title: Canine Distemper Virus Fusion Activation: Critical Role of Residue E123 of CD150/SLAM.
abstract: Measles virus (MeV) and canine distemper virus (CDV) possess tetrameric attachment proteins (H) and trimeric fusion proteins, which cooperate with either SLAM or nectin 4 receptors to trigger membrane fusion for cell entry. While the MeV H-SLAM cocrystal structure revealed the binding interface, two distinct oligomeric H assemblies were also determined. In one of the conformations, two SLAM units were sandwiched between two discrete H head domains, thus spotlighting two binding interfaces ("front" and "back"). Here, we investigated the functional relevance of both interfaces in activating the CDV membrane fusion machinery. While alanine-scanning mutagenesis identified five critical regulatory residues in the front H-binding site of SLAM, the replacement of a conserved glutamate residue (E at position 123, replaced with A [E123A]) led to the most pronounced impact on fusion promotion. Intriguingly, while determination of the interaction of H with the receptor using soluble constructs revealed reduced binding for the identified SLAM mutants, no effect was recorded when physical interaction was investigated with the full-length counterparts of both molecules. Conversely, although mutagenesis of three strategically selected residues within the back H-binding site of SLAM did not substantially affect fusion triggering, nevertheless, the mutants weakened the H-SLAM interaction recorded with the membrane-anchored protein constructs. Collectively, our findings support a mode of binding between the attachment protein and the V domain of SLAM that is common to all morbilliviruses and suggest a major role of the SLAM residue E123, located at the front H-binding site, in triggering the fusion machinery. However, our data additionally support the hypothesis that other microdomain(s) of both glycoproteins (including the back H-binding site) might be required to achieve fully productive H-SLAM interactions.
  
  
  OBJECTIVE
  A complete understanding of the measles virus and canine distemper virus (CDV) cell entry molecular framework is still lacking, thus impeding the rational design of antivirals. Both viruses share many biological features that partially rely on the use of analogous Ig-like host cell receptors, namely, SLAM and nectin 4, for entering immune and epithelial cells, respectively. Here, we provide evidence that the mode of binding between the membrane-distal V domain of SLAM and the attachment protein (H) of morbilliviruses is very likely conserved. Moreover, although structural information revealed two discrete conformational states of H, one of the structures displayed two H-SLAM binding interfaces ("front" and "back"). Our data not only spotlight the front H-binding site of SLAM as the main determinant of membrane fusion promotion but suggest that the triggering efficiency of the viral entry machinery may rely on a local conformational change within the front H-SLAM interactive site rather than the binding affinity.
authors.affiliation: Division of Neurological Sciences, DCR-VPH, Vetsuisse Faculty, University of Bern, Bern, Switzerland Graduate School for Cellular and Biomedical Sciences, University of Bern, Bern, Switzerland.
authors.forename: Mojtaba
authors.initials: M
authors.lastname: Khosravi
authors.affiliation: Division of Neurological Sciences, DCR-VPH, Vetsuisse Faculty, University of Bern, Bern, Switzerland Graduate School for Cellular and Biomedical Sciences, University of Bern, Bern, Switzerland.
authors.forename: Fanny
authors.initials: F
authors.lastname: Bringolf
authors.affiliation: University Clinic RIA, Department of Immunology, University of Bern, Bern, Switzerland.
authors.forename: Silvan
authors.initials: S
authors.lastname: Röthlisberger
authors.affiliation: Institute for Medical Microbiology and Hospital Hygiene, University of Marburg, Marburg, Germany.
authors.forename: Maria
authors.initials: M
authors.lastname: Bieringer
authors.affiliation: Institute for Virology and Immunobiology, University of Würzburg, Würzburg, Germany.
authors.forename: Jürgen
authors.initials: J
authors.lastname: Schneider-Schaulies
authors.affiliation: Division of Neurological Sciences, DCR-VPH, Vetsuisse Faculty, University of Bern, Bern, Switzerland.
authors.forename: Andreas
authors.initials: A
authors.lastname: Zurbriggen
authors.affiliation: Center for Fish and Wildlife Medicine, DIP, Vetsuisse Faculty, University of Bern, Switzerland Institute of Veterinary Bacteriology, DIP, Vetsuisse Faculty, University of Bern, Switzerland.
authors.forename: Francesco
authors.initials: F
authors.lastname: Origgi
authors.affiliation: Division of Neurological Sciences, DCR-VPH, Vetsuisse Faculty, University of Bern, Bern, Switzerland philippe.plattet@vetsuisse.unibe.ch.
authors.forename: Philippe
authors.initials: P
authors.lastname: Plattet
mesh_terms: D000818:Animals; D015703:Antigens, CD / Q000235:genetics / Q000378:metabolism*; D001665:Binding Sites; D002460:Cell Line; D004252:DNA Mutational Analysis; D004217:Distemper Virus, Canine / Q000502:physiology*; D054884:Host-Pathogen Interactions; D006801:Humans; D050576:Membrane Fusion Proteins / Q000378:metabolism; D050505:Mutant Proteins / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D011956:Receptors, Cell Surface / Q000235:genetics / Q000378:metabolism*; D000071177:Signaling Lymphocytic Activation Molecule Family Member 1; D053586:Virus Internalization
chemical_list: D015703:Antigens, CD; D050576:Membrane Fusion Proteins; D050505:Mutant Proteins; D011956:Receptors, Cell Surface; C000604059:SLAMF1 protein, human; D000071177:Signaling Lymphocytic Activation Molecule Family Member 1