39 results on '"Christine Schaeffer-Reiss"'
Search Results
2. Comparative Study of Secreted Proteins, Enzymatic Activities of Wood Degradation and Stilbene Metabolization in Grapevine Botryosphaeria Dieback Fungi
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Clément Labois, Elodie Stempien, Justine Schneider, Christine Schaeffer-Reiss, Christophe Bertsch, Mary-Lorène Goddard, and Julie Chong
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Botryosphaeriaceae ,grapevine ,stilbene metabolization ,secreted proteins ,Biology (General) ,QH301-705.5 - Abstract
Botryosphaeriaceae fungi are plant pathogens associated with Botryosphaeria dieback. To better understand the virulence factors of these fungi, we investigated the diversity of secreted proteins and extracellular enzyme activities involved in wood degradation and stilbene metabolization in Neofusicoccum parvum and Diplodia seriata, which are two major fungi associated with grapevine B. dieback. Regarding the analysis of proteins secreted by the two fungi, our study revealed that N. parvum, known to be more aggressive than D. seriata, was characterized by a higher quantity and diversity of secreted proteins, especially hydrolases and oxidoreductases that are likely involved in cell wall and lignin degradation. In addition, when fungi were grown with wood powder, the extracellular laccase and Mn peroxidase enzyme activities were significantly higher in D. seriata compared to N.parvum. Importantly, our work also showed that secreted Botryosphaeriaceae proteins produced after grapevine wood addition are able to rapidly metabolize the grapevine stilbenes. Overall, a higher diversity of resveratrol and piceatannol metabolization products was found with enzymes of N. parvum compared to D. seriata. This study emphasizes the diversity of secreted virulence factors found in B. dieback fungi and suggests that some resveratrol oligomers produced in grapevine wood after pathogen attack could be formed via pathogenic fungal oxidases.
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- 2021
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3. Unconventional endosome-like compartment and retromer complex in Toxoplasma gondii govern parasite integrity and host infection
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Lamba Omar Sangaré, Tchilabalo Dilezitoko Alayi, Benoit Westermann, Agnes Hovasse, Fabien Sindikubwabo, Isabelle Callebaut, Elisabeth Werkmeister, Frank Lafont, Christian Slomianny, Mohamed-Ali Hakimi, Alain Van Dorsselaer, Christine Schaeffer-Reiss, and Stanislas Tomavo
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Science - Abstract
The retromer complex is a multi-protein component of the endosomal protein sorting machinery. Here, Sangaré et al. identify unique features in the retromer complex of the parasite Toxoplasma gondii, and show that it is crucial for the biogenesis of secretory organelles in this pathogen.
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- 2016
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4. Flowering as the Most Highly Sensitive Period of Grapevine (Vitis vinifera L. cv Mourvèdre) to the Botryosphaeria Dieback Agents Neofusicoccum parvum and Diplodia seriata Infection
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Alessandro Spagnolo, Philippe Larignon, Maryline Magnin-Robert, Agnès Hovasse, Clara Cilindre, Alain Van Dorsselaer, Christophe Clément, Christine Schaeffer-Reiss, and Florence Fontaine
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Botryosphaeria dieback ,Neofusicoccum parvum ,Diplodia seriata ,plant proteomics ,two dimensional gel electrophoresis ,defense-related proteins ,Biology (General) ,QH301-705.5 ,Chemistry ,QD1-999 - Abstract
Botryosphaeria dieback is a fungal grapevine trunk disease that currently represents a threat for viticulture worldwide because of the important economical losses due to reduced yield of affected plants and their premature death. Neofusicoccum parvum and Diplodia seriata are among the causal agents. Vine green stems were artificially infected with N. parvum or D. seriata at the onset of three different phenological stages (G stage (separated clusters), flowering and veraison). Highest mean lesion lengths were recorded at flowering. Major proteome changes associated to artificial infections during the three different phenological stages were also reported using two dimensional gel electrophoresis (2D)-based analysis. Twenty (G stage), 15 (flowering) and 13 (veraison) differentially expressed protein spots were subjected to nanoLC-MS/MS and a total of 247, 54 and 25 proteins were respectively identified. At flowering, a weaker response to the infection was likely activated as compared to the other stages, and some defense-related proteins were even down regulated (e.g., superoxide dismutase, major latex-like protein, and pathogenesis related protein 10). Globally, the flowering period seemed to represent the period of highest sensitivity of grapevine to Botryosphaeria dieback agent infection, possibly being related to the high metabolic activity in the inflorescences.
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- 2014
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5. Proteomic insights into changes in grapevine wood in response to esca proper and apoplexy
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Maryline MAGNIN-ROBERT, Alessandro SPAGNOLO, Tchilabalo DILEZITOKO ALAYI, Clara CILINDRE, Laurence MERCIER, Christine SCHAEFFER-REISS, Alain VAN DORSSELAER, Christophe CLÉMENT, and Florence FONTAINE
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grapevine ,proteomics ,qRT-PCR ,trunk diseases ,Botany ,QK1-989 - Abstract
Among fungal grapevine trunk diseases, esca proper poses a significant threat for viticulture. Apoplexy, mainly occurring on grapevines affected by esca proper, is also a threat. To verify if different responses are activated in the woody tissues of apoplectic (A) and esca proper-affected (E) vines, two-dimensional gel electrophoresis coupled to mass spectrometry analysis was used to examine changes in the trunk wood of E and A field-grown plants. Asymptomatic and black streaked trunk (BST) wood from A and E plants were compared to asymptomatic and BST wood of control plants. Twenty-seven differentially expressed protein spots were identified. For eleven targeted proteins, expression of the relative transcripts was also monitored by qRT-PCR. Hierarchical tree clustering revealed differences in the distribution of spots containing carbohydrate metabolism proteins and heat shock proteins between asymptomatic- and BST-wood of grapevine, irrespective of the type of plant examined (control or diseased grapevines). Asymptomatic wood was mainly characterized by down-expression of proteins involved in cell growth and defense responses. The proteome of BST wood, characterized by extensive presence of grapevine trunk disease agents, revealed over-expression of proteins involved in defense. There was no evidence of strong different response in the trunk wood of apoplectic and esca proper affected plants. This could mean that, despite the different feature of these external crown symptoms, grapevine responses at trunk wood level is very similar in both cases. This plant response might therefore either simply be due to the fact that plants can react in the same way to different stresses, or that apoplexy represents a different effect provoked by the same cause or causal agent(s).
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- 2014
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6. The Pollen Coat Proteome: At the Cutting Edge of Plant Reproduction
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Juan David Rejón, François Delalande, Christine Schaeffer-Reiss, Juan de Dios Alché, María Isabel Rodríguez-García, Alain Van Dorsselaer, and Antonio Jesús Castro
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olive ,pollen coat ,proteomics ,self-incompatibility ,tapetum ,Microbiology ,QR1-502 - Abstract
The tapetum is a single layer of secretory cells which encloses the anther locule and sustains pollen development and maturation. Upon apoptosis, the remnants of the tapetal cells, consisting mostly of lipids and proteins, fill the pits of the sculpted exine to form the bulk of the pollen coat. This extracellular matrix forms an impermeable barrier that protects the male gametophyte from water loss and UV light. It also aids pollen adhesion and hydration and retains small signaling compounds involved in pollen–stigma communication. In this study, we have updated the list of the pollen coat’s protein components and also discussed their functions in the context of sexual reproduction
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- 2016
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7. Detection of prion protein in urine-derived injectable fertility products by a targeted proteomic approach.
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Alain Van Dorsselaer, Christine Carapito, François Delalande, Christine Schaeffer-Reiss, Daniele Thierse, Hélène Diemer, Douglas S McNair, Daniel Krewski, and Neil R Cashman
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Medicine ,Science - Abstract
BackgroundIatrogenic transmission of human prion disease can occur through medical or surgical procedures, including injection of hormones such as gonadotropins extracted from cadaver pituitaries. Annually, more than 300,000 women in the United States and Canada are prescribed urine-derived gonadotropins for infertility. Although menopausal urine donors are screened for symptomatic neurological disease, incubation of Creutzfeldt-Jakob disease (CJD) is impossible to exclude by non-invasive testing. Risk of carrier status of variant CJD (vCJD), a disease associated with decades-long peripheral incubation, is estimated to be on the order of 100 per million population in the United Kingdom. Studies showing infectious prions in the urine of experimental animals with and without renal disease suggest that prions could be present in asymptomatic urine donors. Several human fertility products are derived from donated urine; recently prion protein has been detected in preparations of human menopausal gonadotropin (hMG).Methodology/principal findingsUsing a classical proteomic approach, 33 and 34 non-gonadotropin proteins were identified in urinary human chorionic gonadotropin (u-hCG) and highly-purified urinary human menopausal gonadotropin (hMG-HP) products, respectively. Prion protein was identified as a major contaminant in u-hCG preparations for the first time. An advanced prion protein targeted proteomic approach was subsequently used to conduct a survey of gonadotropin products; this approach detected human prion protein peptides in urine-derived injectable fertility products containing hCG, hMG and hMG-HP, but not in recombinant products.Conclusions/significanceThe presence of protease-sensitive prion protein in urinary-derived injectable fertility products containing hCG, hMG, and hMG-HP suggests that prions may co-purify in these products. Intramuscular injection is a relatively efficient route of transmission of human prion disease, and young women exposed to prions can be expected to survive an incubation period associated with a minimal inoculum. The risks of urine-derived fertility products could now outweigh their benefits, particularly considering the availability of recombinant products.
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- 2011
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8. A novel Toxoplasma gondii nuclear factor TgNF3 is a dynamic chromatin-associated component, modulator of nucleolar architecture and parasite virulence.
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Alejandro Olguin-Lamas, Edwige Madec, Agnes Hovasse, Elisabeth Werkmeister, Isabelle Callebaut, Christian Slomianny, Stephane Delhaye, Thomas Mouveaux, Christine Schaeffer-Reiss, Alain Van Dorsselaer, and Stanislas Tomavo
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Immunologic diseases. Allergy ,RC581-607 ,Biology (General) ,QH301-705.5 - Abstract
In Toxoplasma gondii, cis-acting elements present in promoter sequences of genes that are stage-specifically regulated have been described. However, the nuclear factors that bind to these cis-acting elements and regulate promoter activities have not been identified. In the present study, we performed affinity purification, followed by proteomic analysis, to identify nuclear factors that bind to a stage-specific promoter in T. gondii. This led to the identification of several nuclear factors in T. gondii including a novel factor, designated herein as TgNF3. The N-terminal domain of TgNF3 shares similarities with the N-terminus of yeast nuclear FK506-binding protein (FKBP), known as a histone chaperone regulating gene silencing. Using anti-TgNF3 antibodies, HA-FLAG and YFP-tagged TgNF3, we show that TgNF3 is predominantly a parasite nucleolar, chromatin-associated protein that binds specifically to T. gondii gene promoters in vivo. Genome-wide analysis using chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-seq) identified promoter occupancies by TgNF3. In addition, TgNF3 has a direct role in transcriptional control of genes involved in parasite metabolism, transcription and translation. The ectopic expression of TgNF3 in the tachyzoites revealed dynamic changes in the size of the nucleolus, leading to a severe attenuation of virulence in vivo. We demonstrate that TgNF3 physically interacts with H3, H4 and H2A/H2B assembled into bona fide core and nucleosome-associated histones. Furthermore, TgNF3 interacts specifically to histones in the context of stage-specific gene silencing of a promoter that lacks active epigenetic acetylated histone marks. In contrast to virulent tachyzoites, which express the majority of TgNF3 in the nucleolus, the protein is exclusively located in the cytoplasm of the avirulent bradyzoites. We propose a model where TgNF3 acts essentially to coordinate nucleolus and nuclear functions by modulating nucleosome activities during the intracellular proliferation of the virulent tachyzoites of T. gondii.
