Identification of a novel consensus sequence at the cleavage site of the Lassa virus glycoprotein.
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The Lassa virus glycoprotein consists of an amino-terminal and a carboxy-terminal cleavage fragment designated GP-1 and GP-2, respectively, that are derived by proteolysis from the precursor GP-C. The membrane-anchored GP-2 obtained from purified virions of the Josiah strain revealed the N-terminal tripeptide GTF(262) when analyzed by Edman degradation. Upstream of this site, GP-C contains the tetrapeptide sequence RRLL(259), which is conserved in all Lassa virus isolates published to date. Systematic mutational analysis of vector-expressed GP-C revealed that the motif R-X (L/I/V)-L(259) (where X stands for L, I, or V) is essential for cleavage of the peptide bond between leucine(259) and glycine(260). This cleavage motif is homologous to the consensus sequence recognized by a novel class of cellular endoproteases which have so far not been implicated in the processing of viral glycoproteins. (+info)
Evidence of two Lyssavirus phylogroups with distinct pathogenicity and immunogenicity.
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The genetic diversity of representative members of the Lyssavirus genus (rabies and rabies-related viruses) was evaluated using the gene encoding the transmembrane glycoprotein involved in the virus-host interaction, immunogenicity, and pathogenicity. Phylogenetic analysis distinguished seven genotypes, which could be divided into two major phylogroups having the highest bootstrap values. Phylogroup I comprises the worldwide genotype 1 (classic Rabies virus), the European bat lyssavirus (EBL) genotypes 5 (EBL1) and 6 (EBL2), the African genotype 4 (Duvenhage virus), and the Australian bat lyssavirus genotype 7. Phylogroup II comprises the divergent African genotypes 2 (Lagos bat virus) and 3 (Mokola virus). We studied immunogenic and pathogenic properties to investigate the biological significance of this phylogenetic grouping. Viruses from phylogroup I (Rabies virus and EBL1) were found to be pathogenic for mice when injected by the intracerebral or the intramuscular route, whereas viruses from phylogroup II (Mokola and Lagos bat viruses) were only pathogenic by the intracerebral route. We showed that the glycoprotein R333 residue essential for virulence was naturally replaced by a D333 in the phylogroup II viruses, likely resulting in their attenuated pathogenicity. Moreover, cross-neutralization distinguished the same phylogroups. Within each phylogroup, the amino acid sequence of the glycoprotein ectodomain was at least 74% identical, and antiglycoprotein virus-neutralizing antibodies displayed cross-neutralization. Between phylogroups, the identity was less than 64.5% and the cross-neutralization was absent, explaining why the classical rabies vaccines (phylogroup I) cannot protect against lyssaviruses from phylogroup II. Our tree-axial analysis divided lyssaviruses into two phylogroups that more closely reflect their biological characteristics than previous serotypes and genotypes. (+info)
Mucosal immunization with Salmonella typhimurium expressing Lassa virus nucleocapsid protein cross-protects mice from lethal challenge with lymphocytic choriomeningitis virus.
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OBJECTIVES: Lassa fever virus (LAS) is transmitted to man by rodent carriers and is fatal in a third of untreated cases. Our goal is to provide immune protection from Lassa fever by mucosal vaccination. STUDY DESIGN/METHODS: Mice were vaccinated intragastrically with control vectors or with vectors (vaccinia or Salmonella) expressing LAS nucleocapsid protein (NP). Mice were challenged intracranially with a lethal dose of the related arenavirus, lymphocytic choriomeningitis virus (LCMV), as a measure of the vaccine's ability to elicit cross-protection. RESULTS: Salmonella and vaccinia vectors expressing LAS NP each protected a third of the mice from lethal challenge with LCMV. All mice vaccinated with a vector expressing LCMV NP were protected as expected. CONCLUSIONS: The LAS recombinant Salmonella vector is comparable to the LAS recombinant vaccinia vector in its ability to cross-protect mice from lethal challenge. Nucleocapsid protein is an inadequate immunogen on its own, but provides sufficient cross-protection to make it a useful component of a broadly reactive arenavirus vaccine. (+info)
Lassa fever encephalopathy: Lassa virus in cerebrospinal fluid but not in serum.
