Phospholipid Interactions of a Peptide from the Fusion-Related Domain of the Glycoprotein of VHSV, a Fish Rhabdovirus

Núñez, Elena; Fernández, Asia M.; Estepa, A.; González-Ros, José M.; Gavilanes, Francisco; Coll, Julio

doi:10.1006/viro.1998.9076

Cited by 26 publications

(20 citation statements)

References 53 publications

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“…It occurs at pH 6.0 but not at pH 7.5, and depends on the presence of PS on the target membrane. The data obtained with p2 peptide of VHSV also indicate that this peptide might play an active role in fusion: it mediates phospholipid vesicle fusion, lipid mixing, and leakage of liposome contents and inserts itself into liposome membranes (50). These results together suggest that p2-like peptides directly participate in membrane fusion mediated by rhabdoviruses probably through the protonation of their His residues.…”

Section: Determination Of the Rhabdovirus Fusion Peptidementioning

confidence: 71%

Viral membrane fusion: is glycoprotein G of rhabdoviruses a representative of a new class of viral fusion proteins?

Poian

Carneiro

Stauffer

2005

Braz J Med Biol Res

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Enveloped viruses always gain entry into the cytoplasm by fusion of their lipid envelope with a cell membrane. Some enveloped viruses fuse directly with the host cell plasma membrane after virus binding to the cell receptor. Other enveloped viruses enter the cells by the endocytic pathway, and fusion depends on the acidification of the endosomal compartment. In both cases, virus-induced membrane fusion is triggered by conformational changes in viral envelope glycoproteins. Two different classes of viral fusion proteins have been described on the basis of their molecular architecture. Several structural data permitted the elucidation of the mechanisms of membrane fusion mediated by class I and class II fusion proteins. In this article, we review a number of results obtained by our laboratory and by others that suggest that the mechanisms involved in rhabdovirus fusion are different from those used by the two well-studied classes of viral glycoproteins. We focus our discussion on the electrostatic nature of virus binding and interaction with membranes, especially through phosphatidylserine, and on the reversibility of the conformational changes of the rhabdovirus glycoprotein involved in fusion. Taken together, these data suggest the existence of a third class of fusion proteins and support the idea that new insights should emerge from studies of membrane fusion mediated by the G protein of rhabdoviruses. In particular, the elucidation of the three-dimensional structure of the G protein or even of the fusion peptide at different pH's might provide valuable information for understanding the fusion mechanism of this new class of fusion proteins. Correspondence

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Section: Determination Of the Rhabdovirus Fusion Peptidementioning

confidence: 71%

Viral membrane fusion: is glycoprotein G of rhabdoviruses a representative of a new class of viral fusion proteins?

Poian

Carneiro

Stauffer

2005

Braz J Med Biol Res

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“…This segment was better characterized for viral hemorrhagic septicemia virus, a rhabdovirus of salmonids, and it was named p2 peptide (14,15). Viral hemorrhagic septicemia virus p2 peptide mediates phospholipid vesicle fusion, lipid mixing, and leakage of liposome contents and inserts itself into liposome membranes by adopting a ␤-sheet conformation (16). p2-like peptide was found among all rhabdoviruses and contains two histidyl residues in VSV G protein (17).…”

Section: Vesicular Stomatitis Virus (Vsv)mentioning

confidence: 99%

Membrane Fusion Induced by Vesicular Stomatitis Virus Depends on Histidine Protonation

Carneiro

Stauffer

Lima

et al. 2003

Journal of Biological Chemistry

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Entry of enveloped animal viruses into their host cells always depends on a step of membrane fusion triggered by conformational changes in viral envelope glycoproteins. Vesicular stomatitis virus (VSV) infection is mediated by virus spike glycoprotein G, which induces membrane fusion at the acidic environment of the endosomal compartment. VSV-induced membrane fusion occurs at a very narrow pH range, between 6.2 and 5.8, suggesting that His protonation is required for this process. To investigate the role of His in VSV fusion, we chemically modified these residues using diethylpyrocarbonate (DEPC). We found that DEPC treatment inhibited membrane fusion mediated by VSV in a concentration-dependent manner and that the complete inhibition of fusion was fully reversed by incubation of modified virus with hydroxylamine. Fluorescence measurements showed that VSV modification with DEPC abolished pHinduced conformational changes in G protein, suggesting that His protonation drives G protein interaction with the target membrane at acidic pH. Mass spectrometry analysis of tryptic fragments of modified G protein allowed the identification of the putative active His residues. Using synthetic peptides, we showed that the modification of His-148 and His-149 by DEPC, as well as the substitution of these residues by Ala, completely inhibited peptide-induced fusion, suggesting the direct participation of these His in VSV fusion.Membrane fusion is an essential step in the entry of enveloped viruses into their host cells (1-3). Virus-induced fusion is always mediated by viral surface glycoprotein and may occur through two different general mechanisms: (i) surface fusion between viral envelope and host cell plasma membrane after virus interaction with its cellular receptor, and (ii) fusion of endosomal membrane with viral envelope after virus particle internalization by receptor-mediated endocytosis. In the latter case, fusion is triggered by conformational changes in viral glycoproteins induced by the decrease in the pH of the endosomal medium.Vesicular stomatitis virus (VSV) 1 is a member of Rhabdoviridae family, genus Vesiculovirus. Rhabdoviruses contain helically wound ribonucleocapisid surrounded by a lipid bilayer through which spikes project. These spikes are formed by trimers of a single type of glycoprotein, named G protein. VSV enters into the cell by endocytosis followed by low pH-induced membrane fusion in the endosome (4, 5), which is catalyzed by VSV G protein (6). A common feature of viral fusion proteins is that they bear a highly conserved hydrophobic fusion domain, which is most often located at the N terminus of the polypeptide chain (7). However, VSV G protein does not contain an apolar amino acid sequence similar to the fusion peptides found in other viruses, suggesting alternative mechanisms involved in VSV-induced membrane fusion.We have shown recently (8) that VSV-induced fusion depends on a dramatic structure reorganization of G protein, which occurs within a very narrow pH range, close to 6.0. In addition, ...