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- 2011
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9. Surface charge influences protein corona, cell uptake and biological effects of carbon dots
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Yasmin Arezki, François Delalande, Christine Schaeffer-Reiss, Sarah Cianférani, Mickaël Rapp, Luc Lebeau, Françoise Pons, and Carole Ronzani
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Proteomics ,Surface Properties ,Aucun ,Carbon ,Citric Acid ,Albumins ,Nanoparticles ,Protein Corona ,General Materials Science ,Adiponectin ,Vitronectin ,Amines ,Apolipoproteins C ,Fetuins ,Apolipoproteins B - Abstract
Carbon dots are emerging nanoparticles (NPs) with tremendous applications, especially in the biomedical field. Herein is reported the first quantitative proteomic analysis of the protein corona formed on CDs with different surface charge properties. Four CDs were synthesized from citric acid and various amine group-containing passivation reagents, resulting in cationic NPs with increasing zeta (ζ)-potential and density of positive charges. After CD contact with serum, we show that protein corona identity is influenced by CD surface charge properties, which in turn impacts CD uptake and viability loss in macrophages. In particular, CDs with high ζ-potential (+30 mV) and charge density (2 μmol mg
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- 2022
10. Differential Phosphoproteomics Deciphers Physiopathology of High-Risk Mantle Cell Lymphoma
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Charline Fuseau, Mathieu Baldacini, Alina Nicolae, Laurent Miguet, Laurent Mauvieux, Laurent Vallat, Claire Domon-Dell, Manuela Tavian, Jean-Noel Freund, Sarah Cianferani, Christine Schaeffer-Reiss, Luc-Matthieu Fornecker, and Delphine C.M. Rolland
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Immunology ,Cell Biology ,Hematology ,Biochemistry - Published
- 2022
11. A class of valuable (pro-)activity-based protein profiling probes: application to the redox-active antiplasmodial agent, plasmodione
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Sarah Cianférani, Vrushali Khobragade, Christine Schaeffer-Reiss, Leandro Cotos, Maxime Donzel, Mourad Elhabiri, Elisabeth Davioud-Charvet, Bogdan Adam Cichocki, Stéphanie Blandin, Jean-Marc Strub, Gaillard, Brigitte, Laboratoire d'innovation moléculaire et applications (LIMA), Université de Strasbourg (UNISTRA)-Université de Haute-Alsace (UHA) Mulhouse - Colmar (Université de Haute-Alsace (UHA))-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), Réponse immunitaire et developpement chez les insectes (RIDI - UPR 9002), 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)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), 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), 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), Modèles Insectes de l'Immunité Innée (M3I), Université de Strasbourg (UNISTRA)-Centre National de la Recherche Scientifique (CNRS)-Université de Strasbourg (UNISTRA)-Centre National de la Recherche Scientifique (CNRS)-Université de Strasbourg (UNISTRA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-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), and ANR-11-LABX-0024,ParaFrap,Alliance française contre les maladies parasitaires(2011)
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quinone ,[SDV]Life Sciences [q-bio] ,[CHIM.THER]Chemical Sciences/Medicinal Chemistry ,010402 general chemistry ,01 natural sciences ,Interactome ,Article ,electrophile ,chemistry.chemical_compound ,3-benz(o)ylmenadione ,[CHIM] Chemical Sciences ,[CHIM]Chemical Sciences ,Binding site ,Mode of action ,photoaffinity labelling ,Heme ,QD1-999 ,antimalarial ,Photoaffinity labeling ,010405 organic chemistry ,Activity-based proteomics ,Combinatorial chemistry ,photoredox ,0104 chemical sciences ,3. Good health ,Quinone ,[SDV] Life Sciences [q-bio] ,Chemistry ,chemistry ,Activity-based protein profiling ,Chimie/Chimie thérapeutique ,CuAAC ,photoaffinity labeling ,Protein ligand - Abstract
International audience; Plasmodione (PD) is a potent antimalarial redox-active drug acting at low nM range concentrations on different malaria parasite stages. In this study, in order to determine the precise PD protein interactome in parasites, we developed a class of (pro-)activity-based protein profiling probes (ABPP) as precursors of photoreactive benzophenonelike probes based on the skeleton of PD metabolites (PDO) generated in a cascade of redox reactions. Under UV-photoirradiation, we clearly demonstrate that benzylic oxidation of 3-benzylmenadione 11 produces the 3-benzoylmenadione probe 7, allowing investigation of the proof-of-concept of the ABPP strategy with 3-benzoylmenadiones 7-10. The synthesized 3-benzoylmenadiones, probe 7 with an alkyne group or probe 9 with-NO2 in para position of the benzoyl chain, were found to be the most efficient photoreactive and clickable probes. In the presence of various H-donor partners, the UV-irradiation of the photo-reactive ABPP probes generates different adducts, the expected 'benzophenone-like' adducts (pathway 1) in addition to 'benzoxanthone' adducts (via two other pathways, 2 and 3). Using both human and Plasmodium falciparum glutathione reductases three protein ligand binding sites were identified following photolabeling with probes 7 or 9. The photoreduction of 3-benzoylmenadiones (PDO and probe 9) promoting the formation of both the corresponding benzoxanthone and the derived enone could be replaced by the glutathione reductase-catalyzed reduction step. In particular, the electrophilic character of the benzoxanthone was evidenced by its ability to alkylate heme, as a relevant event supporting the antimalarial mode of action of PD. This work provides a proof-of-principle that (pro-)ABPP probes can generate benzophenonelike metabolites enabling optimized activity-based protein profiling conditions that will be instrumental to analyse the interactome of early-lead antiplasmodial 3-benzylmenadiones displaying an original and innovative mode of action.
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- 2021
12. Correction: An essential role for α4A-tubulin in platelet biogenesis
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Catherine Strassel, Maria M Magiera, Arnaud Dupuis, Morgane Batzenschlager, Agnès Hovasse, Irina Pleines, Paul Guéguen, Anita Eckly, Sylvie Moog, Léa Mallo, Quentin Kimmerlin, Stéphane Chappaz, Jean-Marc Strub, Natarajan Kathiresan, Henri de la Salle, Alain Van Dorsselaer, Claude Ferec, Jean-Yves Py, Christian Gachet, Christine Schaeffer-Reiss, Benjamin T Kile, Carsten Janke, and François Lanza
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Ecology ,biology ,Health, Toxicology and Mutagenesis ,Published Erratum ,macromolecular substances ,Plant Science ,Computational biology ,Biochemistry, Genetics and Molecular Biology (miscellaneous) ,Blot ,Tubulin ,biology.protein ,Platelet ,Research Articles ,Biogenesis ,Research Article - Abstract
Alpha4A-tubulin is the predominant α-tubulin isotype in platelets. Mutations in α4A-tubulin cause abnormal platelet biogenesis and marginal band formation in mice and in a patient, establishing an essential role of this tubulin isotype., During platelet biogenesis, microtubules (MTs) are arranged into submembranous structures (the marginal band) that encircle the cell in a single plane. This unique MT array has no equivalent in any other mammalian cell, and the mechanisms responsible for this particular mode of assembly are not fully understood. One possibility is that platelet MTs are composed of a particular set of tubulin isotypes that carry specific posttranslational modifications. Although β1-tubulin is known to be essential, no equivalent roles of α-tubulin isotypes in platelet formation or function have so far been reported. Here, we identify α4A-tubulin as a predominant α-tubulin isotype in platelets. Similar to β1-tubulin, α4A-tubulin expression is up-regulated during the late stages of megakaryocyte differentiation. Missense mutations in the α4A-tubulin gene cause macrothrombocytopenia in mice and humans. Defects in α4A-tubulin lead to changes in tubulin tyrosination status of the platelet tubulin pool. Ultrastructural defects include reduced numbers and misarranged MT coils in the platelet marginal band. We further observed defects in megakaryocyte maturation and proplatelet formation in Tuba4a-mutant mice. We have, thus, discovered an α-tubulin isotype with specific and essential roles in platelet biogenesis.
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- 2021
13. A spatial vascular transcriptomic, proteomic, and phosphoproteomic atlas unveils an angiocrine Tie–Wnt signaling axis in the liver
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Jingjing Shi, Moritz Jakab, Mathias Heikenwalder, Shalev Itzkovitz, Martin Schneider, Indrabahadur Singh, Shubhada Rajabhau Kulkarni, Paula Argos Vélez, Michael Boutros, Maria Riedel, Ki Hong Lee, Thomas Ruppert, Sudhakar Chintharlapalli, Hellmut G. Augustin, Dominic Helm, Carleen Spegg, Guanxiong Wang, Marziyeh Komeili, Christine Schaeffer-Reiss, Shani Ben-Moshe, and Donato Inverso
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Proteomics ,Resource ,Quantitative proteomics ,Biology ,General Biochemistry, Genetics and Molecular Biology ,Receptor tyrosine kinase ,transcriptomics ,Wnt ,03 medical and health sciences ,chemistry.chemical_compound ,0302 clinical medicine ,liver endothelial cell (L-EC) ,Humans ,Regeneration ,Endothelium ,RNA-Seq ,Phosphorylation ,Wnt Signaling Pathway ,Molecular Biology ,030304 developmental biology ,0303 health sciences ,Phosphoproteomics ,Wnt signaling pathway ,Endothelial Cells ,Gene Expression Regulation, Developmental ,phosphoproteomics ,vascular zonation ,Tyrosine phosphorylation ,Cell Biology ,Flow Cytometry ,Phosphoproteins ,Liver Regeneration ,Cell biology ,Tie2 ,Liver ,Tie1 ,chemistry ,angiocrine factors ,Proteome ,Hepatocytes ,biology.protein ,Single-Cell Analysis ,Transcriptome ,030217 neurology & neurosurgery ,Developmental Biology - Abstract
Summary Single-cell transcriptomics (scRNA-seq) has revolutionized the understanding of the spatial architecture of tissue structure and function. Advancing the “transcript-centric” view of scRNA-seq analyses is presently restricted by the limited resolution of proteomics and genome-wide techniques to analyze post-translational modifications. Here, by combining spatial cell sorting with transcriptomics and quantitative proteomics/phosphoproteomics, we established the spatially resolved proteome landscape of the liver endothelium, yielding deep mechanistic insight into zonated vascular signaling mechanisms. Phosphorylation of receptor tyrosine kinases was detected preferentially in the central vein area, resulting in an atypical enrichment of tyrosine phosphorylation. Prototypic biological validation identified Tie receptor signaling as a selective and specific regulator of vascular Wnt activity orchestrating angiocrine signaling, thereby controlling hepatocyte function during liver regeneration. Taken together, the study has yielded fundamental insight into the spatial organization of liver endothelial cell signaling. Spatial sorting may be employed as a universally adaptable strategy for multiomic analyses of scRNA-seq-defined cellular (sub)-populations., Graphical abstract, Highlights • ScRNA-seq-guided spatial sort enables multiomic dissection of the liver vasculature • Liver sinusoidal endothelial cells have a hybrid vascular-lymphatic phenotype • Tyrosine phosphorylation of endothelial cell molecules is enriched on central vein • Endothelial Tie1 shapes hepatic Wnt signal zonation and promotes liver regeneration, Inverso, Shi et al. generate a multiomic encyclopedia of liver endothelial cells (L-ECs) with spatial resolution of transcriptome, proteome, and phosphoproteome. The study provides insight into liver vascular zonation and a template for scRNA-seq-data-guided spatial proteome and phosphoproteome analyses.
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- 2021
14. Phosphomimetic substitution at Ser-33 of the chloroquine resistance transporter PfCRT reconstitutes drug responses in
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Cecilia P, Sanchez, Sonia, Moliner Cubel, Britta, Nyboer, Monika, Jankowska-Döllken, Christine, Schaeffer-Reiss, Daniel, Ayoub, Gabrielle, Planelles, and Michael, Lanzer
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Antimalarials ,Kinetics ,Parasitic Sensitivity Tests ,parasitic diseases ,Plasmodium falciparum ,Drug Resistance ,Protozoan Proteins ,Serine ,Membrane Transport Proteins ,Chloroquine ,Phosphorylation ,Microbiology - Abstract
The chloroquine resistance transporter PfCRT of the human malaria parasite Plasmodium falciparum confers resistance to the former first-line antimalarial drug chloroquine, and it modulates the responsiveness to a wide range of quinoline and quinoline-like compounds. PfCRT is post-translationally modified by phosphorylation, palmitoylation, and, possibly, ubiquitination. However, the impact of these post-translational modifications on P. falciparum biology and, in particular, the drug resistance–conferring activity of PfCRT has remained elusive. Here, we confirm phosphorylation at Ser-33 and Ser-411 of PfCRT of the chloroquine-resistant P. falciparum strain Dd2 and show that kinase inhibitors can sensitize drug responsiveness. Using CRISPR/Cas9 genome editing to generate genetically engineered PfCRT variants in the parasite, we further show that substituting Ser-33 with alanine reduced chloroquine and quinine resistance by ∼50% compared with the parental P. falciparum strain Dd2, whereas the phosphomimetic amino acid aspartic acid could fully and glutamic acid could partially reconstitute the level of chloroquine/quinine resistance. Transport studies conducted in the parasite and in PfCRT-expressing Xenopus laevis oocytes linked phosphomimetic substitution at Ser-33 to increased transport velocity. Our data are consistent with phosphorylation of Ser-33 relieving an autoinhibitory intramolecular interaction within PfCRT, leading to a stimulated drug transport activity. Our findings shed additional light on the function of PfCRT and suggest that chloroquine could be reevaluated as an antimalarial drug by targeting the kinase in P. falciparum that phosphorylates Ser-33 of PfCRT.
