14 small nucleolar RNA is a conserved non-coding RNA found in yeast and vertebrates required for the pre-rRNA maturation and ribose methylation. Here, we report the solution structure of the free U14 snoRNA K-turn motif (kt-U14) as determined by Nuclear Magnetic Resonance. We demonstrate that a major fraction of free kt-U14 adopts a pre-folded conformation similar to protein bound K-turn, even in the absence of divalent ions. In contrast to the kt-U4 or tyrS RNA, kt-U14 displays a sharp bent in the phosphodiester backbone. The U.U and G.A tandem base pairs are formed through weak hydrogen bonds. Finally, we show that the structure of kt-U14 is stabilized upon Snu13p binding. The structure of the free U14 RNA is the first reference example for the canonical motifs of the C/D box snoRNA family. (C) 2019 Elsevier B.V. and Societe Francaise de Biochimie et Biologie Moleculaire (SFBBM). All rights reserved.
Published: 2019
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16. A structure-function study shows a possible role for the R2TP complex in RISC assembly and/or function

Author: Yoann Abel, Valerie Bourguignon-Igel, Christophe Charron, Marc Quinternet, Christiane Branlant, Edouard Bertrand, Céline Verheggen, Xavier Manival, Bruno Charpentier, Mathieu Rederstorff, Ingénierie Moléculaire et Physiopathologie Articulaire (IMoPA), Université de Lorraine (UL)-Centre National de la Recherche Scientifique (CNRS), and Rederstorff, Mathieu
Subjects: [SDV] Life Sciences [q-bio], [SDV]Life Sciences [q-bio], [SDV.BBM] Life Sciences [q-bio]/Biochemistry, Molecular Biology, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, ComputingMilieux_MISCELLANEOUS
Abstract: International audience
Published: 2018

17. CRD SAT generated by pCARGHO: A new efficient lectin-based affinity tag method for safe, simple and low-cost protein purification

Author: Hélène Le Cordier, Guillaume Groshenry, François Talfournier, Alexandre Kriznik, Sandrine Boschi-Muller, Jean-Christophe Lec, Pascal Reboul, Jean-Yves Jouzeau, Marc Quinternet, Christophe Charron, Melissa Yelehe-Okouma, Hortense Mazon, Ingénierie Moléculaire et Physiopathologie Articulaire (IMoPA), Université de Lorraine (UL)-Centre National de la Recherche Scientifique (CNRS), Département de Pharmacologie Clinique et Toxicologie, Centre Hospitalier Régional Universitaire, Nancy, France, Ingénierie, Biologie et Santé en Lorraine (IBSLor), Université de Lorraine (UL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Service de Pharmacologie Clinique et Toxicologie [CHRU Nancy], Centre Hospitalier Régional Universitaire de Nancy (CHRU Nancy), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lorraine (UL)-Centre National de la Recherche Scientifique (CNRS), and Reboul, Pascal
Subjects: 0301 basic medicine, [SDV.BIO]Life Sciences [q-bio]/Biotechnology, Protein purification based on new tag technology, medicine.disease_cause, Affinity chromatography, Applied Microbiology and Biotechnology, 03 medical and health sciences, Protein purification, [SDV.BBM] Life Sciences [q-bio]/Biochemistry, Molecular Biology, TEV protease, medicine, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, Escherichia coli, galectin-3-derived CRD-tag, Expression vector, Downstream processing, 030102 biochemistry & molecular biology, Chemistry, General Medicine, Fusion protein, [SDV.BIO] Life Sciences [q-bio]/Biotechnology, 030104 developmental biology, Isoelectric point, Biochemistry, Molecular Medicine
Abstract: International audience; Purification of recombinant proteins remains a bottleneck for downstream processing. We engineered a new galectin 3 truncated form (CRDSAT ), functionally and structurally characterized, with preserved solubility and lectinic activity. Taking advantage of these properties, we designed an expression vector (pCARGHO), suitable for CRDSAT -tagged protein expression in prokaryotes. CRDSAT binds to lactose-Sepharose with a high specificity and facilitates solubilization of fusion proteins. This tag is structurally stable and can be easily removed from fusion proteins using TEV protease. Furthermore, due to their basic isoelectric point (pI), CRDSAT and TEV are efficiently eliminated using cationic exchange chromatography. When pI of the protein of interest and CRDSAT were close, other chromatographic methods were successfully tested. Using CRDSAT tag, we purified several proteins from prokaryote and eukaryote origin and demonstrated as examples, the preservation of both Escherichia coli Thioredoxin 1 and human CDC25Bcd activities. Overall, yields of proteins obtained after tag removal were about 5 to 50 mg per litre of bacterial culture. Our purification method displays various advantages described herein that may greatly interest academic laboratories, biotechnology and pharmaceutical companies.
Published: 2018
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18. Deep Structural Analysis of RPAP3 and PIH1D1, Two Components of the HSP90 Co-chaperone R2TP Complex