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The pathogenesis of neurologic complications of Lassa fever is poorly understood. A Nigerian patient had fever, disorientation, seizures, and blood-brain barrier dysfunction, and Lassa virus was found in cerebrospinal fluid (CSF) but not in serum. The concentration of Lassa virus RNA in CSF corresponded to 1 x 10(3) pfu/mL, as determined by a quantitative real-time polymerase chain reaction assay. To characterize the Lassa virus in CSF, the 3.5-kb S RNA was sequenced. In the S RNA coding sequences, the CSF strain differed between 20% and 24.6% from all known prototype strains. These data suggest that Lassa virus or specific Lassa virus strains can persist in the central nervous system and thus contribute to neuropathogenesis. Lassa virus infection should be considered in West African patients or in travelers returning from this area who present only with fever and neurologic signs. (+info)
The Lassa virus glycoprotein precursor GP-C is proteolytically processed by subtilase SKI-1/S1P.
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The surface glycoprotein of the Lassa virus, a member of the arenaviridae family, is synthesized as a 76-kDa precursor (GP-C) that is posttranslationally cleaved into an N-terminal 44-kDa subunit and a C-terminal membrane-anchored 36-kDa subunit. Cleavage occurs at the C-terminal end of the unusual recognition motif R-R-L-L. We show here that GP-C is cleaved in the endoplasmic reticulum by the cellular subtilase SKI-1/S1P, an enzyme that has so far been observed to be involved in cholesterol metabolism. Furthermore, we present evidence that only cleaved glycoprotein is incorporated into virions and that this is necessary for the formation of infectious virus. To our knowledge, there have been no previous reports of this type of viral glycoprotein processing, one that may be an interesting target for antiviral therapy. (+info)
Experimental Lassa virus infection in the squirrel monkey.
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Experimental Lassa virus infection was investigated in a nonhuman primate in order to elucidate the target organs of the viral infection and the course of pathologic events. Four squirrel monkeys (Saimiri scirreus) were inoculated intramuscularly with Lassa virus and sacrificed for organ titrations and histopathology, one each day, on Days 7, 12, 14, and 28 after inoculation. The animals showed a variable clinical course, with an incubation period of 8 to 18 days. The virus was demonstrated to be virtually pantropic; however, lymph node, liver, and kidney were key early targets. After the onset of overt disease, patterns of lymphoreticulotropism, hepatotropism, nephrotropism, adrenotropism, and persistent viremia were evident. Complement-fixing antibody failed to develop after 28 days of infection. Histopathologic findings included germinal center necrosis in spleen and lymph node; myocarditis; acute arteritis; renal tubular necrosis and regeneration; hepatocytic regeneration; chronic inflammation of choroid plexus, ependyma, and meninges; and cerebral perivascular cuffing. There is a relationship between many of these lesions and certain features of other arenavirus infections. The model offers the opportunity to pursue investigations of experimental pathogenesis, transmissibility, and efficacy of immunotherapy. (+info)
Interaction of lyssaviruses with the low-affinity nerve-growth factor receptor p75NTR.
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The low-affinity nerve-growth factor receptor p75NTR interacts in vitro with the rabies virus (RV) glycoprotein and serves as a receptor for RV. The Lyssavirus genus comprises seven genotypes (GTs) of rabies and rabies-related viruses. The ability of p75NTR to interact with the glycoprotein of representative lyssaviruses from each GT was investigated. This investigation was based on a specific binding assay between BSR cells infected with a lyssavirus and Spodoptera frugiperda (Sf21) cells expressing p75NTR on the cell surface. A specific interaction was observed with the glycoprotein of GT 1 RV (challenge virus standard or Pasteur virus strains) as well as wild-type RV and the glycoprotein of GT 6 European bat lyssavirus type 2. In contrast, no interaction was detected with the glycoprotein of lyssaviruses of GTs 2-5 and 7. Therefore, p75NTR is only a receptor for some lyssavirus glycoproteins, indicating that the other GTs must use an alternative specific receptor. (+info)
Self-assembly properties of a model RING domain.
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RING domains act in a variety of essential cellular processes but have no general function ascribed to them. Here, we observe that purified arenaviral protein Z, constituted almost entirely by its RING domain, self-assembles in vitro into spherical structures that resemble functional bodies formed by Z in infected cells. By using a variety of biophysical methods we provide a thermodynamic and kinetic framework for the RING-dependent self-assembly of Z. Assembly appears coupled to substantial conformational reorganization and changes in zinc coordination of site II of the RING. Thus, the rate-limiting nature of conformational reorganization observed in the folding of monomeric proteins can also apply to the assembly of macromolecular scaffolds. These studies describe a unique mechanism of nonfibrillar homogeneous self-assembly and suggest a general function of RINGs in the formation of macromolecular scaffolds that are positioned to integrate biochemical processes in cells. (+info)