- Published
- 2019
15. An essential role for α4A-tubulin in platelet biogenesis
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Alain Van Dorsselaer, Claude Férec, Arnaud Dupuis, Benjamin T. Kile, Christine Schaeffer-Reiss, Carsten Janke, Morgane Batzenschlager, Irina Pleines, François Lanza, Paul Gueguen, Léa Mallo, Anita Eckly, Maria M. Magiera, Jean Marc Strub, Catherine Strassel, Christian Gachet, Sylvie Moog, Jean-Yves Py, Agnès Hovasse, Quentin Kimmerlin, Stephane Chappaz, Natarajan Kathiresan, Biologie et pharmacologie des plaquettes sanguines: hémostase, thrombose, transfusion, Université de Strasbourg (UNISTRA)-EFS-Institut National de la Santé et de la Recherche Médicale (INSERM), Institut Curie [Paris], Stress génotoxiques et cancer, Université Paris-Sud - Paris 11 (UP11)-Institut Curie [Paris]-Centre National de la Recherche Scientifique (CNRS), Laboratoire de chimie de coordination (LCC), Institut National Polytechnique (Toulouse) (Toulouse INP), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Institut de Chimie de Toulouse (ICT-FR 2599), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Centre National de la Recherche Scientifique (CNRS)-Institut de Recherche pour le Développement (IRD)-Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Institut de Recherche pour le Développement (IRD)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), Laboratoire d'Analyses de Biologie Médicale (LABM), Institut National de la Santé et de la Recherche Médicale (INSERM)-EFS Alsace, 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), Génétique moléculaire et génétique épidémiologique, Université de Brest (UBO)-Institut National de la Santé et de la Recherche Médicale (INSERM), Division of Developmental Immunology (DKBW), University of Basel (Unibas), Etablissement Français du Sang Bretagne, EFS, Institut de Chimie de Toulouse (ICT), Institut de Recherche pour le Développement (IRD)-Université Toulouse III - Paul Sabatier (UT3), Université de Toulouse (UT)-Université de Toulouse (UT)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Institut National Polytechnique (Toulouse) (Toulouse INP), Université de Toulouse (UT)-Institut de Recherche pour le Développement (IRD)-Université Toulouse III - Paul Sabatier (UT3), Université de Toulouse (UT)-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)-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 de Chimie de Toulouse (ICT-FR 2599), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Institut National Polytechnique (Toulouse) (Toulouse INP), Université Fédérale Toulouse Midi-Pyrénées-Institut de Recherche pour le Développement (IRD)-Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Centre National de la Recherche Scientifique (CNRS), 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)
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Blood Platelets ,Male ,0301 basic medicine ,Alkylating Agents ,Health, Toxicology and Mutagenesis ,Megakaryocyte differentiation ,Mutation, Missense ,Antigens, CD34 ,Plant Science ,macromolecular substances ,[SDV.BC.BC]Life Sciences [q-bio]/Cellular Biology/Subcellular Processes [q-bio.SC] ,Microtubules ,Corrections ,Biochemistry, Genetics and Molecular Biology (miscellaneous) ,Thrombopoiesis ,Mice ,03 medical and health sciences ,0302 clinical medicine ,Megakaryocyte ,Tubulin ,Microtubule ,[SDV.BC.IC]Life Sciences [q-bio]/Cellular Biology/Cell Behavior [q-bio.CB] ,medicine ,Animals ,Humans ,Platelet ,[SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM] ,Cells, Cultured ,Mice, Inbred BALB C ,Ecology ,biology ,Platelet Count ,Chemistry ,Correction ,Thrombocytopenia ,Isotype ,Tissue Donors ,3. Good health ,Cell biology ,030104 developmental biology ,medicine.anatomical_structure ,Ethylnitrosourea ,030220 oncology & carcinogenesis ,biology.protein ,Megakaryocytes ,Biogenesis - Abstract
During platelet biogenesis, microtubules (MTs) are arranged into submembranous structures (the marginal band) that encircle the cell in a single plane. This unique MT array has no equivalent in any other mammalian cell, and the mechanisms responsible for this particular mode of assembly are not fully understood. One possibility is that platelet MTs are composed of a particular set of tubulin isotypes that carry specific posttranslational modifications. Although β1-tubulin is known to be essential, no equivalent roles of α-tubulin isotypes in platelet formation or function have so far been reported. Here, we identify α4A-tubulin as a predominant α-tubulin isotype in platelets. Similar to β1-tubulin, α4A-tubulin expression is up-regulated during the late stages of megakaryocyte differentiation. Missense mutations in the α4A-tubulin gene cause macrothrombocytopenia in mice and humans. Defects in α4A-tubulin lead to changes in tubulin tyrosination status of the platelet tubulin pool. Ultrastructural defects include reduced numbers and misarranged MT coils in the platelet marginal band. We further observed defects in megakaryocyte maturation and proplatelet formation inTuba4a-mutant mice. We have, thus, discovered an α-tubulin isotype with specific and essential roles in platelet biogenesis.
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- 2019
16. Use of an innovative system and nanotechnology-based strategy for therapeutic applications of Gla-rich protein (GRP)
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Justine Schneider, Marta S. Rafael, Christine Schaeffer-Reiss, Arnaud Poterszman, António Alves de Matos, Nuna Araújo, Dina C. Simes, Carla Viegas, Anjos L. Macedo, and Evelina Edelweiss
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Vitamin ,Inflammation ,Calcification inhibitor ,business.industry ,Free Communications Biology and Biochemistry ,General Medicine ,Therapeutics ,030204 cardiovascular system & hematology ,medicine.disease ,3. Good health ,Calcification ,03 medical and health sciences ,chemistry.chemical_compound ,0302 clinical medicine ,chemistry ,medicine ,Cancer research ,030212 general & internal medicine ,Extracellular vesicles (EVs) ,medicine.symptom ,business ,hormones, hormone substitutes, and hormone antagonists - Abstract
Introduction: Gla-rich protein (GRP) is a vitamin K-dependent protein (VKDP) acting as a calcification inhibitor and anti-inflammatory agent in cardiovascular and articular systems, and THP1 monocyte/macrophage cells [1,2]. Calcification and inflammation processes are known to be involved in the etiology of several calcification-related chronic inflammatory diseases such as atherosclerosis, CKD and osteoarthritis, in a complex bi-directional interplay that drives disease progression. Here, we developed an innovative system to produce human c-carboxylated GRP (cGRP), and a nanotechnology strategy based on GRP loading into extracellular vesicles (EVs) as a gold standard delivery system for GRP in therapeutic applications. Materials and methods: Human GRP protein was co-expressed with c-carboxylase enzyme (GGCX), vitamin K oxidoreductase (GGCX) and furin, in the insect cell baculovirus system in the presence of vitamin K. GRP released in the cell culture media was characterized by mass spectrometry based techniques and Western blot analysis. EVs released by the insect cells overexpressing GRP were isolated by ultracentrifugation, and characterized for GRP content through TEM-immunogold staining, Western blot, ELISA, qPCR. Functional assays using isolated EVs containing GRP were performed in primary vascular smooth muscle cells (VSMCs) and THP1 monocyte/macrophage cells, for anti-mineralizing and anti-inflammatory screening.Results: GRP released in the cell culture media when co-expressed with GGCX, VKOR and furin in the presence of vitamin K, is processed at the pro-peptide and contain Gla residues. EVs released by the insect cells in this system were shown to be loaded with GRP protein and mRNA, and capable of reducing ECM calcium deposition of calcifying VSMCs and the production of TNFa in THP1 monocyte/macrophage cells stimulated with LPS. Discussion and conclusions: While the successful production of human cGRP constitutes a major achievement, this innovative methodology will open new opportunities for the production of other biological active VKDPs. Furthermore, EVs loaded with GRP were shown to have anti-mineralizing and anti-inflammatory properties, with promising therapeutic potentialities for calcification-related chronic inflammatory diseases. Portuguese Foundation for Science and Technology (EU/PID1003201) info:eu-repo/semantics/publishedVersion
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- 2019
17. Phosphomimetic substitution at Ser-33 of the chloroquine resistance transporter PfCRT reconstitutes drug responses in Plasmodium falciparum
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Michael Lanzer, Cecilia P. Sanchez, Daniel Ayoub, Christine Schaeffer-Reiss, Britta Nyboer, Monika Jankowska-Döllken, Gabrielle Planelles, Sonia Moliner Cubel, Heidelberg University Hospital [Heidelberg], 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), Métabolisme et physiologie rénales (ERL 8228), Centre de Recherche des Cordeliers (CRC (UMR_S_1138 / U1138)), École pratique des hautes études (EPHE), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Université Paris Diderot - Paris 7 (UPD7)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU)-École pratique des hautes études (EPHE), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Université Paris Diderot - Paris 7 (UPD7)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU), Sciences Analytiques et Interactions Ioniques et Biomoléculaires (DSA-IPHC), Centre National de la Recherche Scientifique (CNRS)-Université de Strasbourg (UNISTRA)-Centre National de la Recherche Scientifique (CNRS)-Université de Strasbourg (UNISTRA), and École pratique des hautes études (EPHE)-Université Paris Diderot - Paris 7 (UPD7)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU)-École pratique des hautes études (EPHE)-Université Paris Diderot - Paris 7 (UPD7)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU)
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0301 basic medicine ,Plasmodium ,PfCRT ,kinase inhibitor ,malaria ,Drug resistance ,Pharmacology ,Biochemistry ,virulence factor ,03 medical and health sciences ,Palmitoylation ,Chloroquine ,parasitic diseases ,medicine ,[CHIM]Chemical Sciences ,genome editing ,transport velocity ,[SDV.MP.PAR]Life Sciences [q-bio]/Microbiology and Parasitology/Parasitology ,Molecular Biology ,Quinine ,posttranslational modification ,drug resistance ,030102 biochemistry & molecular biology ,biology ,Kinase ,Chemistry ,phosphorylation ,Plasmodium falciparum ,Transporter ,Cell Biology ,biology.organism_classification ,3. Good health ,030104 developmental biology ,[SDV.SP.PHARMA]Life Sciences [q-bio]/Pharmaceutical sciences/Pharmacology ,Phosphorylation ,medicine.drug - Abstract
International audience; The chloroquine resistance transporter PfCRT of the human malaria parasite Plasmodium falciparum confers resistance to the former first-line antimalarial drug chloroquine, and it modulates the responsiveness to a wide range of quinoline and quinoline-like compounds. PfCRT is post-translationally modified by phosphorylation, palmitoylation, and, possibly, ubiquitination. However, the impact of these post-translational modifications on P. falciparum biology and, in particular, the drug resistance–conferring activity of PfCRT has remained elusive. Here, we confirm phosphorylation at Ser-33 and Ser-411 of PfCRT of the chloroquine-resistant P. falciparum strain Dd2 and show that kinase inhibitors can sensitize drug responsiveness. Using CRISPR/Cas9 genome editing to generate genetically engineered PfCRT variants in the parasite, we further show that substituting Ser-33 with alanine reduced chloroquine and quinine resistance by ∼50% compared with the parental P. falciparum strain Dd2, whereas the phosphomimetic amino acid aspartic acid could fully and glutamic acid could partially reconstitute the level of chloroquine/quinine resistance. Transport studies conducted in the parasite and in PfCRT-expressing Xenopus laevis oocytes linked phosphomimetic substitution at Ser-33 to increased transport velocity. Our data are consistent with phosphorylation of Ser-33 relieving an autoinhibitory intramolecular interaction within PfCRT, leading to a stimulated drug transport activity. Our findings shed additional light on the function of PfCRT and suggest that chloroquine could be reevaluated as an antimalarial drug by targeting the kinase in P. falciparum that phosphorylates Ser-33 of PfCRT.