Author: Marie-Eve Chagot, Marc Quinternet, Christelle Aigueperse, Guillaume Terral, Céline Verheggen, Rénette Saint-Fort, Chloé Maurizy, Franck Vandermoere, Florian Georgescauld, Maxime Bourguet, Julien Henri, Bruno Charpentier, Xavier Manival, Yoann Abel, Bérengère Pradet-Balade, Philippe Meyer, Sarah Cianférani, Edouard Bertrand, Isabelle Behm-Ansmant, Laboratoire de Biologie Moléculaire et Cellulaire des Eucaryotes (LBMCE), Institut de biologie physico-chimique (IBPC (FR_550)), Centre National de la Recherche Scientifique (CNRS)-Sorbonne Université (SU)-Centre National de la Recherche Scientifique (CNRS)-Sorbonne Université (SU)-Centre National de la Recherche Scientifique (CNRS), Ingénierie Moléculaire et Physiopathologie Articulaire (IMoPA), Université de Lorraine (UL)-Centre National de la Recherche Scientifique (CNRS), Laboratoire de Spectrométrie de Masse BioOrganique [Strasbourg] (LSMBO), Département Sciences Analytiques et Interactions Ioniques et Biomoléculaires (DSA-IPHC), Institut Pluridisciplinaire Hubert Curien (IPHC), Université de Strasbourg (UNISTRA)-Institut National de Physique Nucléaire et de Physique des Particules du CNRS (IN2P3)-Centre National de la Recherche Scientifique (CNRS)-Université de Strasbourg (UNISTRA)-Institut National de Physique Nucléaire et de Physique des Particules du CNRS (IN2P3)-Centre National de la Recherche Scientifique (CNRS)-Institut Pluridisciplinaire Hubert Curien (IPHC), Université de Strasbourg (UNISTRA)-Institut National de Physique Nucléaire et de Physique des Particules du CNRS (IN2P3)-Centre National de la Recherche Scientifique (CNRS)-Université de Strasbourg (UNISTRA)-Institut National de Physique Nucléaire et de Physique des Particules du CNRS (IN2P3)-Centre National de la Recherche Scientifique (CNRS), Institut de Génétique Moléculaire de Montpellier (IGMM), Centre National de la Recherche Scientifique (CNRS)-Université de Montpellier (UM), Institut de Génomique Fonctionnelle (IGF), Université de Montpellier (UM)-Université Montpellier 1 (UM1)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Montpellier 2 - Sciences et Techniques (UM2)-Centre National de la Recherche Scientifique (CNRS), Centre de recherche en Biologie Cellulaire (CRBM), Université Montpellier 2 - Sciences et Techniques (UM2)-Centre National de la Recherche Scientifique (CNRS)-Université de Montpellier (UM)-Université Montpellier 1 (UM1), Ingénierie, Biologie et Santé en Lorraine (IBSLor), Université de Lorraine (UL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Centre National de la Recherche Scientifique (CNRS)-Centre National de la Recherche Scientifique (CNRS)-Sorbonne Université (SU)-Centre National de la Recherche Scientifique (CNRS), Institut National de Physique Nucléaire et de Physique des Particules du CNRS (IN2P3)-Université de Strasbourg (UNISTRA)-Centre National de la Recherche Scientifique (CNRS)-Institut National de Physique Nucléaire et de Physique des Particules du CNRS (IN2P3)-Université de Strasbourg (UNISTRA)-Centre National de la Recherche Scientifique (CNRS)-Institut Pluridisciplinaire Hubert Curien (IPHC), Institut National de Physique Nucléaire et de Physique des Particules du CNRS (IN2P3)-Université de Strasbourg (UNISTRA)-Centre National de la Recherche Scientifique (CNRS)-Institut National de Physique Nucléaire et de Physique des Particules du CNRS (IN2P3)-Université de Strasbourg (UNISTRA)-Centre National de la Recherche Scientifique (CNRS), Centre de résonance magnétique biologique et médicale (CRMBM), and Aix Marseille Université (AMU)-Assistance Publique - Hôpitaux de Marseille (APHM)-Centre National de la Recherche Scientifique (CNRS)
Subjects: Models, Molecular, 0301 basic medicine, Protein Conformation, [SDV]Life Sciences [q-bio], Protein domain, HSP72 Heat-Shock Proteins, Computational biology, Plasma protein binding, RUVBL, Cell Line, 03 medical and health sciences, Protein structure, Protein Domains, Structural Biology, Chaperones, Humans, Protein Isoforms, Machinery assembly, HSP90, PIH1D1, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, HSP90 Heat-Shock Proteins, Binding site, Molecular Biology, R2TP, ComputingMilieux_MISCELLANEOUS, HSP70, R2TP complex, Binding Sites, biology, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], Chemistry, RPAP3, Molecular machine, Co-chaperone, 030104 developmental biology, Chaperone (protein), biology.protein, Protein Multimerization, Apoptosis Regulatory Proteins, Carrier Proteins, Protein Binding
Abstract: RPAP3 and PIH1D1 are part of the HSP90 co-chaperone R2TP complex involved in the assembly process of many molecular machines. In this study, we performed a deep structural investigation of the HSP binding abilities of the two TPR domains of RPAP3. We combined 3D NMR, non-denaturing MS, and ITC techniques with Y2H, IP-LUMIER, FRET, and ATPase activity assays and explain the fundamental role played by the second TPR domain of RPAP3 in the specific recruitment of HSP90. We also established the 3D structure of an RPAP3:PIH1D1 sub-complex demonstrating the need for a 34-residue insertion, specific of RPAP3 isoform 1, for the tight binding of PIH1D1. We also confirm the existence of a complex lacking PIH1D1 in human cells (R2T), which shows differential binding to certain clients. These results highlight similarities and differences between the yeast and human R2TP complexes, and document the diversification of this family of co-chaperone complexes in human.
Published: 2018
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19. CRD

Author: Alexandre, Kriznik, Mélissa, Yéléhé-Okouma, Jean-Christophe, Lec, Guillaume, Groshenry, Hélène, Le Cordier, Christophe, Charron, Marc, Quinternet, Hortense, Mazon, François, Talfournier, Sandrine, Boschi-Muller, Jean-Yves, Jouzeau, and Pascal, Reboul
Subjects: Thioredoxins, Gene Expression Regulation, Solubility, Galectin 3, Lectins, Endopeptidases, Genetic Vectors, Escherichia coli, Humans, cdc25 Phosphatases, Chromatography, Ion Exchange, Recombinant Proteins
Abstract: Purification of recombinant proteins remains a bottleneck for downstream processing. The authors engineered a new galectin 3 truncated form (CRD
Published: 2018

20. Structural Features of the Box C/D snoRNP Pre-assembly Process Are Conserved through Species

Author: Marie-Eve Chagot, Decebal Tiotiu, Bruno Charpentier, Marc Quinternet, Xavier Manival, Benjamin Rothé, Bioingénierie Moléculaire, Cellulaire et Thérapeutique (BMCT), Université de Lorraine (UL)-Centre National de la Recherche Scientifique (CNRS), Ingénierie Moléculaire et Physiopathologie Articulaire (IMoPA), and Centre National de la Recherche Scientifique (CNRS)-Université de Lorraine (UL)
Subjects: Models, Molecular, 0301 basic medicine, Magnetic Resonance Spectroscopy, Saccharomyces cerevisiae Proteins, Protein Conformation, Saccharomyces cerevisiae, RNA-binding protein, Context (language use), Plasma protein binding, Biology, Conserved sequence, Evolution, Molecular, 03 medical and health sciences, Protein structure, Ribonucleoproteins, Small Nucleolar, Structural Biology, Humans, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, Nuclear protein, Small nucleolar RNA, Molecular Biology, Conserved Sequence, ComputingMilieux_MISCELLANEOUS, Genetics, Nuclear Proteins, RNA-Binding Proteins, Ribonucleoproteins, Small Nuclear, biology.organism_classification, Cell biology, 030104 developmental biology, Protein Multimerization, Protein Binding, Transcription Factors
Abstract: Box C/D small nucleolar ribonucleoparticles (snoRNPs) support 2'-O-methylation of several target RNAs. They share a common set of four core proteins (SNU13, NOP58, NOP56, and FBL) that are assembled on different guide small nucleolar RNAs. Assembly of these entities involves additional protein factors that are absent in the mature active particle. In this context, the platform protein NUFIP1/Rsa1 establishes direct and simultaneous contacts with core proteins and with the components of the assembly machinery. Here, we solve the nuclear magnetic resonance (NMR) structure of a complex resulting from interaction between protein fragments of human NUFIP1 and its cofactor ZNHIT3, and emphasize their imbrication. Using yeast two-hybrid and complementation assays, protein co-expression, isothermal titration calorimetry, and NMR, we demonstrate that yeast and human complexes involving NUFIP1/Rsa1p, ZNHIT3/Hit1p, and SNU13/Snu13p share strong structural similarities, suggesting that the initial steps of the box C/D snoRNP assembly process are conserved among species.
Published: 2016
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21. Functional and Structural Insights of the Zinc-Finger HIT protein family members Involved in Box C/D snoRNP Biogenesis

