title: A single amino acid substitution in the core protein of West Nile virus increases resistance to acidotropic compounds.
abstract: West Nile virus (WNV) is a worldwide distributed mosquito-borne flavivirus that naturally cycles between birds and mosquitoes, although it can infect multiple vertebrate hosts including horses and humans. This virus is responsible for recurrent epidemics of febrile illness and encephalitis, and has recently become a global concern. WNV requires to transit through intracellular acidic compartments at two different steps to complete its infectious cycle. These include fusion between the viral envelope and the membrane of endosomes during viral entry, and virus maturation in the trans-Golgi network. In this study, we followed a genetic approach to study the connections between viral components and acidic pH. A WNV mutant with increased resistance to the acidotropic compound NH4Cl, which blocks organelle acidification and inhibits WNV infection, was selected. Nucleotide sequencing revealed that this mutant displayed a single amino acid substitution (Lys 3 to Glu) on the highly basic internal capsid or core (C) protein. The functional role of this replacement was confirmed by its introduction into a WNV infectious clone. This single amino acid substitution also increased resistance to other acidification inhibitor (concanamycin A) and induced a reduction of the neurovirulence in mice. Interestingly, a naturally occurring accompanying mutation found on prM protein abolished the resistant phenotype, supporting the idea of a genetic crosstalk between the internal C protein and the external glycoproteins of the virion. The findings here reported unveil a non-previously assessed connection between the C viral protein and the acidic pH necessary for entry and proper exit of flaviviruses.
authors.affiliation: Departamento de Biotecnología, Instituto Nacional de Investigación y Tecnología Agraria y Alimentaria (INIA), Madrid, Spain. martin.mangel@inia.es
authors.forename: Miguel A
authors.initials: MA
authors.lastname: Martín-Acebes
authors.forename: Ana-Belén
authors.initials: AB
authors.lastname: Blázquez
authors.forename: Nereida Jiménez
authors.initials: NJ
authors.lastname: de Oya
authors.forename: Estela
authors.initials: E
authors.lastname: Escribano-Romero
authors.forename: Pei-Yong
authors.initials: PY
authors.lastname: Shi
authors.forename: Juan-Carlos
authors.initials: JC
authors.lastname: Saiz
mesh_terms: D019943:Amino Acid Substitution / Q000235:genetics*; D000643:Ammonium Chloride / Q000494:pharmacology; D000704:Analysis of Variance; D000818:Animals; D015153:Blotting, Western; D002522:Chlorocebus aethiops; D006224:Cricetinae; D024882:Drug Resistance, Viral / Q000235:genetics*; D005455:Fluorescent Antibody Technique; D006863:Hydrogen-Ion Concentration; D018942:Macrolides / Q000494:pharmacology; D051379:Mice; D060888:Real-Time Polymerase Chain Reaction; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014709:Vero Cells; D014758:Viral Core Proteins / Q000235:genetics*; D014774:Virulence; D014902:West Nile virus / Q000235:genetics* / Q000472:pathogenicity
chemical_list: D018942:Macrolides; D014758:Viral Core Proteins; D000643:Ammonium Chloride; C036978:concanamycin A