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- 2019
18. Combinatorial regulation of hepatic cytoplasmic signaling and nuclear transcriptional events by the OGT/REV-ERBα complex
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Agnès Hovasse, Manjula Vinod, Jérôme Eeckhoute, Jeremy Alexandre, Alexandre Berthier, Julie Dubois-Chevalier, Fabrice Bray, Maheul Ploton, Xavier Marechal, Bart Staels, Christine Schaeffer-Reiss, Agata Steenackers, Hélène Duez, Philippe Lefebvre, Geoffrey Porez, Sarah Cianférani, Christian Rolando, Nao Yamakawa, Céline Gheeraert, Tony Lefebvre, Récepteurs nucléaires, maladies cardiovasculaires et diabète - U 1011 (RNMCD), Institut Pasteur de Lille, Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lille-Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille), Unité de Glycobiologie Structurale et Fonctionnelle UMR 8576 (UGSF), Université de Lille-Centre National de la Recherche Scientifique (CNRS), Laboratoire de Spectrométrie de Masse BioOrgan [Université de Strasbourg], Université de Strasbourg (UNISTRA), 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), Miniaturisation pour la Synthèse, l’Analyse et la Protéomique - UAR 3290 (MSAP), Biochimie Structurale et Fonctionnelle des Assemblages Biomoléculaires - CNRS FR3688 (FRABio), Institut Michel Eugène Chevreul - FR 2638 (IMEC), Université d'Artois (UA)-Centrale Lille-Institut de Chimie du CNRS (INC)-Université de Lille-Centre National de la Recherche Scientifique (CNRS)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), This work was supported by Agence Nationale de la Recherche Grants OGlcRev and ANR-10-LABX-46, Fondation pour la Recherche Médicale Grant Equipe labellisée FRM 2015 DEQ20150331724, European Foundation for the Study of Diabetes/Lilly European Diabetes Research Program, European Commission EuRhythDia FP7-health Grant 278397, and French Proteomic Infrastructure Grant ANR-10-INBS-08–03. B.S. is a recipient of Advanced European Council Grant 694717. OGlcNAcylated protein identification was done at 'Plateforme Analyses Glycoconjugués' (FR3688 FRABio, CNRS, Université de Lille)., ANR-10-LABX-0046,EGID,EGID Diabetes Pole(2010), European Project: 278397,EC:FP7:HEALTH,FP7-HEALTH-2011-two-stage,EURHYTHDIA(2011), European Project: 694717,H2020-EU.1.1. - EXCELLENT SCIENCE - European Research Council (ERC) ,ImmunoBile(2016), Inserm, Université de Lille, CHU Lille, CNRS, Récepteurs nucléaires, maladies cardiovasculaires et diabète - U 1011 [RNMCD], Unité de Glycobiologie Structurale et Fonctionnelle UMR 8576 [UGSF], Institut Pluridisciplinaire Hubert Curien [IPHC], Laboratoire de Spectrométrie de Masse BioOrganique [Strasbourg] [LSMBO], Miniaturisation pour la Synthèse, l’Analyse et la Protéomique - UAR 3290 [MSAP], Récepteurs nucléaires, maladies cardiovasculaires et diabète (EGID), Université de Lille, Droit et Santé-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut Pasteur de Lille, Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP)-Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille), Unité de Glycobiologie Structurale et Fonctionnelle - UMR 8576 (UGSF), Université de Lille-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Recherche Agronomique (INRA), Université de Strasbourg (UNISTRA)-Centre National de la Recherche Scientifique (CNRS), Miniaturisation pour l'Analyse, la Synthèse & la Protéomique (USR CNRS 3290), Centre National de la Recherche Scientifique (CNRS), Fédération de Recherche Biochimie Structurale et Fonctionnelle des Assemblages Biomoléculaires (FRABio - CNRS FR3688), Institut Michel Eugène Chevreul [Villeneuve d’Ascq] (CNRS FR 2638), Centre National de la Recherche Scientifique (CNRS)-Université de Lille, ANR-10-LABX-0046/10-LABX-0046,EGID,EGID Diabetes Pole(2010), Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP)-Université de Lille-Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille)-Institut National de la Santé et de la Recherche Médicale (INSERM), Institut National de la Recherche Agronomique (INRA)-Université de Lille-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), Miniaturisation pour la Synthèse, l’Analyse et la Protéomique - USR 3290 (MSAP), Centre National de la Recherche Scientifique (CNRS)-Université de Lille-Institut de Chimie du CNRS (INC), Université d'Artois (UA)-Institut de Chimie du CNRS (INC)-Université de Lille-Centre National de la Recherche Scientifique (CNRS)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE)-Centrale Lille Institut (CLIL), Derudas, Marie-Hélène, EGID Diabetes Pole - - EGID2010 - ANR-10-LABX-0046 - LABX - VALID, Chronotherapeutic lifestyle intervention for diabetes and obesity to reset the circadian rhythm and improve cardiometabolic risk in the European working population - EURHYTHDIA - - EC:FP7:HEALTH2011-10-01 - 2017-06-30 - 278397 - VALID, Bile acid, immune-metabolism, lipid and glucose homeostasis - ImmunoBile - - H2020-EU.1.1. - EXCELLENT SCIENCE - European Research Council (ERC) 2016-09-01 - 2021-08-31 - 694717 - VALID, 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)
- Subjects
0301 basic medicine ,viruses ,Circadian clock ,N-Acetylglucosaminyltransferases ,Biochemistry ,Mice ,03 medical and health sciences ,O-GlcNAcylation ,REV-ERBα ,Cell Line, Tumor ,Circadian Clocks ,Gene expression ,[SDV.BBM] Life Sciences [q-bio]/Biochemistry, Molecular Biology ,Animals ,Humans ,Insulin ,[SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology ,Phosphorylation ,Nuclear protein ,Protein kinase B ,Mice, Knockout ,Multidisciplinary ,biology ,Kinase ,Chemistry ,Hep G2 Cells ,Biological Sciences ,Lipid Metabolism ,Cell biology ,Mice, Inbred C57BL ,Insulin receptor ,Glucose ,HEK293 Cells ,030104 developmental biology ,PNAS Plus ,Gene Expression Regulation ,Liver ,Nuclear receptor ,Regulatory sequence ,epigenomics ,metabolism ,Signal Transduction ,Nuclear Receptor Subfamily 1, Group D, Member 1 ,biology.protein ,Sterol Regulatory Element Binding Protein 1 ,Proto-Oncogene Proteins c-akt ,signal transduction - Abstract
Significance Using an interactomic approach, we have identified the nuclear receptor REV-ERBα as a O-GlcNAc transferase (OGT) protein partner. REV-ERBα protects cytoplasmic OGT from proteasomal degradation and facilitates cytosolic and nuclear protein O-GlcNAcylation while REV-ERα ligands decreased cytoplasmic OGT activity. REV-ERBα thus exerts pleiotropic activities through OGT, coordinating signal transduction, epigenomic programming, and transcriptional response in the liver., The nuclear receptor REV-ERBα integrates the circadian clock with hepatic glucose and lipid metabolism by nucleating transcriptional comodulators at genomic regulatory regions. An interactomic approach identified O-GlcNAc transferase (OGT) as a REV-ERBα−interacting protein. By shielding cytoplasmic OGT from proteasomal degradation and favoring OGT activity in the nucleus, REV-ERBα cyclically increased O-GlcNAcylation of multiple cytoplasmic and nuclear proteins as a function of its rhythmically regulated expression, while REV-ERBα ligands mostly affected cytoplasmic OGT activity. We illustrate this finding by showing that REV-ERBα controls OGT-dependent activities of the cytoplasmic protein kinase AKT, an essential relay in insulin signaling, and of ten-of-eleven translocation (TET) enzymes in the nucleus. AKT phosphorylation was inversely correlated to REV-ERBα expression. REV-ERBα enhanced TET activity and DNA hydroxymethylated cytosine (5hmC) levels in the vicinity of REV-ERBα genomic binding sites. As an example, we show that the REV-ERBα/OGT complex modulates SREBP-1c gene expression throughout the fasting/feeding periods by first repressing AKT phosphorylation and by epigenomically priming the Srebf1 promoter for a further rapid response to insulin. Conclusion: REV-ERBα regulates cytoplasmic and nuclear OGT-controlled processes that integrate at the hepatic SREBF1 locus to control basal and insulin-induced expression of the temporally and nutritionally regulated lipogenic SREBP-1c transcript.
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- 2018
19. An evolutionary conserved zinc finger protein is involved in Toxoplasma gondii mRNA nuclear export
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Mathieu, Gissot, Agnès, Hovasse, Laurent, Chaloin, Christine, Schaeffer-Reiss, Alain, Van Dorsselaer, Stanislas, Tomavo, Centre d’Infection et d’Immunité de Lille - INSERM U 1019 - UMR 9017 - UMR 8204 (CIIL), Centre National de la Recherche Scientifique (CNRS)-Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille)-Université de Lille-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut Pasteur de Lille, Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP), 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), Institut de Recherche en Infectiologie de Montpellier (IRIM), Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS), This work was supported by Centre National de la Recherche Scientifique (CNRS), Institut National de la Santé et de la Recherche Médicale (INSERM), a grant from the French National Research Agency (ANR) [grant number ANR‐13‐JSV3‐0006‐01 to MG] and the Laboratoire d'Excellence (LabEx) ParaFrap [ANR‐11‐LABX‐0024 to ST]., ANR-11-LABX-0024,ParaFrap,Alliance française contre les maladies parasitaires(2011), Institut Pasteur de Lille, Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lille-Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille)-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)
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Gene Knockdown Techniques ,Genetic Complementation Test ,Plasmodium falciparum ,parasitic diseases ,Active Transport, Cell Nucleus ,Kruppel-Like Transcription Factors ,CYS2-HIS2 Zinc Fingers ,Humans ,RNA, Viral ,[CHIM]Chemical Sciences ,Cell Cycle Checkpoints ,RNA, Messenger ,Toxoplasma - Abstract
International audience; Apicomplexan parasites are responsible for some of the most deadly parasitic diseases affecting humans and livestock. There is an urgent need for new medicines that will target apicomplexan-specific pathways. We characterized a Toxoplasma gondii C2H2 zinc finger protein, named TgZNF2, which is conserved among eukaryotes. We constructed an inducible KO strain (iKO-TgZNF2) for this gene where the tgznf2 gene expression is repressed in the presence of a tetracycline analog (ATc). We showed that the iKO-TgZNF2 parasites are unable to proliferate after depletion of the TgZNF2 protein. Complementation with a full length copy of the gene restores the phenotype Moreover, the homolog of this protein in the related apicomplexan Plasmodium falciparum was shown to efficiently rescue the phenotype, suggesting that this pathway is likely conserved among apicomplexan parasites. We demonstrated that the iKO-mutant lacking TgZNF2 are arrested during the cell cycle during the G1 phase. We identified potential protein partners of this protein among which are spliceosomal complex and mRNA nuclear export components. We confirmed that TgZNF2 is able to bind in vivo to transcripts but splicing is not perturbed in the ATc-treated parasites. Instead, we demonstrated that TgZNF2 depletion leads to the sequestration of polyA+ mRNAs in the nucleus while ribosomal RNAs are not affected. We discovered a conserved protein with specific apicomplexan functional properties that is essential for the survival of T. gondii. TgZNF2 may be crucial to ensure the correct polyA+ mRNA nuclear export, a function that is conserved in P. falciparum.
- Published
- 2017
20. Flowering as the Most Highly Sensitive Period of Grapevine (Vitis vinifera L. cv Mourvèdre) to the Botryosphaeria Dieback Agents Neofusicoccum parvum and Diplodia seriata Infection
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Christophe Clément, Alessandro Spagnolo, Alain Van Dorsselaer, Christine Schaeffer-Reiss, Philippe Larignon, Clara Cilindre, Agnès Hovasse, Florence Fontaine, and Maryline Magnin-Robert
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Proteomics ,Proteome ,two dimensional gel electrophoresis ,Diplodia seriata ,Article ,Catalysis ,Veraison ,lcsh:Chemistry ,Inorganic Chemistry ,defense-related proteins ,Ascomycota ,Seriata ,Tandem Mass Spectrometry ,Botany ,Electrophoresis, Gel, Two-Dimensional ,Vitis ,Physical and Theoretical Chemistry ,Botryosphaeria dieback ,lcsh:QH301-705.5 ,Molecular Biology ,Botryosphaeria ,Spectroscopy ,Plant Diseases ,Plant Proteins ,Pathogenesis-related protein ,biology ,Spots ,Organic Chemistry ,plant proteomics ,food and beverages ,General Medicine ,biology.organism_classification ,Neofusicoccum parvum ,Computer Science Applications ,lcsh:Biology (General) ,lcsh:QD1-999 ,Inflorescence ,Viticulture - Abstract
Botryosphaeria dieback is a fungal grapevine trunk disease that currently represents a threat for viticulture worldwide because of the important economical losses due to reduced yield of affected plants and their premature death. Neofusicoccum parvum and Diplodia seriata are among the causal agents. Vine green stems were artificially infected with N. parvum or D. seriata at the onset of three different phenological stages (G stage (separated clusters), flowering and veraison). Highest mean lesion lengths were recorded at flowering. Major proteome changes associated to artificial infections during the three different phenological stages were also reported using two dimensional gel electrophoresis (2D)-based analysis. Twenty (G stage), 15 (flowering) and 13 (veraison) differentially expressed protein spots were subjected to nanoLC-MS/MS and a total of 247, 54 and 25 proteins were respectively identified. At flowering, a weaker response to the infection was likely activated as compared to the other stages, and some defense-related proteins were even down regulated (e.g., superoxide dismutase, major latex-like protein, and pathogenesis related protein 10). Globally, the flowering period seemed to represent the period of highest sensitivity of grapevine to Botryosphaeria dieback agent infection, possibly being related to the high metabolic activity in the inflorescences.
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- 2014
21. H2B ubiquitylation modulates spliceosome assembly and function in budding yeast
- Author
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Catherine Dargemont, Lucas Hérissant, Christine Guthrie, Diego Bertaccini, Christine Schaeffer-Reiss, Erica A. Moehle, and Alain Van Dorsselaer
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Genetics ,Spliceosome ,biology ,Exonic splicing enhancer ,Cell Biology ,General Medicine ,Methylation ,Chromatin ,Cell biology ,Histone ,RNA splicing ,biology.protein ,snRNP ,Chromatin immunoprecipitation - Abstract
Background information Commitment to splicing occurs co-transcriptionally, but a major unanswered question is the extent to which various modifications of chromatin, the template for transcription in vivo, contribute to the regulation of splicing. Results Here, we perform genome-wide analyses showing that inhibition of specific marks – H2B ubiquitylation, H3K4 methylation and H3K36 methylation – perturbs splicing in budding yeast, with each modification exerting gene-specific effects. Furthermore, semi-quantitative mass spectrometry on purified nuclear mRNPs and chromatin immunoprecipitation analysis on intron-containing genes indicated that H2B ubiquitylation, but not Set1-, Set2- or Dot1-dependent H3 methylation, stimulates recruitment of the early splicing factors, namely U1 and U2 snRNPs, onto nascent RNAs. Conclusions These results suggest that histone modifications impact splicing of distinct subsets of genes using distinct pathways.
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- 2014
22. Trafficking of the exported P. falciparum chaperone PfHsp70x
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Paul R. Gilson, Sarah C. Charnaud, Michael Lanzer, Simone Külzer, Jude M. Przyborski, Cecilia G. Sanchez, Christine Schaeffer-Reiss, Brendan S. Crabb, Benoit Westermann, Maja Strecker, Sebastian Müller, Verena Bittl, Manuel Rhiel, and Anke Tribensky
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0301 basic medicine ,Erythrocytes ,Amino Acid Motifs ,Plasmodium falciparum ,030106 microbiology ,Protozoan Proteins ,Article ,03 medical and health sciences ,medicine ,Humans ,HSP70 Heat-Shock Proteins ,Site-directed mutagenesis ,Multidisciplinary ,biology ,Translocon ,Cell biology ,Hsp70 ,Transport protein ,Protein Transport ,Red blood cell ,Cytosol ,030104 developmental biology ,medicine.anatomical_structure ,Biochemistry ,Acetylation ,Chaperone (protein) ,biology.protein - Abstract
Plasmodium falciparum extensively modifies its chosen host cell, the mature human erythrocyte. This remodelling is carried out by parasite-encoded proteins that are exported into the host cell. To gain access to the human red blood cell, these proteins must cross the parasitophorous vacuole, a membrane bound compartment surrounding the parasite that is generated during the invasion process. Many exported proteins carry a so-called PEXEL/HT signal that directs their transport. We recently reported the unexpected finding of a species-restricted parasite-encoded Hsp70, termed PfHsp70x, which is exported into the host erythrocyte cytosol. PfHsp70x lacks a classical PEXEL/HT motif, and its transport appears to be mediated by a 7 amino acid motif directly following the hydrophobic N-terminal secretory signal. In this report, we analyse this short targeting sequence in detail. Surprisingly, both a reversed and scrambled version of the motif retained the capacity to confer protein export. Site directed mutagenesis of glutamate residues within this region leads to a block of protein trafficking within the lumen of the PV. In contrast to PEXEL-containing proteins, the targeting signal is not cleaved, but appears to be acetylated. Furthermore we show that, like other exported proteins, trafficking of PfHsp70x requires the vacuolar translocon, PTEX.