Author: Sarah Cianférani, Jean-Michel Saliou, Hélène Marty, Bruno Charpentier, Decebal Tiotiu, Xavier Manival, Marc Quinternet, Benjamin Rothé, Benoit Bragantini, Ingénierie Moléculaire et Physiopathologie Articulaire (IMoPA), Université de Lorraine (UL)-Centre National de la Recherche Scientifique (CNRS), Laboratoire de Spectrométrie de Masse BioOrganique [Strasbourg] (LSMBO), Département Sciences Analytiques et Interactions Ioniques et Biomoléculaires (DSA-IPHC), Institut Pluridisciplinaire Hubert Curien (IPHC), Université de Strasbourg (UNISTRA)-Institut National de Physique Nucléaire et de Physique des Particules du CNRS (IN2P3)-Centre National de la Recherche Scientifique (CNRS)-Université de Strasbourg (UNISTRA)-Institut National de Physique Nucléaire et de Physique des Particules du CNRS (IN2P3)-Centre National de la Recherche Scientifique (CNRS)-Institut Pluridisciplinaire Hubert Curien (IPHC), Université de Strasbourg (UNISTRA)-Institut National de Physique Nucléaire et de Physique des Particules du CNRS (IN2P3)-Centre National de la Recherche Scientifique (CNRS)-Université de Strasbourg (UNISTRA)-Institut National de Physique Nucléaire et de Physique des Particules du CNRS (IN2P3)-Centre National de la Recherche Scientifique (CNRS), Université de Strasbourg (UNISTRA)-Institut National de Physique Nucléaire et de Physique des Particules du CNRS (IN2P3)-Centre National de la Recherche Scientifique (CNRS), Institut Pasteur de Lille, Réseau International des Instituts Pasteur (RIIP), and Bioingénierie Moléculaire, Cellulaire et Thérapeutique (BMCT)
Subjects: Models, Molecular, 0301 basic medicine, Protein Folding, Hot Temperature, Magnetic Resonance Spectroscopy, Saccharomyces cerevisiae Proteins, Protein family, Protein Conformation, Saccharomyces cerevisiae, Biology, Chromatin remodeling, 03 medical and health sciences, Protein structure, Ribonucleoproteins, Small Nucleolar, Stress, Physiological, Structural Biology, Kruppel-Like Factor 6, Small nucleolar RNA, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], Molecular Biology, Zinc finger, Genetics, 030102 biochemistry & molecular biology, Protein Stability, Zinc Fingers, biology.organism_classification, Cell biology, 030104 developmental biology, Protein folding, [SDV.MHEP]Life Sciences [q-bio]/Human health and pathology
Abstract: International audience; Zf-HIT family members share the zf-HIT domain (ZHD), which is characterized by a fold in ``treble-clef' through interleaved CCCC and CCHC ZnF motifs that both bind a zinc atom. Six proteins containing ZHD are present in human and three in yeast proteome, all belonging to multimodular RNA/protein complexes involved in gene regulation, chromatin remodeling, and snoRNP assembly. An interesting characteristic of the cellular complexes that ensure these functions is the presence of the RuvBL1/2/Rvb1/2 ATPases closely linked with zf-HIT proteins. Human ZNHIT6/BCD1 and its counterpart in yeast Bcd1p were previously characterized as assembly factors of the box C/D snoRNPs. Our data reveal that the ZHD of Bcd1p is necessary but not sufficient for yeast growth and that the motif has no direct RNA-binding capacity but helps Bcd1p maintain the box C/D snoRNAs level in steady state. However, we demonstrated that Bcd1p interacts nonspecifically with RNAs depending on their length. Interestingly, the ZHD of Bcd1p is functionally interchangeable with that of Hit1p, another box C/D snoRNP assembly factor belonging to the zf-HIT family. This prompted us to use NMR to solve the 3D structures of ZHD from yeast Bcd1p and Hit1p to highlight the structural similarity in the zf-HIT family. We identified structural features associated with the requirement of Hit1p and Bcd1p ZHD for cell growth and box C/D snoRNA stability under heat stress. Altogether, our data suggest an important role of ZHD could be to maintain functional folding to the rest of the protein, especially under heat stress conditions. (C) 2016 Elsevier Ltd. All rights reserved.
Published: 2016
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22. Vinexinβ, an atypical 'sensor' of retinoic acid receptor γ signaling: union and sequestration, separation, and phosphorylation

Author: Marc Vitorino, Gaétan Bour, Pascal Kessler, Marc Quinternet, Nathalie Bruck, Marc-André Delsuc, Sébastien Lalevée, Cécile Rochette-Egly, Bruno Kieffer, Jean-Luc Vonesch, and Eric Samarut
Subjects: Receptors, Retinoic Acid, Molecular Sequence Data, Retinoic acid, Biology, Biochemistry, SH3 domain, Mice, chemistry.chemical_compound, Cell Line, Tumor, Chlorocebus aethiops, Genetics, Animals, Amino Acid Sequence, Phosphorylation, Promoter Regions, Genetic, Molecular Biology, Adaptor Proteins, Signal Transducing, Promoter, Molecular biology, Recombinant Proteins, Cell biology, Retinoic acid receptor, chemistry, Nuclear receptor, COS Cells, Mutation, Corepressor, Chromatin immunoprecipitation, Protein Binding, Signal Transduction, Biotechnology
Abstract: The transcriptional activity of nuclear retinoic acid receptors (RARs) relies on the association/dissociation of coregulators at the ligand-binding domain. However, we determined that the N-terminal domain (NTD) also plays a role through its phosphorylation, and we isolated vinexinβ, a cytoskeleton protein with three SH3 domains, as a new partner of the RARγ NTD. Here we deciphered the mechanism of the interaction and its role in RARγ-mediated transcription. By combining molecular and biophysical (surface plasmon resonance, NMR, and fluorescence resonance energy transfer) approaches, we demonstrated that the third SH3 domain of vinexinβ interacts with a proline-rich domain (PRD) located in RARγ NTD and that phosphorylation at a serine located in the PRD abrogates the interaction. The affinity of the interaction was also evaluated. In vivo, vinexinβ represses RARγ-mediated transcription and we dissected the underlying mechanism in chromatin immunoprecipitation experiments performed with F9 cells expressing RARγ wild type or mutated at the phosphorylation site. In the absence of retinoic acid (RA), vinexinβ does not occupy RARγ target gene promoters and sequesters nonphosphorylated RARγ out of promoters. In response to RA, RARγ becomes phosphorylated and dissociates from vinexinβ. This separation allows RARγ to occupy promoters. This is the first report of an RAR corepressor association/dissociation out of promoters and regulated by phosphorylation.
Published: 2010
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23. Solution Structure and Backbone Dynamics of the Cysteine 103 to Serine Mutant of the N-Terminal Domain of DsbD from Neisseria meningitides

Author: Marc Quinternet, Manh-Thong Cung, Laure Selme, Marie-Christine Averlant-Petit, Pascale Tsan, Christophe Jacob, Guy Branlant, Chrystel Beaufils, Sandrine Boschi-Muller, Laboratoire de Chimie Physique Macromoléculaire (LCPM), Institut de Chimie du CNRS (INC)-Université de Lorraine (UL)-Centre National de la Recherche Scientifique (CNRS), Maturation des ARN et enzymologie moléculaire (MAEM), and Cancéropôle du Grand Est-Université Henri Poincaré - Nancy 1 (UHP)-IFR111-Centre National de la Recherche Scientifique (CNRS)
Subjects: Models, Molecular, Meningitides, Magnetic Resonance Spectroscopy, MESH: Mutation, Stereochemistry, MESH: Catalytic Domain, Neisseria meningitidis, MESH: Solutions, 010402 general chemistry, 01 natural sciences, Biochemistry, MESH: Neisseria meningitidis, Serine, MESH: Mutant Proteins, MESH: Protein Structure, Tertiary, 03 medical and health sciences, Oxidoreductase, Catalytic Domain, Cysteine, MESH: Oxidoreductases, MESH: Serine, 030304 developmental biology, chemistry.chemical_classification, 0303 health sciences, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], biology, MESH: Magnetic Resonance Spectroscopy, Active site, Periplasmic space, MESH: Cysteine, Transmembrane protein, Protein Structure, Tertiary, 0104 chemical sciences, Solutions, chemistry, Mutation, biology.protein, Mutant Proteins, Thioredoxin, Oxidoreductases, MESH: Models, Molecular
Abstract: International audience; The DsbD protein is essential for electron transfer from the cytoplasm to the periplasm of Gram-negative bacteria. Its N-terminal domain dispatches electrons coming from cytoplasmic thioredoxin (Trx), via its central transmembrane and C-terminal domains, to its periplasmic partners: DsbC, DsbE/CcmG, and DsbG. Previous structural studies described the latter proteins as Trx-like folds possessing a characteristic C-X-X-C motif able to generate a disulfide bond upon oxidation. The Escherichia coli nDsbD displays an immunoglobulin-like fold in which two cysteine residues (Cys103 and Cys109) allow a disulfide bond exchange with its biological partners.We have determined the structure in solution and the backbone dynamics of the C103S mutant of the N-terminal domain of DsbD from Neisseria meningitidis. Our results highlight significant structural changes concerning the beta-sheets and the local topology of the active site compared with the oxidized form of the E. coli nDsbD. The structure reveals a "cap loop" covering the active site, similar to the oxidized E. coli nDsbD X-ray structure. However, regions featuring enhanced mobility were observed both near to and distant from the active site, revealing a capacity of structural adjustments in the active site and in putative interaction areas with nDsbD biological partners. Results are discussed in terms of functional consequences.
Published: 2008
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24. Structure/Function Analysis of Protein-Protein Interactions Developed by the Yeast Pih1 Platform Protein and Its Partners in Box C/D snoRNP Assembly

