Structural Virology. Near-Atomic Cryo-EM Structure of the Helical Measles Virus Nucleocapsid
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Structural virology. Near-atomic cryo-EM structure of the helical measles virus nucleocapsid. Irina Gutsche, Ambroise Desfosses, Grégory Effantin, Wai-Li Ling, Melina Haupt, Rob W H Ruigrok, Carsten Sachse, Guy Schoehn To cite this version: Irina Gutsche, Ambroise Desfosses, Grégory Effantin, Wai-Li Ling, Melina Haupt, et al.. Structural virology. Near-atomic cryo-EM structure of the helical measles virus nucleocapsid.. Science, American Association for the Advancement of Science, 2015, 348 (6235), pp.704-7. hal-01162615 HAL Id: hal-01162615 https://hal.univ-grenoble-alpes.fr/hal-01162615 Submitted on 24 Nov 2020 HAL is a multi-disciplinary open access L’archive ouverte pluridisciplinaire HAL, est archive for the deposit and dissemination of sci- destinée au dépôt et à la diffusion de documents entific research documents, whether they are pub- scientifiques de niveau recherche, publiés ou non, lished or not. The documents may come from émanant des établissements d’enseignement et de teaching and research institutions in France or recherche français ou étrangers, des laboratoires abroad, or from public or private research centers. publics ou privés. RESEARCH | REPORTS STRUCTURAL VIROLOGY The left-handed MeV Ncore-RNA helix is com- posed of 12.34 nucleoprotein subunits per turn, with a pitch of 49.54 Å and an outer diameter of 190 Å, in good agreement with previous studies Near-atomic cryo-EM structure of the (3–5)(tableS1).TheRNAthreadwindsaround the nucleoprotein bobbin, accommodated inside helical measles virus nucleocapsid the cleft between the outward-pointing NTD and the inward-oriented CTD, andshieldedfromabove Irina Gutsche,1,2* Ambroise Desfosses,3† Grégory Effantin,1,2 Wai Li Ling,4,5,6 by the N subunits from the successive helical turn Melina Haupt,7 Rob W. H. Ruigrok,1,2 Carsten Sachse,3 Guy Schoehn1,2,4,5,6 (Fig. 1B). As in the nsNSV N-RNA rings, MeV Ncore oligomerization is mainly mediated via the Measles is a highly contagious human disease. We used cryo–electron microscopy and single exchange subdomains called the NTD arm (resi- particle–based helical image analysis to determine the structure of the helical nucleocapsid dues 1 to 36) and the CTD arm (residues 373 to formed by the folded domain of the measles virus nucleoprotein encapsidating an RNA 391). Our 3D reconstruction shows how the NTD at a resolution of 4.3 angstroms. The resulting pseudoatomic model of the measles virus arm of the Ni protomer inserts into a groove in nucleocapsid offers important insights into the mechanism of the helical polymerization of the CTD of the Ni+1 subunit, whereas the CTD nucleocapsids of negative-strand RNA viruses, in particular via the exchange subdomains of the arm lies on top of the NCTD of the Ni–1 subunit, nucleoprotein. The structure reveals the mode of the nucleoprotein-RNA interaction and generating a repeated helical structure (Fig. 1, C explains why each nucleoprotein of measles virus binds six nucleotides, whereas the respiratory to F). Unlike in the model of the RSV nucleocapsid syncytial virus nucleoprotein binds seven. It provides a rational basis for further analysis of (15), the Ni–1 and Ni+1 subunits in the MeV Ncore- measles virus replication and transcription, and reveals potential targets for drug design. RNA helix do not interact directly. Downloaded from The domain organization of the MeV Ncore (Fig. 1D) corroborates the assumption that all onsegmented negative-strand RNA viruses to 525) (Fig. 1A) that confers to the nucleocapsids nsNSV nucleoproteins share the same global fold (nsNSVs, or Mononegavirales)causeepi- a structural plasticity hampering their analysis at (2, 16) (fig. S2A). In particular, the CTD fold appears demics of serious respiratory tract illnesses better than ~2 nm resolution (3, 4). conserved among the Paramyxoviridae (MeV, [e.g., measles virus (MeV) and respiratory Removal of the Ntail tightens the MeV nucleo- NiV, RSV) (7, 12). The major difference between N http://science.sciencemag.org/ syncytial virus (RSV)] and outbreaks of capsid helix, decreasing both its diameter and the pneumovirus RSV and the Paramyxovirinae lethal zoonotic diseases [e.g., Ebola virus and pitch (3–5). Our previous cryo–electron micros- MeV and NiV is located, as predicted (7, 12), at Nipah virus (NiV)]. Although preventable by vac- copy (cryo-EM) map of the resulting Ncore-RNA the distal tip of the NTD (residues 91 to 159 in cination, measles still remains one of the leading helix at 12 Å resolution (5) provided precise heli- MeV N). The local resolution of the cryo-EM map sources of death among young children worldwide cal parameters and the domain organization in this solvent-exposed region, known as an anti- (1). MeV belongs to the Paramyxoviridae family of the MeV Ncore, and a number of low- and genic site of MeV N (residues 122 to 150) (17), of Mononegavirales, which is further divided in medium-resolution EM reconstructions of other