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- 2016
23. Hemoglobin Kirklareli (α H58L), a New Variant Associated with Iron Deficiency and Increased CO Binding
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Tchilabalo Dilezitoko Alayi, Emmanuel Bissé, Thomas Epting, Karl Winkler, John S. Olson, Christine Schaeffer-Reiss, Premila P. Samuel, Andres S. Benitez Cardenas, Ivan Birukou, Jayashree Soman, Alain Van Dorsselaer, Freiburg University Medical Center, 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), Rice University [Houston], and 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)
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0301 basic medicine ,Adult ,Male ,[SDV]Life Sciences [q-bio] ,Hemoglobins, Abnormal ,Mutant ,Static Electricity ,Crystallography, X-Ray ,Biochemistry ,Mass Spectrometry ,law.invention ,03 medical and health sciences ,chemistry.chemical_compound ,Young Adult ,law ,Catalytic Domain ,Humans ,Denaturation (biochemistry) ,Molecular Biology ,Heme ,Chromatography, High Pressure Liquid ,Carbon Monoxide ,Chromatography, Reverse-Phase ,biology ,Red Cell ,Anemia, Iron-Deficiency ,Active site ,Molecular Bases of Disease ,Cell Biology ,Chromatography, Ion Exchange ,3. Good health ,Oxygen ,030104 developmental biology ,chemistry ,biology.protein ,Recombinant DNA ,Female ,Hemoglobin ,Oxidation-Reduction ,Hemin - Abstract
International audience; Mutations in hemoglobin can cause a wide range of pheno-typic outcomes, including anemia due to protein instability and red cell lysis. Uncovering the biochemical basis for these phenotypes can provide new insights into hemoglobin structure and function as well as identify new therapeutic opportunities. We report here a new hemoglobin ␣ chain variant in a female patient with mild anemia, whose father also carries the trait and is from the Turkish city of Kirklareli. Both the patient and her father had a His-58(E7) 3 Leu mutation in ␣1. Surprisingly, the patient's father is not anemic, but he is a smoker with high levels of HbCO (ϳ16%). To understand these phenotypes, we examined recombinant human Hb (rHb) Kirklareli containing the ␣ H58L replacement. Mutant ␣ subunits containing Leu-58(E7) autoxidize ϳ8 times and lose hemin ϳ200 times more rapidly than native ␣ subunits, causing the oxygenated form of rHb Kirklareli to denature very rapidly under physiological conditions. The crystal structure of rHb Kirklareli shows that the ␣ H58L replacement creates a completely apolar active site, which prevents electrostatic stabilization of bound O 2 , promotes autoxidation, and enhances hemin dissociation by inhibiting water coordination to the Fe(III) atom. At the same time, the mutant ␣ subunit has an ϳ80,000-fold higher affinity for CO than O 2 , causing it to rapidly take up and retain carbon monoxide , which prevents denaturation both in vitro and in vivo and explains the phenotypic differences between the father, who is a smoker, and his daughter.
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- 2016
24. Unconventional endosome-like compartment and retromer complex in Toxoplasma gondii govern parasite integrity and host infection
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Mohamed-Ali Hakimi, Frank Lafont, Elisabeth Werkmeister, Fabien Sindikubwabo, Lamba Omar Sangaré, Alain Van Dorsselaer, Tchilabalo Dilezitoko Alayi, Christian Slomianny, Isabelle Callebaut, Agnès Hovasse, Christine Schaeffer-Reiss, Benoit Westermann, Stanislas Tomavo, Centre d’Infection et d’Immunité de Lille - INSERM U 1019 - UMR 9017 - UMR 8204 (CIIL), Institut Pasteur de Lille, Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lille-Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille)-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), Protéomique et Peptides Modifiés (P3M), Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lille-Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille)-Centre National de la Recherche Scientifique (CNRS)-Institut Pasteur de Lille, Institute for Advanced Biosciences / Institut pour l'Avancée des Biosciences (Grenoble) (IAB), Centre Hospitalier Universitaire [Grenoble] (CHU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Etablissement français du sang - Auvergne-Rhône-Alpes (EFS)-Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019]), Institut de minéralogie, de physique des matériaux et de cosmochimie (IMPMC), Muséum national d'Histoire naturelle (MNHN)-Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut de recherche pour le développement [IRD] : UR206-Centre National de la Recherche Scientifique (CNRS), Institut de biologie de Lille - UMS 3702 (IBL), Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP)-Université de Lille-Centre National de la Recherche Scientifique (CNRS), Laboratoire de Physiologie Cellulaire : Canaux ioniques, inflammation et cancer - U 1003 (PHYCELL), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lille, Centre National de la Recherche Scientifique (CNRS)-Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille)-Université de Lille-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut Pasteur de Lille, Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP), Université de Strasbourg (UNISTRA)-Institut National de Physique Nucléaire et de Physique des Particules du CNRS (IN2P3)-Centre National de la Recherche Scientifique (CNRS), Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP)-Centre National de la Recherche Scientifique (CNRS)-Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille)-Université de Lille-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut Pasteur de Lille, Laboratoire Adaptation et pathogénie des micro-organismes [Grenoble] (LAPM), Centre National de la Recherche Scientifique (CNRS)-Université Joseph Fourier - Grenoble 1 (UJF), Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut de recherche pour le développement [IRD] : UR206-Muséum national d'Histoire naturelle (MNHN)-Centre National de la Recherche Scientifique (CNRS), Bioimaging platform (IBL), Université de Lille, HAL UPMC, Gestionnaire, and Université Joseph Fourier - Grenoble 1 (UJF)-Centre National de la Recherche Scientifique (CNRS)
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0301 basic medicine ,Genetics ,Multidisciplinary ,Retromer ,Endosome ,Science ,[SDV]Life Sciences [q-bio] ,General Physics and Astronomy ,Endocytic recycling ,General Chemistry ,Biology ,General Biochemistry, Genetics and Molecular Biology ,Transmembrane protein ,Major facilitator superfamily ,3. Good health ,Cell biology ,Retromer complex ,[SDV] Life Sciences [q-bio] ,03 medical and health sciences ,030104 developmental biology ,parasitic diseases ,Organelle biogenesis ,Biogenesis - Abstract
Membrane trafficking pathways play critical roles in Apicomplexa, a phylum of protozoan parasites that cause life-threatening diseases worldwide. Here we report the first retromer-trafficking interactome in Toxoplasma gondii. This retromer complex includes a trimer Vps35–Vps26–Vps29 core complex that serves as a hub for the endosome-like compartment and parasite-specific proteins. Conditional ablation of TgVps35 reveals that the retromer complex is crucial for the biogenesis of secretory organelles and for maintaining parasite morphology. We identify TgHP12 as a parasite-specific and retromer-associated protein with functions unrelated to secretory organelle formation. Furthermore, the major facilitator superfamily homologue named TgHP03, which is a multiple spanning and ligand transmembrane transporter, is maintained at the parasite membrane by retromer-mediated endocytic recycling. Thus, our findings highlight that both evolutionarily conserved and unconventional proteins act in concert in T. gondii by controlling retrograde transport that is essential for parasite integrity and host infection.
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- 2016
25. Toxoplasma Sortilin-like Receptor Regulates Protein Transport and Is Essential for Apical Secretory Organelle Biogenesis and Host Infection
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Christine Schaeffer-Reiss, Tchilabalo Dilezitoko Alayi, Alain Van Dorsselear, Agnès Hovasse, Isabelle Callebaut, Rajshekhar Y. Gaji, Pierre-Julien Sloves, Thomas Mouveaux, Vern B. Carruthers, Stéphane Delhaye, Elisabeth Werkmeister, Christian Slomianny, Stanislas Tomavo, Centre d’Infection et d’Immunité de Lille - INSERM U 1019 - UMR 9017 - UMR 8204 (CIIL), Institut Pasteur de Lille, Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lille-Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille)-Centre National de la Recherche Scientifique (CNRS), Laboratoire de Physiologie Cellulaire : Canaux ioniques, inflammation et cancer - U 1003 (PHYCELL), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lille, 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)-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), Institut de minéralogie et de physique des milieux condensés (IMPMC), Université Pierre et Marie Curie - Paris 6 (UPMC)-Université Paris Diderot - Paris 7 (UPD7)-Institut de Physique du Globe de Paris (IPG Paris)-Centre National de la Recherche Scientifique (CNRS), Department of Microbiology and Immunology, University of Michigan Medical School, University of Michigan [Ann Arbor], University of Michigan System-University of Michigan System, Centre National de la Recherche Scientifique (CNRS)-Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille)-Université de Lille-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut Pasteur de Lille, Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP), 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), and Université Pierre et Marie Curie - Paris 6 (UPMC)-IPG PARIS-Université Paris Diderot - Paris 7 (UPD7)-Centre National de la Recherche Scientifique (CNRS)
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Cancer Research ,Biology ,Models, Biological ,Microbiology ,Microneme ,Mice ,03 medical and health sciences ,0302 clinical medicine ,Virology ,Immunology and Microbiology(all) ,Protein Interaction Mapping ,Animals ,Humans ,Molecular Biology ,Cells, Cultured ,030304 developmental biology ,Organelles ,Mice, Inbred BALB C ,0303 health sciences ,Rhoptry ,Membrane Proteins ,Signal transducing adaptor protein ,Survival Analysis ,Transmembrane protein ,3. Good health ,Cell biology ,Transport protein ,Adaptor Proteins, Vesicular Transport ,Protein Transport ,Toxoplasmosis, Animal ,Cytoplasm ,Female ,Parasitology ,Organelle biogenesis ,Toxoplasma ,030217 neurology & neurosurgery ,Biogenesis ,Protein Binding - Abstract
International audience; Apicomplexan parasites have an assortment of unique apical secretory organelles (rhoptries and micronemes), which have crucial functions in host infection. Here, we show that a Toxoplasma gondii sortilin-like receptor (TgSORTLR) is required for the subcellular localization and formation of apical secretory organelles.TgSORTLR is a transmembrane protein that resides within Golgi-endosomal related compartments. The lumenal domain specifically interacts with rhoptry and microneme proteins, while the cytoplasmic tail of TgSORTLR recruits cytosolic sorting machinery involved in anterograde and retrograde protein transport. Ectopic expression of the N-terminal TgSORTLR lumenal domain results in dominant negative effects with the mislocalization of both endogenous TgSORTLR as well as rhoptry and microneme proteins. Conditional ablation of TgSORTLR disrupts rhoptry and microneme biogenesis, inhibits parasite motility, and blocks both invasion into and egress from host cells. Thus, the sortilin-like receptor is essential for protein trafficking and the biogenesis of key secretory organelles in Toxoplasma.
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- 2012
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26. The Circulating Protease Persephone Is an Immune Sensor for Microbial Proteolytic Activities Upstream of the Drosophila Toll Pathway
- Author
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Nicolas Matt, Najwa Issa, Florian Veillard, Emilie Lauret, Jean-Marc Reichhart, Alain Van Dorsselaer, Christine Schaeffer-Reiss, Nina Guillaumot, Modèles Insectes de l'Immunité Innée (M3I), 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)-Université de Strasbourg (UNISTRA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Institut Pluridisciplinaire Hubert Curien (IPHC), and Institut National de Physique Nucléaire et de Physique des Particules du CNRS (IN2P3)-Université de Strasbourg (UNISTRA)-Centre National de la Recherche Scientifique (CNRS)
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Male ,0301 basic medicine ,Proteases ,medicine.medical_treatment ,virulence factors ,Virulence ,Immune receptor ,danger signal ,Article ,Virulence factor ,03 medical and health sciences ,0302 clinical medicine ,medicine ,[CHIM]Chemical Sciences ,Animals ,Drosophila Proteins ,Beauveria ,Receptors, Immunologic ,innate immunity ,Molecular Biology ,NF-κB pathway ,Serine protease ,bacterial proteases ,Protease ,Innate immune system ,cysteine cathepsin ,immune receptor ,biology ,Serine Endopeptidases ,Toll-Like Receptors ,Pattern recognition receptor ,cathepsin 26-29-p ,fungal proteases ,Cell Biology ,Immunity, Innate ,Cell biology ,Drosophila melanogaster ,030104 developmental biology ,clip-serine protease ,Proteolysis ,biology.protein ,Female ,Serine Proteases ,030217 neurology & neurosurgery ,Signal Transduction - Abstract
Summary Microbial or endogenous molecular patterns as well as pathogen functional features can activate innate immune systems. Whereas detection of infection by pattern recognition receptors has been investigated in details, sensing of virulence factors activities remains less characterized. In Drosophila, genetic evidences indicate that the serine protease Persephone belongs to a danger pathway activated by abnormal proteolytic activities to induce Toll signaling. However, neither the activation mechanism of this pathway nor its specificity has been determined. Here, we identify a unique region in the pro-domain of Persephone that functions as bait for exogenous proteases independently of their origin, type, or specificity. Cleavage in this bait region constitutes the first step of a sequential activation and licenses the subsequent maturation of Persephone to the endogenous cysteine cathepsin 26-29-p. Our results establish Persephone itself as an immune receptor able to sense a broad range of microbes through virulence factor activities rather than molecular patterns., Graphical Abstract, Highlights • All pathogen-secreted proteases activate the danger-sensing arm of the Toll pathway • The protease Persephone is the immune sensor for microbial proteolytic activities • A sensitive region in Persephone zymogen functions as a bait for exogenous proteases • Bait-region hydrolysis primes maturation of Persephone by the host cathepsin 26-29-p, Innate immune systems are activated by microbial molecular patterns or pathogen functional features. Issa et al. show that the Drosophila Toll pathway senses pathogen proteases through a hydrolysis-sensitive region localized in the Persephone pro-domain. Cleavage of this bait region primes maturation of Persephone and activation of the pathway by the host cathepsin 26-29-p.