Author: Marc Quinternet, Benjamin Rothé, Muriel Barbier, Clémence Jacquemin, Marie-Eve Chagot, Régis Back, Claude Bobo, Jean-Michel Saliou, Bruno Charpentier, Christiane Branlant, Xavier Manival, Philippe Meyer, Sarah Cianférani, Bioingénierie Moléculaire, Cellulaire et Thérapeutique (BMCT), Centre National de la Recherche Scientifique (CNRS)-Université de Lorraine (UL), Ingénierie Moléculaire et Physiopathologie Articulaire (IMoPA), Université de Lorraine (UL)-Centre National de la Recherche Scientifique (CNRS), Département Sciences Analytiques et Interactions Ioniques et Biomoléculaires (DSA-IPHC), Institut Pluridisciplinaire Hubert Curien (IPHC), Université de Strasbourg (UNISTRA)-Institut National de Physique Nucléaire et de Physique des Particules du CNRS (IN2P3)-Centre National de la Recherche Scientifique (CNRS)-Université de Strasbourg (UNISTRA)-Institut National de Physique Nucléaire et de Physique des Particules du CNRS (IN2P3)-Centre National de la Recherche Scientifique (CNRS), Laboratoire de Biologie Moléculaire et Cellulaire des Eucaryotes (LBMCE), Université Pierre et Marie Curie - Paris 6 (UPMC)-Centre National de la Recherche Scientifique (CNRS), and Université de Strasbourg (UNISTRA)-Université de Haute-Alsace (UHA) Mulhouse - Colmar (Université de Haute-Alsace (UHA))-Institut National de Physique Nucléaire et de Physique des Particules du CNRS (IN2P3)-Centre National de la Recherche Scientifique (CNRS)-Université de Strasbourg (UNISTRA)-Université de Haute-Alsace (UHA) Mulhouse - Colmar (Université de Haute-Alsace (UHA))-Institut National de Physique Nucléaire et de Physique des Particules du CNRS (IN2P3)-Centre National de la Recherche Scientifique (CNRS)
Subjects: Ribosomal Proteins, Protein Folding, Saccharomyces cerevisiae Proteins, Saccharomyces cerevisiae, Methylation, Ribosome assembly, Protein–protein interaction, Structural Biology, Ribonucleoproteins, Small Nucleolar, Escherichia coli, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, Guide RNA, HSP90 Heat-Shock Proteins, Protein Interaction Maps, Molecular Biology, Nuclear Magnetic Resonance, Biomolecular, R2TP complex, biology, RNA, Nuclear Proteins, Ribonucleoproteins, Small Nuclear, Protein Structure, Tertiary, Biochemistry, RNA, Ribosomal, Chaperone (protein), Box C/D snoRNP assembly, biology.protein, Biophysics, Protein folding, Hydrophobic and Hydrophilic Interactions, [SDV.MHEP]Life Sciences [q-bio]/Human health and pathology, Molecular Chaperones, Protein Binding
Abstract: International audience; In eukaryotes, nucleotide post-transcriptional modifications in RNAs play an essential role in cell proliferation by contributing to pre-ribosomal RNA processing, ribosome assembly and activity. Box C/D small nucleolar ribonucleoparticles catalyze site-specific 2'-O-methylation of riboses, one of the most prevalent RNA modifications. They contain one guide RNA and four core proteins and their in vivo assembly requires numerous factors including (HUMAN/Yeast) BCD1/Bcd1p, NUFIP1/Rsa1p, ZNHIT3/Hit1p, the R2TP complex composed of protein PIH1D1/Pih1p and RPAP3/Tah1p that bridges the R2TP complex to the HSP90/Hsp82 chaperone and two AAA+ATPases. We show that Tah1p can stabilize Pih1p in the absence of Hsp82 activity during the stationary phase of growth and consequently that the Tah1p:Pih1p interaction is sufficient for Pih1p stability. This prompted us to establish the solution structure of the Tah1p:Pih1p complex by NMR. The C-terminal tail S-93-S-111 of Tah1p snakes along Pih1p(264-344) folded in a CS domain to form two intermolecular beta-sheets and one covering loop. However, a thorough inspection of the NMR and crystal structures revealed structural differences that may be of functional importance. In addition, our NMR and isothermal titration calorimetry data revealed the formation of direct contacts between Pih1p(257-344) and the H5p82MC domain in the presence of Tah1p. By co-expression in Escherichia coli, we demonstrate that Pih1p has two other direct partners, the Rsa1p assembly factor and the Nop58p core protein, and in vivo and in vitro experiments mapped the required binding domains. Our data suggest that these two interactions are mutually exclusive. The implication of this finding for box C/D small nucleolar ribonucleoparticle assembly is discussed. (C) 2015 Elsevier Ltd. All rights reserved.
Published: 2015
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25. Accurate protein-peptide titration experiments by nuclear magnetic resonance using low-volume samples

Author: Christian, Köhler, Raphaël, Recht, Marc, Quinternet, Frederic, de Lamotte, Marc-André, Delsuc, and Bruno, Kieffer
Subjects: Models, Molecular, Magnetic Resonance Spectroscopy, Receptors, Retinoic Acid, Molecular Sequence Data, Humans, Muscle Proteins, Proteins, Amino Acid Sequence, Phosphorylation, Peptides, Adaptor Proteins, Signal Transducing, Protein Structure, Tertiary
Abstract: NMR spectroscopy allows measurements of very accurate values of equilibrium dissociation constants using chemical shift perturbation methods, provided that the concentrations of the binding partners are known with high precision and accuracy. The accuracy and precision of these experiments are improved if performed using individual capillary tubes, a method enabling full automation of the measurement. We provide here a protocol to set up and perform these experiments as well as a robust method to measure peptide concentrations using tryptophan as an internal standard.
Published: 2015

