Characterization of the Interaction Domains between the Phosphoprotein and the Nucleoprotein of Human Metapneumovirus
| Autor: | Jean-François Eléouët, Monika Bajorek, Luis Checa Ruano, Marie Galloux, Jenna Fix, Charles-Adrien Richard, Hortense Decool, Olivier Sperandio, Irina Gutsche, Benjamin Bardiaux |
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| Přispěvatelé: | Virologie et Immunologie Moléculaires (VIM (UR 0892)), Université de Versailles Saint-Quentin-en-Yvelines (UVSQ)-Université Paris-Saclay-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), Bioinformatique structurale - Structural Bioinformatics, Institut Pasteur [Paris] (IP)-Centre National de la Recherche Scientifique (CNRS)-Université Paris Cité (UPCité), Molécules Thérapeutiques in silico (MTI), Université Paris Diderot - Paris 7 (UPD7)-Institut National de la Santé et de la Recherche Médicale (INSERM), Institut de biologie structurale (IBS - UMR 5075), Centre National de la Recherche Scientifique (CNRS)-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes (UGA), ANR-19-CE11-0017, Agence Nationale de la Recherche, ANR, This work was carried out with the financial support of the French Agence Nationale de la Recherche, specific program ANR DecRisP, grant no. ANR-19-CE11-0017. We declare that we have no conflicts of interest with the contents of this article., ANR-19-CE11-0017,DecRisP,Décrypter les synthèses d'ARN par les pneumovirus(2019) |
| Jazyk: | angličtina |
| Rok vydání: | 2022 |
| Předmět: |
Models
Molecular viruses MESH: Cricetinae Virus Replication chemistry.chemical_compound Protein-protein interaction Transcription (biology) MESH: Metapneumovirus RNA polymerase Cricetinae Structural modeling MESH: Animals Polymerase Inclusion Bodies [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry Molecular Biology/Structural Biology [q-bio.BM] HMPV Nucleocapsid Proteins Phosphoprotein MESH: RNA Viral MESH: RNA-Dependent RNA Polymerase [SDV.MP.VIR]Life Sciences [q-bio]/Microbiology and Parasitology/Virology RNA Viral MESH: Models Molecular Protein Binding MESH: Mutation Immunology Biology MESH: Phosphoproteins Microbiology Cell Line Human metapneumovirus Virology MESH: Protein Binding Animals Protein Interaction Domains and Motifs Nucleoprotein MESH: Protein Interaction Domains and Motifs Binding Sites Structure and Assembly MESH: Virus Replication RNA MESH: Nucleocapsid Proteins biology.organism_classification Phosphoproteins RNA-Dependent RNA Polymerase MESH: Inclusion Bodies MESH: Cell Line MESH: Binding Sites Viral replication chemistry Insect Science Mutation biology.protein Metapneumovirus |
| Zdroj: | J Virol Journal of Virology Journal of Virology, 2022, 96 (2), pp.e0090921. ⟨10.1128/JVI.00909-21⟩ |
| ISSN: | 0022-538X 1098-5514 |
| DOI: | 10.1128/JVI.00909-21⟩ |
| Popis: | Human metapneumovirus (HMPV) causes severe respiratory diseases in young children. The HMPV RNA genome is encapsidated by the viral nucleoprotein (N), forming an RNA-N complex (N(Nuc)), which serves as the template for genome replication and mRNA transcription by the RNA-dependent RNA polymerase (RdRp). The RdRp is formed by the association of the large polymerase subunit (L), which has RNA polymerase, capping, and methyltransferase activities, and the tetrameric phosphoprotein (P). P plays a central role in the RdRp complex by binding to N(Nuc) and L, allowing the attachment of the L polymerase to the N(Nuc) template. During infection these proteins concentrate in cytoplasmic inclusion bodies (IBs) where viral RNA synthesis occurs. By analogy to the closely related pneumovirus respiratory syncytial virus (RSV), it is likely that the formation of IBs depends on the interaction between HMPV P and N(Nuc), which has not been demonstrated yet. Here, we finely characterized the binding P-N(Nuc) interaction domains by using recombinant proteins, combined with a functional assay for the polymerase complex activity, and the study of the recruitment of these proteins to IBs by immunofluorescence. We show that the last 6 C-terminal residues of HMPV P are necessary and sufficient for binding to N(Nuc) and that P binds to the N-terminal domain of N (N(NTD)), and we identified conserved N residues critical for the interaction. Our results allowed us to propose a structural model for the HMPV P-N(Nuc) interaction. IMPORTANCE Human metapneumovirus (HMPV) is a leading cause of severe respiratory infections in children but also affects human populations of all ages worldwide. Currently, no vaccine or efficient antiviral treatments are available for this pneumovirus. A better understanding of the molecular mechanisms involved in viral replication could help the design or discovery of specific antiviral compounds. In this work, we have investigated the interaction between two major viral proteins involved in HMPV RNA synthesis, the N and P proteins. We finely characterized their domains of interaction and identified a pocket on the surface of the N protein, a potential target of choice for the design of compounds interfering with N-P complexes and inhibiting viral replication. |
| Databáze: | OpenAIRE |
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