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- 2018
27. Characterization of Soluble and Membrane-Bound Proteins of Toxoplasma gondii as Diagnostic Markers of Infection
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Benoit Westermann, Aïda Bouratbine, F. Saghrouni, Alain Van Dorsselaer, Sami Lakhal, Moncef Ben Said, Alia Benkahla, Christine Schaeffer Reiss, and Imen Khammari
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Gel electrophoresis ,biology ,ved/biology ,ved/biology.organism_classification_rank.species ,Acquired Toxoplasmosis ,Toxoplasma gondii ,Diagnostic marker ,biology.organism_classification ,medicine.disease ,Molecular biology ,Toxoplasmosis ,Homology (biology) ,Hammondia hammondi ,Antigen ,medicine - Abstract
In the present study, we applied the combination of one-dimensional gel electrophoresis, immunoblot and nanoliquid chromatography-tandem mass spectrometry (nanoLC-MS/MS) to identify potential immunogenic proteins of Toxoplasma gondii tachyzoites that can be used for the development of reliable assays in the serodiagnosis of acquired toxoplasmosis in immunocompetent subjects. For this purpose, we developed an immunoblot using soluble and membrane extracts of GT1 Toxoplasma gondii tachyzoites and tested 194 positive and 100 negative sera obtained from pregnant women. Five bands of soluble antigens (98 kDa, 36 kDa, 33 kDa, 32 kDa and 21 kDa) and 4 bands of membrane antigens (41 kDa, 35 kDa, 32 kDa and 30 kDa) were selected as the most valuable in terms of sensitivity and specificity. Among these bands, only 2 bands of soluble antigen (33 kDa and 32 kDa) and 2 bands of membrane antigen (32 kDa and 30 kDa) showed a specificity ≥ 90%. After mass spectrometry and bioinformatics analysis, 7 proteins were identified as potential markers for serodiagnosis of toxoplasmosis. These proteins are: SRS34A, GRA7, GRA1, DG32, MIC5, ROP5 and Toxofilin. These proteins showed a 86% to 100% homology with proteins of both VEG and ME49 strains of T. gondii and a 58% to 87% homology with Hammondia hammondi; and can be considered as attractive candidates for the development of an immunochromatography test that can be used for the rapid diagnosis of toxoplasmosis and as a confirmatory test when routine techniques give equivocal results.
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- 2015
28. H2B ubiquitylation modulates spliceosome assembly and function in budding yeast
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Lucas, Hérissant, Erica A, Moehle, Diego, Bertaccini, Alain, Van Dorsselaer, Christine, Schaeffer-Reiss, Christine, Guthrie, and Catherine, Dargemont
- Subjects
Histones ,Spliceosomes ,Ubiquitination ,Saccharomyces cerevisiae ,Article - Abstract
Commitment to splicing occurs co-transcriptionally, but a major unanswered question is the extent to which various modifications of chromatin, the template for transcription in vivo, contribute to the regulation of splicing.Here, we perform genome-wide analyses showing that inhibition of specific marks - H2B ubiquitylation, H3K4 methylation and H3K36 methylation - perturbs splicing in budding yeast, with each modification exerting gene-specific effects. Furthermore, semi-quantitative mass spectrometry on purified nuclear mRNPs and chromatin immunoprecipitation analysis on intron-containing genes indicated that H2B ubiquitylation, but not Set1-, Set2- or Dot1-dependent H3 methylation, stimulates recruitment of the early splicing factors, namely U1 and U2 snRNPs, onto nascent RNAs.These results suggest that histone modifications impact splicing of distinct subsets of genes using distinct pathways.
- Published
- 2014
29. Selected Expression and Functional Importance of α4a-Tubulin in Platelet Biogenesis
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Sylvie Moog, Catherine Strassel, Carsten Janke, François Lanza, Christian Gachet, Christine Schaeffer-Reiss, Morgane Batzenschlager, Magda Mageira, Benjamin T. Kile, and Agnès Hovasse
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Gene isoform ,Immunology ,Cell Biology ,Hematology ,Biology ,Biochemistry ,Isotype ,Cell biology ,Tubulin ,Microtubule ,biology.protein ,Platelet ,Biogenesis ,Megakaryopoiesis ,Megakaryocytopoiesis - Abstract
Platelets are produced from mature megakaryocytes (MK) following a profound cellular reorganization. This includes the assembly of microtubules (MT) into a unique submenbranous coiled structure, the marginal band (MB). This process is thought to depend on a specific αβ-tubulin isotype repertoire. The MK-restricted-β1-tubulin, the predominant isoform of the MB, is already known to be important for platelet biogenesis but the implication of other isotypes is currently unknown. Our goal was to establish the αβ-tubulin repertoire in platelets and during megakaryopoiesis and to evaluate the implication of selected isotypes in platelet formation. To establish an exhaustive list of the tubulin isotypes, we used combination of RT PCR and proteomic analyses to quantify the expression of each isotype in human platelets and in human MK differentiated in culture from CD34+ hematopoietic progenitors. Information gained on the hierarchical combination of tubulin isoforms in the course of platelet biogenesis has been extended at the functional level to investigate both their role in marginal band formation and platelet functions β6-, β5- and α1c-tubulin transcripts were already present in CD34+ cells and decreased during the final stages of megakaryopoiesis. On the other hand, β1-, α4A- and α8-tubulin transcripts were only observed later during MK differentiation and in platelets. Quantitative LC-SRM mass spectrometry confirmed the predominant expression of β1 and α4A-isotypes in platelets. A functional role of the newly identified α4a-tubulin was supported by the thrombocytopenia and enlarged platelets with a decreased number of MT coils (1-3) comprising less-acetylated tubulin in mice carrying a point mutation in tuba4a. Additionally, a tendency to increased responses to several agonists was observed in these platelets. This study reveals new information on the evolution of the tubulin isotype repertoire in platelet formation pointing to a role of less-widely expressed α-isotypes. Disclosures No relevant conflicts of interest to declare.
- Published
- 2016
30. Proteomics profiling reveals novel proteins and functions of the plant stigma exudate
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María Isabel Rodríguez-García, Christine Carapito, Juan David Rejón, Juan de Dios Alché, Krzysztof Zienkiewicz, Antonio J. Castro, Christine Schaeffer-Reiss, François Delalande, Alain Van Dorsselaer, Ministerio de Ciencia e Innovación (España), Junta de Andalucía, European Commission, Centre National de la Recherche Scientifique (France), and Agence Nationale de la Recherche (France)
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Exudate ,Proteomics ,Programmed cell death ,Pollination ,Physiology ,secretome ,Plant Exudates ,Plant Science ,Flowers ,Pollen Tube ,Biology ,medicine.disease_cause ,olive ,Cell Wall ,Polysaccharides ,Pollen ,Olea ,Botany ,medicine ,Extracellular ,Plant Proteins ,Secretome ,Olive ,exudate ,Eastern lily ,Cell biology ,Stigma ,Secretory protein ,stigma ,Pollen tube ,Lilium ,medicine.symptom ,Research Paper - Abstract
Proteomic analysis of the stigmatic exudate of Lilium longiflorum and Olea europaea led to the identification of 51 and 57 proteins, respectively, most of which are described for the first time in this secreted fluid. These results indicate that the stigmatic exudate is an extracellular environment metabolically active, participating in at least 80 different biological processes and 97 molecular functions. The stigma exudate showed a markedly catabolic profile and appeared to possess the enzyme machinery necessary to degrade large polysaccharides and lipids secreted by papillae to smaller units, allowing their incorporation into the pollen tube during pollination. It may also regulate pollen-tube growth in the pistil through the selective degradation of tube-wall components. Furthermore, some secreted proteins were involved in pollen-tube adhesion and orientation, as well as in programmed cell death of the papillae cells in response to either compatible pollination or incompatible pollen rejection. Finally, the results also revealed a putative cross-talk between genetic programmes regulating stress/defence and pollination responses in the stigma., This work was supported by the Spanish Ministry of Science and Innovation (ERDF-cofinanced projects AGL2008-00517 and PIE-200840I186) and Junta de Andalucía (ERDF-cofinanced project P2010-CVI5767). This work was also funded by the CNRS, the ‘Agence National de la Recherche’ (ANR) and the ‘Region Alsace’. JDR thanks the MICIIN for providing FPU grant funding.
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- 2013
31. Toxoplasma transcription factor TgAP2XI-5 regulates the expression of genes involved in parasite virulence and host invasion
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Guillemette Marot, Tchilabalo Dilezitoko Alayi, Mathieu Gissot, Ludovic Huot, Robert A. Walker, Alain Van Dorsselaer, David Hot, Kami Kim, Stanislas Tomavo, Christine Schaeffer-Reiss, Centre d’Infection et d’Immunité de Lille - INSERM U 1019 - UMR 9017 - UMR 8204 (CIIL), Centre National de la Recherche Scientifique (CNRS)-Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille)-Université de Lille-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut Pasteur de Lille, Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP), Université de Strasbourg (UNISTRA), 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), MOdel for Data Analysis and Learning (MODAL), Inria Lille - Nord Europe, Institut National de Recherche en Informatique et en Automatique (Inria)-Institut National de Recherche en Informatique et en Automatique (Inria)-Laboratoire Paul Painlevé - UMR 8524 (LPP), Centre National de la Recherche Scientifique (CNRS)-Université de Lille-Centre National de la Recherche Scientifique (CNRS)-Université de Lille-Evaluation des technologies de santé et des pratiques médicales - ULR 2694 (METRICS), Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille)-Université de Lille-Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille)-Université de Lille-École polytechnique universitaire de Lille (Polytech Lille)-Université de Lille, Sciences et Technologies, Evaluation des technologies de santé et des pratiques médicales - ULR 2694 (METRICS), Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille)-Université de Lille, Albert Einstein College of Medicine [New York], Institut Pasteur de Lille, Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lille-Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille)-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), Laboratoire Paul Painlevé (LPP), Université de Lille-Centre National de la Recherche Scientifique (CNRS)-Université de Lille-Centre National de la Recherche Scientifique (CNRS)-Université de Lille, Sciences et Technologies-Inria Lille - Nord Europe, Institut National de Recherche en Informatique et en Automatique (Inria)-Institut National de Recherche en Informatique et en Automatique (Inria)-Evaluation des technologies de santé et des pratiques médicales - ULR 2694 (METRICS), Université de Lille-Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille)-Université de Lille-Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille)-École polytechnique universitaire de Lille (Polytech Lille), and Université de Lille-Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille)
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TBX1 ,Transcription, Genetic ,animal diseases ,[SDV]Life Sciences [q-bio] ,Genes, Protozoan ,Protozoan Proteins ,Virulence ,Biology ,Response Elements ,Biochemistry ,03 medical and health sciences ,parasitic diseases ,Gene Regulation ,Molecular Biology ,Transcription factor ,030304 developmental biology ,Genetics ,Regulation of gene expression ,0303 health sciences ,Rhoptry ,General transcription factor ,030306 microbiology ,fungi ,Promoter ,Cell Biology ,TCF4 ,Cell biology ,Gene Expression Regulation ,Toxoplasma ,Toxoplasmosis ,Transcription Factors - Abstract
International audience; Gene regulation in apicomplexan parasites, a phylum containing important protozoan parasites such as Plasmodium and Toxoplasma, is poorly understood. The life cycle of T. gondii is complex, with multiple proliferation and differentiation steps of which tachyzoite proliferation is the most relevant to pathogenesis in humans and animals. Tachyzoites express invasion and virulence factors that are crucial for their survival and manipulation of host cell functions. The expression of those factors is tightly controlled during the tachyzoite cell cycle to permit their correct packaging in newly formed apical secretory organelles named micronemes and rhoptries in the daughter cells. However, little is known about the factors that control the expression of genes encoding the virulence factors present in these parasite-specific secretory organelles. We report that the plant-like nuclear factor TgAP2XI-5 targets more than 300 gene promoters and actively controls the transcription of these genes. Most of these target genes, including those that are essential for parasite virulence, showed a peak of expression in S and M phases of the cell cycle. Furthermore, we identified the cis-regulatory element recognized by TgAP2XI-5 and demonstrated its ability to actively drive gene transcription. Our results demonstrated that TgAP2XI-5 is a novel DNA sequence-specific transcription factor associated with promoter activation. TgAP2XI-5 may regulate gene transcription of crucial virulence factors in T. gondii.