26. Accurate protein-peptide titration experiments by nuclear magnetic resonance using low-volume samples

Author: Bruno Kieffer, Raphaël Recht, Christian Köhler, Marc-André Delsuc, Marc Quinternet, Frédéric De Lamotte, Kieffer, Bruno, Institut de Génétique et de Biologie Moléculaire et Cellulaire (IGBMC), Université de Strasbourg (UNISTRA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Centre National de la Recherche Scientifique (CNRS), Amélioration génétique et adaptation des plantes méditerranéennes et tropicales (UMR AGAP), Centre de Coopération Internationale en Recherche Agronomique pour le Développement (Cirad)-Institut National de la Recherche Agronomique (INRA)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro), and Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)
Subjects: Accuracy and precision, Materials science, Capillary action, méthode de mesure, Analytical chemistry, Peptide, 010402 general chemistry, 01 natural sciences, 03 medical and health sciences, Autre (Chimie), 030304 developmental biology, chemistry.chemical_classification, 0303 health sciences, Nuclear magnetic resonance spectroscopy, affinity measurements, NMR, 0104 chemical sciences, Dissociation constant, Low volume, protein-peptide interactions, chemistry, Titration, equilibrium binding constants, Other, spectrographie, [CHIM.OTHE]Chemical Sciences/Other
Abstract: AGAP : équipe ID; International audience; NMR spectroscopy allows measurements of very accurate values of equilibrium dissociation constants using chemical shift perturbation methods, provided that the concentrations of the binding partners are known with high precision and accuracy. The accuracy and precision of these experiments are improved if performed using individual capillary tubes, a method enabling full automation of the measurement. We provide here a protocol to set up and perform these experiments as well as a robust method to measure peptide concentrations using tryptophan as an internal standard.
Published: 2015
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27. (1)H, (15)N and (13)C resonance assignments of the yeast Pih1 and Tah1 C-terminal domains complex

Author: Marc Quinternet, Clémence Jacquemin, Bruno Charpentier, Xavier Manival, Ingénierie Moléculaire et Physiopathologie Articulaire (IMoPA), Centre National de la Recherche Scientifique (CNRS)-Université de Lorraine (UL), Service de néphrologie, Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpital Bicêtre-Institut National de la Santé et de la Recherche Médicale (INSERM), Institut Pasteur de la Guyane, Réseau International des Instituts Pasteur (RIIP), and Université de Lorraine (UL)-Centre National de la Recherche Scientifique (CNRS)
Subjects: Saccharomyces cerevisiae Proteins, Complex formation, Molecular Sequence Data, education, Saccharomyces cerevisiae, MESH: Amino Acid Sequence, Biology, Biochemistry, MESH: Protein Structure, Tertiary, MESH: Saccharomyces cerevisiae Proteins, Structural Biology, MESH: Nuclear Magnetic Resonance, Biomolecular, Amino Acid Sequence, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], Nuclear Magnetic Resonance, Biomolecular, MESH: Molecular Sequence Data, Nuclear Proteins, [SDV.BBM.BM]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, MESH: Saccharomyces cerevisiae, Yeast, Protein Structure, Tertiary, Crystallography, Chaperone (protein), biology.protein, MESH: Molecular Chaperones, MESH: Nuclear Proteins, [SDV.MHEP]Life Sciences [q-bio]/Human health and pathology, Molecular Chaperones
Abstract: International audience; We report the nearly complete (1)H, (15)N and (13)C resonance assignment of the complex formed by the C-terminal domains of Pih1 and Tah1 from S. cerevisiae and evidence the folding ability of Tah1 under complex formation.
Published: 2015
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28. 1H, 13C, and 15N resonance assignment of the C103S mutant of the N-terminal domain of DsbD from Neisseria meningitidis

Author: Pascale Tsan, Marc Quinternet, Marie-Christine Averlant-Petit, Guy Branlant, Manh-Thong Cung, Christophe Jacob, Chrystel Beaufils, Sandrine Boschi-Muller, Laure Selme, Laboratoire de Chimie Physique Macromoléculaire (LCPM), Institut de Chimie du CNRS (INC)-Université de Lorraine (UL)-Centre National de la Recherche Scientifique (CNRS), Maturation des ARN et enzymologie moléculaire (MAEM), Cancéropôle du Grand Est-Université Henri Poincaré - Nancy 1 (UHP)-IFR111-Centre National de la Recherche Scientifique (CNRS), Laboratoire Hippolyte Fizeau (FIZEAU), Université Nice Sophia Antipolis (... - 2019) (UNS), COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-Institut national des sciences de l'Univers (INSU - CNRS)-Observatoire de la Côte d'Azur, and Université Côte d'Azur (UCA)-COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-Université Côte d'Azur (UCA)-Centre National de la Recherche Scientifique (CNRS)
Subjects: Meningitides, Magnetic Resonance Spectroscopy, Stereochemistry, Molecular Sequence Data, Mutant, MESH: Amino Acid Sequence, Neisseria meningitidis, 010402 general chemistry, medicine.disease_cause, 01 natural sciences, Biochemistry, MESH: Neisseria meningitidis, Microbiology, 03 medical and health sciences, Bacterial Proteins, Structural Biology, medicine, Amino Acid Sequence, MESH: Bacterial Proteins, Peptide sequence, Protein secondary structure, 030304 developmental biology, Antigens, Bacterial, Carbon Isotopes, 0303 health sciences, MESH: Molecular Sequence Data, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], Nitrogen Isotopes, biology, MESH: Magnetic Resonance Spectroscopy, Chemistry, MESH: Molecular Weight, MESH: Carbon Isotopes, Nuclear magnetic resonance spectroscopy, biology.organism_classification, 0104 chemical sciences, Molecular Weight, MESH: Mutagenesis, Site-Directed, Domain (ring theory), Mutagenesis, Site-Directed, MESH: Protons, Neisseria, Protons, MESH: Antigens, Bacterial, MESH: Nitrogen Isotopes
Abstract: International audience; We report the nearly complete 1H, 13C, and 15N resonance assignments of the C103S mutant of the N-terminal domain of DsbD from Neisseria meningitides. Secondary structure determination using CSI method leads to the prediction of nine beta-sheet parts.
Published: 2008
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29. (1)H, (15)N and (13)C resonance assignments of the two TPR domains from the human RPAP3 protein

Author: Marc Quinternet, Christiane Branlant, Clémence Jacquemin, Marie-Eve Chagot, Bruno Charpentier, Xavier Manival, Ingénierie Moléculaire et Physiopathologie Articulaire (IMoPA), Centre National de la Recherche Scientifique (CNRS)-Université de Lorraine (UL), ARN-RNP, structure-fonction-maturation (AREMS), Université Henri Poincaré - Nancy 1 (UHP)-Centre National de la Recherche Scientifique (CNRS), Service de néphrologie, Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpital Bicêtre-Institut National de la Santé et de la Recherche Médicale (INSERM), Institut Pasteur de la Guyane, Réseau International des Instituts Pasteur (RIIP), and Université de Lorraine (UL)-Centre National de la Recherche Scientifique (CNRS)
Subjects: Protein family, Molecular Sequence Data, RNA polymerase II, MESH: Carrier Proteins, MESH: Amino Acid Sequence, Biochemistry, MESH: Protein Structure, Tertiary, Structural Biology, Heat shock protein, MESH: Nuclear Magnetic Resonance, Biomolecular, Humans, Amino Acid Sequence, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], Nuclear Magnetic Resonance, Biomolecular, MESH: Humans, MESH: Molecular Sequence Data, biology, [SDV.BBM.BM]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, Molecular biology, 3. Good health, Protein Structure, Tertiary, Tetratricopeptide, Chaperone (protein), biology.protein, Apoptosis Regulatory Proteins, Carrier Proteins, [SDV.MHEP]Life Sciences [q-bio]/Human health and pathology
Abstract: International audience; We report the nearly complete (1)H, (15)N and (13)C resonance assignments of the two tetratricopeptide-repeat domains of the human RPAP3 protein, a co-chaperone of the heat-shock protein family.
Published: 2014
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30. Characterization of the interaction between protein Snu13p/15.5K and the Rsa1p/NUFIP factor and demonstration of its functional importance for snoRNP assembly