seemstobethelowest,probablyindicatingmo- two subfamilies: Paramyxovirinae (containing Mononegavirales nucleocapsids are available bility with respect to the nucleocapsid core (fig. 0 MeV and NiV) and Pneumovirinae (e.g., RSV). (6–9). However, the only atomic-resolution struc- S1D). In NiV N core-P50 crystals, the RNA-free nu- The genome of negative-strand RNA viruses tural information about Mononegavirales nucleo- cleoprotein is observed in an open state (12). The (NSVs) is enwrapped with the viral nucleoprotein capsids comes from x-ray crystallography of present structure of the MeV Ncore-RNA mono- N. The resulting ribonucleoprotein complex, called N-RNA rings obtained upon nonspecific encap- mer enables accurate modeling of the hinge mo- on March 5, 2018 the nucleocapsid, protects the viral genetic infor- sidation of short cellular RNAs by recombinant tion between NTD and CTD, which has been mation while providing a flexible helical template nucleoproteins of RSV (7) and of two rhabdoviruses proposed to be associated with the open-closed for viral transcription and replication by the viral (10, 11), whereas such rings could not yet be ob- transition accompanying nucleocapsid formation RNA polymerase L, which, in the Paramyxoviridae served for Paramyxovirinae. In addition, the crys- (fig. S2, B and C) (12). 0 0 and Rhabdoviridae families of Mononegavirales, tal structure of the RNA-free NiV Ncore (N core) On the basis of the NiV N core-P50 structure, 0 is associated with the modular phosphoprotein bound to a short N-terminal region of P (N core- the N-terminal domain of Paramyxovirinae P cofactor P (2). As a unique structure in nucleic acid P50)(12), has been solved recently. We therefore was assigned two simultaneous roles (12): (i) biology, the NSV nucleocapsid constitutes an at- focused on the helical state of the MeV Ncore-RNA trapping the nucleoprotein in an open RNA- tractive potential target for antiviral drugs with- nucleocapsid, so as to (i) obtain a high-resolution free conformation by rigidifying the CTD, and out harmful side effects. Like all Mononegavirales three-dimensional (3D) structure of the Ncore (ii) preventing its polymerization by interfering nucleoproteins, MeV N is composed of two globular monomer, (ii) identify the molecular determinants with the binding of the exchange subdomains. 0 domains, the N- and C-terminal domains (NTD of helical nucleocapsid polymerization, and (iii) Indeed, although crystallization of NiV N core-P50 and CTD), which together form a stable peanut- directly visualize the RNA inside the nucleopro- required deletion of both the NTD and CTD shaped nucleoprotein core (Ncore, residues 1 to 391 teinandunderstandwhyeachMeVnucleopro- arms, the first and second a helices of the helix- in MeV N) holding the RNA molecule in the tein binds exactly six ribonucleotides, whereas kink-helix peptide of P (Pa1andPa2) were shown interdomain cleft. In addition, like other Para- the RSV N binds seven. Here, we report the 3D tooverlaywiththeNTDandCTDarmloopsof myxovirinae, MeV N features a long, intrinsi- structure of recombinant MeV Ncore-RNA nucleo- RSV N, respectively (12). In MeV, the NTD arm cally disordered tail domain (Ntail, residues 392 capsids at 4.3 Å resolution, determined by single begins with an a helix (residues 2 to 14) (Fig. 1D particle–based helical image analysis (13, 14) and fig. S2A) that fits into a hydrophobic groove 1 CNRS, Unit for Virus Host-Cell Interactions, 38042 (fig. S1). This cryo-EM map reveals the detailed formed by three conserved CTD a helices of the Ni+1 Grenoble, France. 2Université Grenoble Alpes, Unit for Virus domain architecture of the N ,itssecondary subunit (Fig. 2A) (12). Specifically, four aromatic 3 core Host-Cell Interactions, 38042 Grenoble, France. Structural structure elements and many bulky side chains, residues (Phe11, Phe269,Tyr303,andPhe324)thatare and Computational Biology Unit, European Molecular Biology Laboratory, 69917 Heidelberg, Germany. 4Université Grenoble the rationale of the nucleoprotein packing into a conserved in Paramyxovirinae (12) seem to Alpes, IBS, 38044 Grenoble, France. 5CNRS, IBS, 38044 helix, and the mode of nucleoprotein-RNA inter- stack together, thereby fixing the NTD arm a Grenoble, France. 6CEA, IBS, 38044 Grenoble, France. action. Combined with the atomic structures of helix and rigidifying the helix bundle around it 7 Institut Laue-Langevin, 38000 Grenoble, France. 0 a † the RSV N and NiV N core,this3Dreconstruction (Fig. 2B). Furthermore, the NTD arm helix of *Corresponding author. E-mail: [email protected] Present a address: School of Biological Sciences, University of Auckland, allows us to build a reliable pseudoatomic model MeV Ni−1 perfectly superimposes with P 1from 0 Auckland 1010, New Zealand. of the MeV Ncore-RNA helix. the NiV N core-P50 structure, although the latter 704 8MAY2015• VOL 348 ISSUE 6235 sciencemag.org SCIENCE RESEARCH | REPORTS is actually positioned upside down, and the CTD helix of P and the NTD arm a helix of MeV N The MeV Ncore-RNA structure explains bio- arm loop of the MeV Ni+1 overlaps with Pa2(Fig.