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- 2013
32. An evolutionary conserved zinc finger protein is involved inToxoplasma gondiimRNA nuclear export
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Agnès Hovasse, Laurent Chaloin, Christine Schaeffer-Reiss, Mathieu Gissot, Alain Van Dorsselaer, and Stanislas Tomavo
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0301 basic medicine ,Zinc finger ,Spliceosomal complex ,Genetics ,030102 biochemistry & molecular biology ,Immunology ,Biology ,Microbiology ,3. Good health ,Complementation ,03 medical and health sciences ,Cell nucleus ,030104 developmental biology ,medicine.anatomical_structure ,Virology ,parasitic diseases ,RNA splicing ,Gene expression ,medicine ,Nuclear export signal ,Gene - Abstract
Apicomplexan parasites are responsible for some of the most deadly parasitic diseases affecting humans and livestock. There is an urgent need for new medicines that will target apicomplexan-specific pathways. We characterized a Toxoplasma gondii C2H2 zinc finger protein, named TgZNF2, which is conserved among eukaryotes. We constructed an inducible KO strain (iKO-TgZNF2) for this gene where the tgznf2 gene expression is repressed in the presence of a tetracycline analog (ATc). We showed that the iKO-TgZNF2 parasites are unable to proliferate after depletion of the TgZNF2 protein. Complementation with a full length copy of the gene restores the phenotype Moreover, the homolog of this protein in the related apicomplexan Plasmodium falciparum was shown to efficiently rescue the phenotype, suggesting that this pathway is likely conserved among apicomplexan parasites. We demonstrated that the iKO-mutant lacking TgZNF2 are arrested during the cell cycle during the G1 phase. We identified potential protein partners of this protein among which are spliceosomal complex and mRNA nuclear export components. We confirmed that TgZNF2 is able to bind in vivo to transcripts but splicing is not perturbed in the ATc-treated parasites. Instead, we demonstrated that TgZNF2 depletion leads to the sequestration of polyA+ mRNAs in the nucleus while ribosomal RNAs are not affected. We discovered a conserved protein with specific apicomplexan functional properties that is essential for the survival of T. gondii. TgZNF2 may be crucial to ensure the correct polyA+ mRNA nuclear export, a function that is conserved in P. falciparum.
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- 2016
33. The Pollen Coat Proteome: At the Cutting Edge of Plant Reproduction
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Antonio Castro, François Delalande, Juan David Rejón, María Isabel Rodríguez-García, Christine Schaeffer-Reiss, Juan de Dios Alché, Alain Van Dorsselaer, Consejo Superior de Investigaciones Científicas (España), Ministerio de Economía, Industria y Competitividad (España), Ministerio de Economía y Competitividad (España), Junta de Andalucía, and Agence Nationale de la Recherche (France)
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0106 biological sciences ,0301 basic medicine ,tapetum ,Clinical Biochemistry ,lcsh:QR1-502 ,Stamen ,Context (language use) ,Review ,Biology ,medicine.disease_cause ,Pollen coat ,self-incompatibility ,01 natural sciences ,Biochemistry ,lcsh:Microbiology ,olive ,03 medical and health sciences ,proteomics ,Structural Biology ,Pollen ,Botany ,Locule ,otorhinolaryngologic diseases ,medicine ,pollen coat ,Pollen adhesion ,Molecular Biology ,Tapetum ,food and beverages ,Sexual reproduction ,030104 developmental biology ,010606 plant biology & botany - Abstract
The tapetum is a single layer of secretory cells which encloses the anther locule and sustains pollen development and maturation. Upon apoptosis, the remnants of the tapetal cells, consisting mostly of lipids and proteins, fill the pits of the sculpted exine to form the bulk of the pollen coat. This extracellular matrix forms an impermeable barrier that protects the male gametophyte from water loss and UV light. It also aids pollen adhesion and hydration and retains small signaling compounds involved in pollen–stigma communication. In this study, we have updated the list of the pollen coat’s protein components and also discussed their functions in the context of sexual reproduction, This study was supported by ERDF-confinanced grants PIE-200840I186 and AGL2008-00517/AGR (MICINN), AGL2013-43042-P and BFU2011-22779 (MINECO), and P10-CVI-5767 (Junta de Andalucía). It was also funded by the CNRS, the “Agence National de la Recherche” (ANR), the “Region Alsace” and the French National Proteomic Infrastructure (grant ANR-10-INBS-08; ProFI; the “Infrastructures Nationales en Biologie et Santé” project). We also wish to thank Krysztof Zienkiewicz for kindly providing us with the image in Figure 1b. English proofreading was done by Michael O’Shea., We acknowledge support by the CSIC Open Access Publication Initiative through its Unit of Information Resources for Research (URICI).
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- 2016
34. A Novel Toxoplasma gondii Nuclear Factor TgNF3 Is a Dynamic Chromatin-Associated Component, Modulator of Nucleolar Architecture and Parasite Virulence
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Agnès Hovasse, Christine Schaeffer-Reiss, Alain Van Dorsselaer, Thomas Mouveaux, Alejandro Olguin-Lamas, Stanislas Tomavo, Edwige Madec, Christian Slomianny, Elisabeth Werkmeister, Stéphane Delhaye, Isabelle Callebaut, Centre d’Infection et d’Immunité de Lille - INSERM U 1019 - UMR 9017 - UMR 8204 (CIIL), Centre National de la Recherche Scientifique (CNRS)-Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille)-Université de Lille-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut Pasteur de Lille, Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP), Unité de Glycobiologie Structurale et Fonctionnelle UMR 8576 (UGSF), Institut National de la Recherche Agronomique (INRA)-Université de Lille-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), Institut de minéralogie et de physique des milieux condensés (IMPMC), Université Pierre et Marie Curie - Paris 6 (UPMC)-IPG PARIS-Université Paris Diderot - Paris 7 (UPD7)-Centre National de la Recherche Scientifique (CNRS), Rôle des canaux ioniques membranaires et du calcium intracellulaire dans la physiopathologie de la prostate, Université de Lille, Sciences et Technologies-Institut National de la Santé et de la Recherche Médicale (INSERM), This research was funded by the Centre National de la Recherche Scientifique (CNRS), Institut National de la Sante et de la Recherche Medicale(INSERM), Institut Pasteur de Lille (IPL). The funders had no role in study design, data collection and analysis, decision to publish, or preparationof the manuscript., We would like to thank Dewailly E and Mortuaire M for excellent technical assistance, especially in the earlier phase of this study. We acknowledge Drs Harb O and Roos DS (University of Pennsylvania) for providing the nucleotide sequence of T. gondii rDNA 18S. We are indebted to Drs Hakimi A and Leprince D for providing nucleosome and antibodies specific to core histones, respectively. Special thanks to Drs Gordon Langsley and Robert Walker for critical reading the manuscript and all members of the lab for fruitful discussions., Centre d’Infection et d’Immunité de Lille (CIIL) - U1019 - UMR 8204 (CIIL), Institut Pasteur de Lille, Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lille-Centre National de la Recherche Scientifique (CNRS), Unité de Glycobiologie Structurale et Fonctionnelle - UMR 8576 (UGSF), Université de Lille-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Recherche Agronomique (INRA), Université de Strasbourg (UNISTRA)-Centre National de la Recherche Scientifique (CNRS), Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lille-Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille)-Centre National de la Recherche Scientifique (CNRS), Université de Lille-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), Université Pierre et Marie Curie - Paris 6 (UPMC)-Université Paris Diderot - Paris 7 (UPD7)-Institut de Physique du Globe de Paris (IPG Paris)-Centre National de la Recherche Scientifique (CNRS), and Thomas, Danielle
- Subjects
Proteomics ,MESH: Sequence Analysis, Protein ,Protozoan Proteins ,Antibodies, Protozoan ,MESH: Toxoplasma/pathogenicity ,MESH: DNA-Binding Proteins/chemistry ,Regulatory Sequences, Nucleic Acid ,Mass Spectrometry ,MESH: Protozoan Proteins/chemistry ,Histones ,MESH: Cell Nucleolus/metabolism ,Sequence Analysis, Protein ,MESH: Reverse Transcriptase Polymerase Chain Reaction ,Transcriptional regulation ,MESH: Gene Silencing ,Nuclear protein ,MESH: Ribosomes/metabolism ,Promoter Regions, Genetic ,MESH: High-Throughput Nucleotide Sequencing ,lcsh:QH301-705.5 ,MESH: Promoter Regions, Genetic ,Regulation of gene expression ,MESH: Chromatin Immunoprecipitation ,MESH: Nuclear Proteins/metabolism ,0303 health sciences ,MESH: Histones/metabolism ,Reverse Transcriptase Polymerase Chain Reaction ,MESH: Proteomics ,030302 biochemistry & molecular biology ,High-Throughput Nucleotide Sequencing ,Nuclear Proteins ,Chromatin ,3. Good health ,DNA-Binding Proteins ,MESH: Staining and Labeling ,Histone ,Toxoplasma ,MESH: Nuclear Proteins/genetics ,Cell Nucleolus ,Research Article ,lcsh:Immunologic diseases. Allergy ,Chromatin Immunoprecipitation ,Immunology ,MESH: Protozoan Proteins/genetics ,MESH: Microscopy, Electron ,Biology ,MESH: DNA-Binding Proteins/metabolism ,Microbiology ,MESH: Toxoplasma/metabolism ,Tacrolimus Binding Proteins ,03 medical and health sciences ,Virology ,Genetics ,[SDV.BBM] Life Sciences [q-bio]/Biochemistry, Molecular Biology ,Nucleosome ,MESH: Antibodies, Protozoan ,MESH: Regulatory Sequences, Nucleic Acid ,[SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology ,Gene Silencing ,Molecular Biology/Chromatin Structure ,Molecular Biology ,MESH: Protozoan Proteins/metabolism ,030304 developmental biology ,MESH: Protozoan Proteins/biosynthesis ,MESH: Mass Spectrometry ,MESH: DNA-Binding Proteins/genetics ,Staining and Labeling ,MESH: Nuclear Proteins/biosynthesis ,Infectious Diseases/Protozoal Infections ,Promoter ,Molecular biology ,MESH: Tacrolimus Binding Proteins/chemistry ,MESH: Cell Nucleolus/genetics ,MESH: Nuclear Proteins/chemistry ,Microscopy, Electron ,lcsh:Biology (General) ,Molecular Biology/Nucleolus and Nuclear Bodies ,MESH: Chromatin/metabolism ,biology.protein ,Parasitology ,MESH: Toxoplasma/genetics ,lcsh:RC581-607 ,Chromatin immunoprecipitation ,Ribosomes - Abstract
In Toxoplasma gondii, cis-acting elements present in promoter sequences of genes that are stage-specifically regulated have been described. However, the nuclear factors that bind to these cis-acting elements and regulate promoter activities have not been identified. In the present study, we performed affinity purification, followed by proteomic analysis, to identify nuclear factors that bind to a stage-specific promoter in T. gondii. This led to the identification of several nuclear factors in T. gondii including a novel factor, designated herein as TgNF3. The N-terminal domain of TgNF3 shares similarities with the N-terminus of yeast nuclear FK506-binding protein (FKBP), known as a histone chaperone regulating gene silencing. Using anti-TgNF3 antibodies, HA-FLAG and YFP-tagged TgNF3, we show that TgNF3 is predominantly a parasite nucleolar, chromatin-associated protein that binds specifically to T. gondii gene promoters in vivo. Genome-wide analysis using chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-seq) identified promoter occupancies by TgNF3. In addition, TgNF3 has a direct role in transcriptional control of genes involved in parasite metabolism, transcription and translation. The ectopic expression of TgNF3 in the tachyzoites revealed dynamic changes in the size of the nucleolus, leading to a severe attenuation of virulence in vivo. We demonstrate that TgNF3 physically interacts with H3, H4 and H2A/H2B assembled into bona fide core and nucleosome-associated histones. Furthermore, TgNF3 interacts specifically to histones in the context of stage-specific gene silencing of a promoter that lacks active epigenetic acetylated histone marks. In contrast to virulent tachyzoites, which express the majority of TgNF3 in the nucleolus, the protein is exclusively located in the cytoplasm of the avirulent bradyzoites. We propose a model where TgNF3 acts essentially to coordinate nucleolus and nuclear functions by modulating nucleosome activities during the intracellular proliferation of the virulent tachyzoites of T. gondii., Author Summary Apicomplexa including Toxoplasma gondii are responsible for a variety of deadly infections. These intracellular parasites have complex life cycles within different hosts and their infectivity relies on their capacity to regulate gene expression in response to different environments. However, to date, little is known about nuclear factors that regulate their gene expression. Here, we have characterized parasite nuclear factors that bind to a stage-specific promoter. We identified several nuclear factors including a novel factor, designated herein as TgNF3. The N-terminal domain of TgNF3 shares similarities with the N-terminus of yeast nuclear FK506-binding protein (FKBP), known as a histone chaperone regulating gene silencing. We show that TgNF3 is predominantly a nucleolar, chromatin-associated protein that specifically binds to T. gondii nucleosome-associated histones and promoters. Genome-wide analysis identified promoter occupancies by TgNF3 and we demonstrated a direct role for this factor in transcriptional control of genes involved in parasite metabolism, transcription and translation. Ectopic expression of TgNF3 induces dynamic changes in the size of the nucleolus, and a severe attenuation of parasite virulence in vivo. In avirulent bradyzoites, TgNF3 is found exclusively in the cytoplasm, suggesting a potential role in regulating nucleolar and nuclear functions in the virulent tachyzoites of T. gondii.