Author: Bruno Charpentier, Xavier Manival, Régis Back, Magali Blaud, Edouard Bertrand, Marie Cécile Robert, Marc Quinternet, Jonathan Bizarro, Christophe Romier, Christiane Branlant, Benjamin Rothé, Ingénierie Moléculaire et Physiopathologie Articulaire (IMoPA), Université de Lorraine (UL)-Centre National de la Recherche Scientifique (CNRS), Faculté de Médecine [Nancy], Université de Lorraine (UL), Institut de Génétique Moléculaire de Montpellier (IGMM), Centre National de la Recherche Scientifique (CNRS)-Université de Montpellier (UM), Institut de Génétique et de Biologie Moléculaire et Cellulaire (IGBMC), Université de Strasbourg (UNISTRA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), UL, Imopa, Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS), Ingénierie Moléculaire et Physiopathologie Articulaire ( IMoPA ), Université de Lorraine ( UL ) -Centre National de la Recherche Scientifique ( CNRS ), Université de Lorraine ( UL ), Institut de Génétique Moléculaire de Montpellier ( IGMM ), Université de Montpellier ( UM ) -Centre National de la Recherche Scientifique ( CNRS ), Institut de Génétique et de Biologie Moléculaire et Cellulaire ( IGBMC ), and Université de Strasbourg ( UNISTRA ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Centre National de la Recherche Scientifique ( CNRS )
Subjects: Models, Molecular, Ribosomal Proteins, Saccharomyces cerevisiae Proteins, RNA Stability, Molecular Sequence Data, Static Electricity, Saccharomyces cerevisiae, RNA-binding protein, Plasma protein binding, Protein Structure, Secondary, [ SDV.BBM.BC ] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM], Ribonucleoproteins, Small Nucleolar, Ribosomal protein, [ SDV.MHEP ] Life Sciences [q-bio]/Human health and pathology, Genetics, Humans, Protein Interaction Domains and Motifs, Amino Acid Sequence, Small nucleolar RNA, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], [SDV.BBM.BC] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], Peptide sequence, [SDV.MHEP] Life Sciences [q-bio]/Human health and pathology, biology, Nuclear Proteins, Ribonucleoproteins, Small Nuclear, biology.organism_classification, Biochemistry, Biophysics, RNA, Chaperone complex, Hydrophobic and Hydrophilic Interactions, Small nuclear RNA, [SDV.MHEP]Life Sciences [q-bio]/Human health and pathology, Molecular Chaperones, Protein Binding
Abstract: International audience; The yeast Snu13p protein and its 15.5K human homolog both bind U4 snRNA and box C/D snoRNAs. They also bind the Rsa1p/NUFIP assembly factor, proposed to scaffold immature snoRNPs and to recruit the Hsp90-R2TP chaperone complex. However, the nature of the Snu13p/15.5K-Rsa1p/NUFIP interaction and its exact role in snoRNP assembly remained to be elucidated. By using biophysical, molecular and imaging approaches, here, we identify residues needed for Snu13p/15.5K-Rsa1p/NUFIP interaction. By NMR structure determination and docking approaches, we built a 3D model of the Snup13p-Rsa1p interface, suggesting that residues R-249, R-246 and K-250 in Rsa1p and E-72 and D-73 in Snu13p form a network of electrostatic interactions shielded from the solvent by hydrophobic residues from both proteins and that residue W-253 of Rsa1p is inserted in a hydrophobic cavity of Snu13p. Individual mutations of residues in yeast demonstrate the functional importance of the predicted interactions for both cell growth and snoRNP formation. Using archaeal box C/D sRNP 3D structures as templates, the association of Snu13p with Rsa1p is predicted to be exclusive of interactions in active snoRNPs. Rsa1p and NUFIP may thus prevent premature activity of pre-snoRNPs, and their removal may be a key step for active snoRNP production.
Published: 2014
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31. Heteronuclear NMR provides an accurate assessment of therapeutic insulin's quality

Author: Bruno Kieffer, Jean-Philippe Starck, Marc Quinternet, and Marc-André Delsuc
Subjects: Active ingredient, Magnetic Resonance Spectroscopy, Chemistry, media_common.quotation_subject, Clinical Biochemistry, Molecular Sequence Data, Pharmaceutical Science, Computational biology, Therapeutic Insulin, Analytical Chemistry, Nuclear magnetic resonance, Heteronuclear molecule, Drug Discovery, Humans, Insulin, Quality (business), Amino Acid Sequence, Spectroscopy, media_common
Abstract: New generations of drugs are using more and more often therapeutic proteins as the active ingredient, prompting the regulation agencies to adapt their analytical methods. Fast and unambiguous information on the secondary, tertiary and quaternary structure of the protein should be provided to assess the quality of these biodrugs. Recent developments of heteronuclear NMR methods, enabling their use on pharmaceutical formulated unlabeled proteins, provide an efficient way to perform such analysis, a feature that is illustrated here using various commercial formulations of insulins.
Published: 2012

32. Unraveling complex small-molecule binding mechanisms by using simple NMR spectroscopy

Author: Bruno Kieffer, Marc-André Delsuc, Marc Quinternet, and Jean-Philippe Starck
Subjects: Analytical chemistry, Serum albumin, Ligands, Catalysis, Dissociation (chemistry), Computational chemistry, Animals, Nuclear Magnetic Resonance, Biomolecular, biology, Molecular Structure, Chemistry, Organic Chemistry, Tryptophan, Isothermal titration calorimetry, Serum Albumin, Bovine, General Chemistry, Nuclear magnetic resonance spectroscopy, Ligand (biochemistry), Dissociation constant, Heteronuclear molecule, Models, Chemical, biology.protein, Thermodynamics, Cattle, Small molecule binding, Protein Binding
Abstract: Heteronuclear NMR spec- troscopy provides a unique way to obtain site-specific information about protein-ligand interactions. Usually, such studies rely on the availability of isotopically labeled proteins, thereby allowing both editing of the spectra and ligand signals to be filtered out. Herein, we report that the use of the methyl SOFAST correlation experi- ment enables the determination of site- specific equilibrium binding constants by using unlabeled proteins. By using the binding of l- and d-tryptophan to serum albumin as a test case, we deter- mined very accurate dissociation con- stants for both the high- and low-affini- ty sites present at the protein surface. The values of site-specific dissociation constants were closer to those obtained by isothermal titration calorimetry than those obtained from ligand-ob- served methods, such as saturation transfer difference. The possibility of measuring ligand binding to serum al- bumin at physiological concentrations with unlabeled proteins may open up new perspectives in the field of drug discovery.
Published: 2011

33. Formation of the Complex between DsbD and PilB N-Terminal Domains from Neisseria meningitidis Necessitates an Adaptability of nDsbD