- Published
- 2011
35. Detection of prion protein in urine-derived injectable fertility products by a targeted proteomic approach
- Author
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Christine Carapito, François Delalande, Daniel Krewski, Alain Van Dorsselaer, Hélène Diemer, Douglas S. McNair, Neil R. Cashman, Christine Schaeffer-Reiss, and Danièle Thiersé
- Subjects
Proteomics ,Bacterial Diseases ,Anatomy and Physiology ,Nosocomial Infections ,Physiology ,Urine ,Chorionic Gonadotropin ,Mass Spectrometry ,Human chorionic gonadotropin ,0302 clinical medicine ,Endocrinology ,Electrophoresis, Gel, Two-Dimensional ,0303 health sciences ,education.field_of_study ,030219 obstetrics & reproductive medicine ,Multidisciplinary ,Obstetrics and Gynecology ,Reference Standards ,3. Good health ,Medicine ,Infectious diseases ,Female ,Menotropins ,Gonadotropin ,Research Article ,Infertility ,endocrine system ,Drugs and Devices ,Prion diseases ,medicine.drug_class ,Prions ,Urinary system ,Science ,Population ,Molecular Sequence Data ,Endocrine System ,Biology ,Incubation period ,Fertility Agents ,Injections ,03 medical and health sciences ,Adverse Reactions ,medicine ,Humans ,Reproductive Endocrinology ,Amino Acid Sequence ,education ,030304 developmental biology ,Endocrine Physiology ,medicine.disease ,Female Subfertility ,Hormones ,Creutzfeldt-Jakob disease ,nervous system diseases ,Immunology ,Peptides ,Chromatography, Liquid - Abstract
BackgroundIatrogenic transmission of human prion disease can occur through medical or surgical procedures, including injection of hormones such as gonadotropins extracted from cadaver pituitaries. Annually, more than 300,000 women in the United States and Canada are prescribed urine-derived gonadotropins for infertility. Although menopausal urine donors are screened for symptomatic neurological disease, incubation of Creutzfeldt-Jakob disease (CJD) is impossible to exclude by non-invasive testing. Risk of carrier status of variant CJD (vCJD), a disease associated with decades-long peripheral incubation, is estimated to be on the order of 100 per million population in the United Kingdom. Studies showing infectious prions in the urine of experimental animals with and without renal disease suggest that prions could be present in asymptomatic urine donors. Several human fertility products are derived from donated urine; recently prion protein has been detected in preparations of human menopausal gonadotropin (hMG).Methodology/principal findingsUsing a classical proteomic approach, 33 and 34 non-gonadotropin proteins were identified in urinary human chorionic gonadotropin (u-hCG) and highly-purified urinary human menopausal gonadotropin (hMG-HP) products, respectively. Prion protein was identified as a major contaminant in u-hCG preparations for the first time. An advanced prion protein targeted proteomic approach was subsequently used to conduct a survey of gonadotropin products; this approach detected human prion protein peptides in urine-derived injectable fertility products containing hCG, hMG and hMG-HP, but not in recombinant products.Conclusions/significanceThe presence of protease-sensitive prion protein in urinary-derived injectable fertility products containing hCG, hMG, and hMG-HP suggests that prions may co-purify in these products. Intramuscular injection is a relatively efficient route of transmission of human prion disease, and young women exposed to prions can be expected to survive an incubation period associated with a minimal inoculum. The risks of urine-derived fertility products could now outweigh their benefits, particularly considering the availability of recombinant products.
- Published
- 2010
36. Screening of Ole e 1 polymorphism among olive cultivars by peptide mapping and N-glycopeptide analysis
- Author
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Audrey Bednarczyk, Antonio J. Castro, Alain Van Dorsselaer, Juan de Dios Alché, María Isabel Rodríguez-García, and Christine Schaeffer-Reiss
- Subjects
Silver Staining ,Proteome ,Molecular Sequence Data ,Peptide ,Biology ,medicine.disease_cause ,Biochemistry ,Antibodies ,Mass Spectrometry ,fluids and secretions ,Allergen ,Polysaccharides ,Peptide mapping ,Olea ,Complementary DNA ,medicine ,Electrophoresis, Gel, Two-Dimensional ,Amino Acid Sequence ,Cultivar ,Ole e 1 allergen ,Polymorphism ,Molecular Biology ,Peptide sequence ,Plant Proteins ,Genetics ,chemistry.chemical_classification ,Polymorphism, Genetic ,Plant Extracts ,Glycopeptides ,Allergens ,Antigens, Plant ,equipment and supplies ,biology.organism_classification ,Glycopeptide ,chemistry ,Plant proteomics ,Peptides ,Olive cultivars ,Chromatography, Liquid ,Densitometry ,N-glycopeptide - Abstract
10 páginas, 3 figuras, 3 tablas., In the present paper, we have used 2-DE coupled to MS analysis to examine the molecular variability of the Ole e 1 allergen in three olive cultivars (cvs). Our results confirmed that the predicted polymorphism of Ole e 1 at cDNA level is extended to the expressed protein. The profiles of both the Ole e 1 peptides and the N-glycan variants significantly changed among cvs. We observed that Picual and Arbequina cvs presented the highest and lowest degree of Ole e 1 polymorphism, respectively. Some of these peptides and N-glycans were distributed in a cv-specific manner. The putative implications of this molecular polymorphism in the development of the allergy symptoms are discussed., This work was supported by projects BFU2004-00601/BFI and P06-AGR-01791 from the MEC and the Junta de Andalucía, respectively.
- Published
- 2010
37. The plant stigma exudate: a biochemically active extracellular environment for pollen germination?
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Juan David Rejón, María Isabel Rodríguez-García, Christine Carapito, Antonio J. Castro, Krzysztof Zienkiewicz, Juan de Dios Alché, François Delalande, Christine Schaeffer-Reiss, Alain Van Dorsselaer, Ministerio de Ciencia e Innovación (España), Junta de Andalucía, Centre National de la Recherche Scientifique (France), Agence Nationale de la Recherche (France), Région Alsace, and European Commission
- Subjects
Exudate ,Proteomics ,Pollination ,Plant Exudates ,Plant Science ,Flowers ,Biology ,medicine.disease_cause ,Stigma (anatomy) ,Cell biology ,Sexual reproduction ,Addendum ,Pollen ,Olea ,Botany ,Extracellular ,medicine ,Secretion ,Pollen tube ,Lilium ,medicine.symptom ,Plant Proteins - Abstract
During sexual reproduction, pollen performance is greatly influenced by the female tissues. The stigma exudate, i.e., the extracellular secretion that covers the stigma outermost surface, has been usually regarded as a reservoir of water, secondary metabolites, cell wall precursors and compounds that serve as energy supply for rapid pollen tube growth. In an attempt to identify the proteins present in the stigma secretome, we performed a large-scale analysis in two species (Lilium longiflorum and Olea europaea) following a proteomic-based approach. The resulting data strongly suggest that the stigma exudate is not a mere storage site but also a biochemically active environment with a markedly catabolic nature. Thus, this secretion may modulate early pollen tube growth and contribute to the senescence of stigma after pollination. In addition, a putative cross-talk between genetic programs that regulate stress/defense and pollination responses in the stigma is also suggested. The stigma exudate might also functionally diverge between species on the basis on their ecology and the biochemical, morphological and anatomical features of their stigmas. Unexpectedly, we identified in both exudates some intracellular proteins, suggesting that a mechanism other than the canonical ER-Golgi exocytic pathway may exist in the stigma and contribute to exudate secretion., This work was funded by MICINN (ERDF-cofinanced projects AGL2008-00517 and PIE-200840I186), Junta de Andalucía (ERDF-cofinanced project P2010-CVI5767), the CNRS, the “Agence National de la Recherche” (ANR) and the “Region Alsace.” JDR thanks the MICINN for providing FPU grant funding.
38. New applications and opportunities in proteomics
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Guillaumot, Nina, 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), Université de Strasbourg, Alain Van Dorsselaer, Christine Schaeffer-Reiss, and STAR, ABES
- Subjects
Proteomics ,[CHIM.ANAL] Chemical Sciences/Analytical chemistry ,Mass spectrometry ,Characterization ,Spectrométrie de masse ,N-terminomique ,Complexe ,Protéomique ,Chemical labeling ,Caractérisation ,[CHIM.ANAL]Chemical Sciences/Analytical chemistry ,Quantification ,N-terminomic ,Marquage ,Multiprotein complex - Abstract
The aim of this work was to develop new methods for the identification, characterization and quantification of proteins best suited to a large diversity of proteomics studies, which is nowadays essential to biology. Our work has shown that proteomic analysis based on mass spectrometry can be a valuable and relevant tool to evaluate the isolation strategy efficiency set up for a specific complex and thus guide the biologists in their choice. The N-terminomic labeling strategy developed allowed us to describe a biological maturation process by determining precisely the Persephone protein activation sites using specific labeling of the successively generated N-terminal extremities. This work allowed elucidating a new regulation mechanism in the Drosophila innate immunity system. New chemical labeling reagents to target specific amino acids (cysteine, tyrosine and tryptophan) have been set up for fast mass-spectrometry based proteomics. These labeling strategies combined with proteomic tools will allow developing a robust and quantitative approach essential for biological studies., Les objectifs de mes travaux de thèse étaient de développer de nouvelles méthodes d’identification, de caractérisation et de quantification de protéines, mieux adaptées à la diversité des études en protéomique, ce dont la biologie a besoin aujourd’hui. L’analyse protéomique par spectrométrie de masse est apparue comme un outil précieux et pertinent pour évaluer la qualité de l’isolement d’un complexe spécifique, et pour guider les biologistes dans les choix de la stratégie à adopter. La stratégie de marquage de N-terminomique développée a permis de caractériser un processus de maturation biologique en déterminant précisément les sites d’activation de la protéine Perséphone par marquage spécifique des extrémités N-terminales. Ce travail a permis d’élucider un nouveau mécanisme fin de régulation dans l’immunité innée chez la drosophile. De nouveaux modes de marquages ont été mis au point et les familles chimiques des réactifs de marquage étudiés permettront d’adapter au mieux les études de quantifications protéomiques à la nature et aux contraintes des études biologiques à mener.
- Published
- 2017
39. Advances in analytical methodologies for the characterization and quantification in proteomic analysis
- Author
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Bertaccini, Diego, STAR, ABES, 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, Alain Van Dorsselaer, and Christine Schaeffer-Reiss
- Subjects
Label free quantitation ,Analyse protéomique ,[CHIM.ANAL] Chemical Sciences/Analytical chemistry ,Proteolytic cleavage ,[CHIM.ANAL]Chemical Sciences/Analytical chemistry ,N-terminome ,DIA ,Clivages protéolytiques ,TMPP ,DDA ,Protein N-terminus ,Proteogenomics - Abstract
The objective of this Ph.D. thesis was to develop and optimize new methodologies and analytical approaches to improve the potential of the mass spectrometry based proteomics.The first part of this work focused on the development of the N-Termini proteomics. This topic was addressed with a specific N-Termini chemical derivatization based on TMPP. We have shown that our method allowed both specific N-Terminomics and classical proteomics studies in the same experiment.This N-Terminus methodology was applied to study the proteolytic cleavages of the exported proteins in P. falciparum, a parasite responsible for the malaria.In order to automatize the complex and tedious informatics processsing of the MS/SM data of ourTMPP based N-Terminomics method, we have introduced a new approach (named dN-TOP), based on the use of a stable isotope labeled TMPP which made now N-Terminome proteomics compatible with high throughput studies.The second part addresses quantitative aspects of proteomics. It describes the optimization of quantitative methods at the peptide level or at the protein level for five different proteomic studies in the context of protein complex subunits, targeted SRM based prion, quantification of monoclonal antibodies glycation and hemoglobin HbA2 for reference measurement methods standardization., L’objectif de cette thèse était de développer et d’optimiser de nouvelles méthodologies et approches analytiques afin d’améliorer le potentiel de l’analyse protéomique pour les études biologiques.La première partie de ce travail est consacrée à la détermination massive et exacte de la position N-Terminale des protéines (N-Terminome). Pour cela, nous avons utilisé et développé une approche basée sur une dérivation N-Terminale au TMPP. Cette méthodologie de marquage de la position N-Terminale a permis d’aborder l’étude des clivages protéolytiques des protéines exportées par le parasite P. falciparum (pathogène de la malaria) dans le globule rouge.Afin de permettre une exploitation automatique à haut débit des données de MS/MS, nous avons élaboré une nouvelle méthodologie (dénommée dN-TOP). Celle-Ci repose sur l’utilisation de TMPP portant des isotopes stables et permet ainsi d’accéder à la détermination des positions N-Terminales pour des études de N-Terminome à large échelle.La seconde partie est dédiée aux développements de différentes stratégies analytiques de quantification, aussi bien au niveau peptidique qu’au niveau protéique, appliquées à une série de problématiques biologiques. Ces optimisations ont été réalisées dans le contexte de l’étude des complexes protéiques, du dosage de prion par SRM, de quantification des glycations d’anticorps monoclonaux thérapeutiques et de l’hémoglobine HbA2 pour la standardisation des méthodes de référence.
- Published
- 2014
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