Author: Guy Branlant, Marc Quinternet, Sandrine Boschi-Muller, Pascale Tsan, Christophe Jacob, Laure Selme-Roussel, Manh-Thong Cung, Laboratoire de Chimie Physique Macromoléculaire (LCPM), Institut de Chimie du CNRS (INC)-Université de Lorraine (UL)-Centre National de la Recherche Scientifique (CNRS), ARN-RNP, structure-fonction-maturation (AREMS), and Centre National de la Recherche Scientifique (CNRS)-Université Henri Poincaré - Nancy 1 (UHP)
Subjects: Models, Molecular, MICROBIO, PROTEINS, Electrons, Plasma protein binding, Neisseria meningitidis, Biology, medicine.disease_cause, Protein Structure, Secondary, Bacterial protein, 03 medical and health sciences, Bacterial Proteins, Structural Biology, Oxidoreductase, medicine, Disulfides, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], Nuclear Magnetic Resonance, Biomolecular, Molecular Biology, ComputingMilieux_MISCELLANEOUS, 030304 developmental biology, chemistry.chemical_classification, 0303 health sciences, Binding Sites, 030302 biochemistry & molecular biology, Disulfide bond, Membrane Proteins, Periplasmic space, Transmembrane protein, Protein Structure, Tertiary, Kinetics, Crystallography, chemistry, Periplasm, Biophysics, Oxidoreductases, Protein Binding
Abstract: SummaryDsbD transmembrane protein dispatches electrons to periplasmic Trx/DsbE-like partners via specific interactions with its N-terminal domain, nDsbD. In the present study, PilB N-terminal domain (NterPilB) is shown to efficiently accept electrons coming from nDsbD from Neisseria meningitidis. Using an NMR-driven docking approach, we have modeled the structure of a mixed disulfide complex between NterPilB and nDsbD. We show the needed opening of nDsbD cap-loop whereas NterPilB FLHE loop does not seem essential in the formation and stabilization of the complex. Relaxation analysis performed on backbone amide groups highlights a kind of dynamics transfer from nDsbD cap-loop on NterPilB α1 helix, suggesting that a mobility contribution is required not only for the formation of the mixed disulfide complex, but also for its disruption. Taking into account previous X-ray data on covalent complexes involving nDsbD, a cartoon of interactions between Trx-like partners and nDsbD is proposed that illustrates the adaptability of nDsbD.
Published: 2009
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34. The Spliceosomal Phosphopeptide P140 Controls the Lupus Disease by Interacting with the HSC70 Protein and via a Mechanism Mediated by γδ T Cells

Author: Jean-Marc Strub, Nicolas Schall, Jean-Paul Briand, Alain Van Dorsselaer, Manh Thong Cung, Sylviane Muller, Nicolas Page, Marion Decossas, Marc Quinternet, Olivier Chaloin, Immunologie et chimie thérapeutiques (ICT), Cancéropôle du Grand Est-Centre National de la Recherche Scientifique (CNRS), Laboratoire de Spectrométrie de Masse BioOrganique [Strasbourg] (LSMBO), Département Sciences Analytiques et Interactions Ioniques et Biomoléculaires (DSA-IPHC), Institut Pluridisciplinaire Hubert Curien (IPHC), Université de Strasbourg (UNISTRA)-Institut National de Physique Nucléaire et de Physique des Particules du CNRS (IN2P3)-Centre National de la Recherche Scientifique (CNRS)-Université de Strasbourg (UNISTRA)-Institut National de Physique Nucléaire et de Physique des Particules du CNRS (IN2P3)-Centre National de la Recherche Scientifique (CNRS)-Institut Pluridisciplinaire Hubert Curien (IPHC), Université de Strasbourg (UNISTRA)-Institut National de Physique Nucléaire et de Physique des Particules du CNRS (IN2P3)-Centre National de la Recherche Scientifique (CNRS)-Université de Strasbourg (UNISTRA)-Institut National de Physique Nucléaire et de Physique des Particules du CNRS (IN2P3)-Centre National de la Recherche Scientifique (CNRS), Laboratoire de Chimie Physique Macromoléculaire (LCPM), and Institut de Chimie du CNRS (INC)-Université de Lorraine (UL)-Centre National de la Recherche Scientifique (CNRS)
Subjects: Mice, Inbred MRL lpr, T-Lymphocytes, Immunology/Innate Immunity, Immunology/Immunomodulation, Fluorescent Antibody Technique, Apoptosis, medicine.disease_cause, T-Lymphocytes, Regulatory, Autoimmunity, Interleukin 21, Mice, 0302 clinical medicine, immune system diseases, Biochemistry/Cell Signaling and Trafficking Structures, Cytotoxic T cell, Lupus Erythematosus, Systemic, Biochemistry/Biomacromolecule-Ligand Interactions, 0303 health sciences, B-Lymphocytes, Multidisciplinary, ZAP70, Receptors, Antigen, T-Cell, gamma-delta, 3. Good health, medicine.anatomical_structure, 030220 oncology & carcinogenesis, Medicine, Biochemistry/Drug Discovery, Research Article, T cell, Science, Immunology/Autoimmunity, Down-Regulation, Biology, Models, Biological, Ribonucleoprotein, U1 Small Nuclear, 03 medical and health sciences, Antigen, medicine, Animals, B cell, Rheumatology/Autoimmunity, Autoimmune, and Inflammatory Diseases, 030304 developmental biology, Binding Sites, HSC70 Heat-Shock Proteins, [SDV.IMM.IMM]Life Sciences [q-bio]/Immunology/Immunotherapy, Surface Plasmon Resonance, Molecular biology, Peptide Fragments, Peripheral blood lymphocyte, Rheumatology/Systemic Lupus Erythematosos, Immunology/Immune Response
Abstract: International audience; The phosphopeptide P140 issued from the spliceosomal U1-70K snRNP protein is recognized by lupus CD4(+) T cells, transiently abolishes T cell reactivity to other spliceosomal peptides in P140-treated MRL/lpr mice, and ameliorates their clinical features. P140 modulates lupus patients' T cell response ex vivo and is currently included in phase IIb clinical trials. Its underlying mechanism of action remains elusive. Here we show that P140 peptide binds a unique cell-surface receptor, the constitutively-expressed chaperone HSC70 protein, known as a presenting-protein. P140 induces apoptosis of activated MRL/lpr CD4(+) T cells. In P140-treated mice, it increases peripheral blood lymphocyte apoptosis and decreases B cell, activated T cell, and CD4(-)CD8(-)B220(+) T cell counts via a specific mechanism strictly depending on gammadelta T cells. Expression of inflammation-linked genes is rapidly regulated in CD4(+) T cells. This work led us to identify a powerful pathway taken by a newly-designed therapeutic peptide to immunomodulate lupus autoimmunity.
Published: 2009
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35. Solution structure and dynamics of the reduced and oxidized forms of the N-terminal domain of PilB from Neisseria meningitidis

Author: Marie-Christine Averlant-Petit, Sandrine Boschi-Muller, Chrystel Beaufils, Marc Quinternet, Guy Branlant, Pascale Tsan, Manh-Thong Cung, Fabrice Neiers, Laboratoire de Chimie Physique Macromoléculaire (LCPM), Institut de Chimie du CNRS (INC)-Université de Lorraine (UL)-Centre National de la Recherche Scientifique (CNRS), Maturation des ARN et enzymologie moléculaire (MAEM), and Cancéropôle du Grand Est-Université Henri Poincaré - Nancy 1 (UHP)-IFR111-Centre National de la Recherche Scientifique (CNRS)
Subjects: Models, Molecular, MESH: Oxidation-Reduction, MESH: Hydrogen-Ion Concentration, Magnetic Resonance Spectroscopy, Cytochrome, Protein Conformation, Stereochemistry, Neisseria meningitidis, 010402 general chemistry, medicine.disease_cause, 01 natural sciences, Biochemistry, Redox, MESH: Neisseria meningitidis, 03 medical and health sciences, MESH: Protein Structure, Tertiary, MESH: Protein Conformation, Bacterial Proteins, medicine, MESH: Oxidoreductases, MESH: Bacterial Proteins, 030304 developmental biology, chemistry.chemical_classification, 0303 health sciences, biology, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], MESH: Magnetic Resonance Spectroscopy, Active site, Hydrogen-Ion Concentration, Electron transport chain, Protein Structure, Tertiary, 0104 chemical sciences, Enzyme, chemistry, biology.protein, Methionine sulfoxide reductase, Protein topology, Oxidoreductases, Oxidation-Reduction, MESH: Models, Molecular
Abstract: International audience; The secreted form of the PilB protein was proposed to be involved in pathogen survival fighting against the defensive host's oxidative burst. PilB protein is composed of three domains. The central and the C-terminal domains display methionine sulfoxide reductase A and B activities, respectively. The N-terminal domain, which possesses a CXXC motif, was recently shown to regenerate in vitro the reduced forms of the methionine sulfoxide reductase domains of PilB from their oxidized forms, as does the thioredoxin 1 from E. coli, via a disulfide bond exchange. The thioredoxin-like N-terminal domain belongs to the cytochrome maturation protein structural family, but it possesses a unique additional segment (99)FLHE (102) localized in a loop. This segment covers one edge of the active site in the crystal structure of the reduced form of the N-terminal domain of PilB. We have determined the solution structure and the dynamics of the N-terminal domain from Neisseria meningitidis, in its reduced and oxidized forms. The FLHE loop adopts, in both redox states, a well-defined conformation. Subtle conformational and dynamic changes upon oxidation are highlighted around the active site, as well as in the FLHE loop. The functional consequences of the cytochrome maturation protein topology and those of the presence of FLHE loop are discussed in relation to the enzymatic properties of the N-terminal domain.
Published: 2008
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36. 1H, 13C and 15N resonance assignment of the oxidized form (Cys(67)-Cys (70)) of the N-terminal domain of PilB from Neisseria meningitidis

Author: Fabrice Neiers, Marc Quinternet, Sandrine Boschi-Muller, Marie-Christine Averlant-Petit, Chrystel Beaufils, Manh-Thong Cung, Pascale Tsan, Guy Branlant, Laboratoire de Chimie Physique Macromoléculaire (LCPM), Institut de Chimie du CNRS (INC)-Université de Lorraine (UL)-Centre National de la Recherche Scientifique (CNRS), Maturation des ARN et enzymologie moléculaire (MAEM), and Cancéropôle du Grand Est-Université Henri Poincaré - Nancy 1 (UHP)-IFR111-Centre National de la Recherche Scientifique (CNRS)
Subjects: MESH: Oxidation-Reduction, MESH: Molecular Structure, MESH: Protein Structure, Secondary, Neisseria meningitidis, 010402 general chemistry, medicine.disease_cause, 01 natural sciences, Biochemistry, Protein Structure, Secondary, MESH: Neisseria meningitidis, MESH: Recombinant Proteins, 03 medical and health sciences, MESH: Protein Structure, Tertiary, Protein structure, Bacterial Proteins, Structural Biology, MESH: Nuclear Magnetic Resonance, Biomolecular, medicine, MESH: Oxidoreductases, Protein secondary structure, Nuclear Magnetic Resonance, Biomolecular, MESH: Bacterial Proteins, 030304 developmental biology, 0303 health sciences, biology, Molecular Structure, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], Chemistry, Disulfide bond, Resonance, Oxidation reduction, MESH: Cystine, biology.organism_classification, Recombinant Proteins, 0104 chemical sciences, 3. Good health, Protein Structure, Tertiary, Crystallography, Talos, Domain (ring theory), Cystine, Oxidoreductases, Oxidation-Reduction
Abstract: International audience; We report the nearly complete 1H, 13C and 15N resonance assignments of the oxidized form (Cys(67)-Cys(70)) of the N-terminal domain of PilB from Neisseria meningitidis. Secondary structure determination using CSI method and TALOS leads mainly to the prediction of 7 alpha-helical and 5 beta-sheet parts.
Published: 2007
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37. First case of emergence of atovaquone resistance in Plasmodium falciparum during second-line atovaquone-proguanil treatment in South America

Author: Béatrice Volney, Eric Legrand, Philippe Esterre, Marc Quinternet, Christophe Rogier, Bernard Carme, Odile Puijalon, Magalie Demar, Marie-Thérèse Ekala, Christiane Bouchier, Thierry Fandeur, Institut Pasteur de la Guyane, Réseau International des Instituts Pasteur (RIIP), Unité de Maladies infectieuses et tropicales-Hygiène hospitalière (EA 3593), Centre Hospitalier Andrée Rosemon [Cayenne, Guyane Française], Immunologie moléculaire des parasites, Institut Pasteur [Paris]-Centre National de la Recherche Scientifique (CNRS), Génomique (Plate-Forme) - Genomics Platform, Institut Pasteur [Paris], Immunologie parasitaire (TOURS UMR IMMUNOLOGIE PARASIT), Institut National de la Recherche Agronomique (INRA)-Faculte des Sciences Pharmaceutiques, Unité de Recherche en Biologie et Epidémiologie Parasitaires, Institut de Médecine Tropicale du Service de Santé des Armées-Centre National de la Recherche Scientifique (CNRS), Epidémiologie des parasitoses et mycoses tropicales, Institut National de la Santé et de la Recherche Médicale (INSERM)-Université des Antilles et de la Guyane (UAG), Centre Hospitalier Andrée Rosemon, Génomique (Plate-Forme), Université des Antilles et de la Guyane (UAG)-Institut National de la Santé et de la Recherche Médicale (INSERM), Institut Pasteur [Paris] (IP)-Centre National de la Recherche Scientifique (CNRS), and Institut Pasteur [Paris] (IP)
Subjects: Drug Resistance, Drug resistance, Pharmacology, MESH: Parasitic Sensitivity Tests, Treatment failure, 0302 clinical medicine, Second line, Parasitic Sensitivity Tests, Pharmacology (medical), MESH: Animals, Malaria, Falciparum, Letter to the Editor, ComputingMilieux_MISCELLANEOUS, MESH: Plasmodium falciparum, 0303 health sciences, biology, MESH: Malaria, Falciparum, MESH: Proguanil, 3. Good health, Infectious Diseases, Proguanil, Combination, MESH: Drug Resistance, Drug Therapy, Combination, Atovaquone, medicine.drug, Falciparum, 030231 tropical medicine, Plasmodium falciparum, MESH: Atovaquone, 03 medical and health sciences, Antimalarials, Drug Therapy, parasitic diseases, medicine, Animals, Humans, [SDV.MP.PAR]Life Sciences [q-bio]/Microbiology and Parasitology/Parasitology, MESH: Humans, 030306 microbiology, MESH: South America, South America, medicine.disease, biology.organism_classification, Atovaquone/proguanil, Virology, MESH: Antimalarials, Malaria, MESH: Drug Therapy, Combination, human activities
Abstract: The atovaquone-proguanil combination (Malarone) has been introduced in French Guiana for prophylaxis and second-line treatment for Plasmodium falciparum malaria in 2002. We report here a treatment failure in a patient who was given a second-line atovaquone-proguanil treatment. A nonimmune P.
Published: 2007
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