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“…Alignment of the pG sequence of VSV with those of 14 other animal rhabdoviruses representing vesiculoviruses, lyssaviruses, ephemeroviruses, and novirhabdoviruses, made it possible to predict the locations of hypothetical fusion peptides in these other rhabdoviruses including VHSV (42). According to that model, the expected fusion peptide of VHSV could be located between positions 142 and 159.On the other hand, evidence obtained by the use of synthetic and recombinant peptides of the pG of VHSV suggested that the sequence from positions 56 to 110 (frg11), containing noncanonical heptad repeats (11) and the phospholipid-binding peptide (p2), may also be involved in fusion (14,15,32). For instance, recombinant frg11 showed dramatic changes in both solubility and ␤-sheet conformation at low pH and induced low-pH-dependent cell-cell fusion by itself when added to cell monolayers (15).…”

mentioning

confidence: 99%

“…On the other hand, evidence obtained by the use of synthetic and recombinant peptides of the pG of VHSV suggested that the sequence from positions 56 to 110 (frg11), containing noncanonical heptad repeats (11) and the phospholipid-binding peptide (p2), may also be involved in fusion (14,15,32). For instance, recombinant frg11 showed dramatic changes in both solubility and ␤-sheet conformation at low pH and induced low-pH-dependent cell-cell fusion by itself when added to cell monolayers (15).…”

mentioning

confidence: 99%

Conformation- and Fusion-Defective Mutations in the Hypothetical Phospholipid-Binding and Fusion Peptides of Viral Hemorrhagic Septicemia Salmonid Rhabdovirus Protein G

Rocha¹,

Ruiz²,

Tafalla³

et al. 2004

J Virol

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Fourteen single and two double point mutants in the highly conserved region (positions 56 to 159) of the G gene of viral hemorrhagic septicaemia virus (VHSV), a salmonid rhabdovirus, were selected and obtained in plasmids by site-directed mutagenesis. Fish cell monolayers transfected with the mutant plasmids were then assayed for protein G (pG) expression, conformation-dependent monoclonal antibody (MAb) reactivity, and cell-cell fusion. Some mutations located in the phospholipid-binding p2 peptide (positions 82 to 110; mutants P86A, A96E, G98A, and R107A) abolished both MAb recognition and fusion activity, while others (P79A, L85S, and R103A) abolished MAb recognition but retained fusion at similar or lower pHs compared to those for the wild type. Phospholipid-binding assays of p2-derived synthetic peptides suggested that phosphatidylserine binding was not affected by the mutations studied. On the other hand, three (P79A, L85S, and T135E) of the four mutants retaining fusion activity mapped around two locations showing amino acid variation in 22 VHSV isolates and in neutralizing MAb-resistant mutants described previously. Mutations located in the hypothetical fusion peptide (positions 142 to 159; mutants F147K, P148K, and W154K) abolished both MAb recognition and fusion activity. The existence of mutants with altered conformation and defective fusion in both p2 and fusion peptides provides further evidence in favor of the participation of these and adjacent regions in some of the steps of the VHSV fusion processes, as suggested by previous studies. In addition, because the studied region induced strong immunological responses in trout, some of the mutants described here might be used to design attenuated VHSV vaccines.Rhabdoviruses cause highly damaging diseases in fish reared by the worldwide salmon culture industry. Among fish rhabdoviruses (novirhabdoviruses), the viral hemorrhagic septicemia virus (VHSV), which originated in Europe but has recently been found in America (26, 41), affects not only salmonids but also cod, turbot (38), sea bass, eels, flatfish, and shrimp (8,24,27). Despite many efforts, including successful DNA vaccines at the laboratory level, a commercial vaccine against VHSV is not yet available (1,18,25,30).The whole genome of VHSV has been sequenced (23), and the disulfide structure of its protein G (pG) has been elucidated (12). Reverse genetic methods have been developed for the VHSV-related rhabdovirus infectious hematopoietic necrosis virus (5-7), which could allow the design of new vaccines. The study of rhabdovirus fusion in the VHSV model could be important in the design of alternative vaccines to fight these diseases.The pG mutants of vesicular stomatitis virus (VSV), a wellstudied mammalian rhabdovirus, with an altered extent of fusion and/or optimal fusion pH in vitro, had mutations located either in the fusion peptide or in carboxy-terminal regions affecting the low-pH conformational changes required for fusion (39,40,43). Alignment of the pG sequence of VSV with those of 14 o...

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Phospholipid Interactions of a Peptide from the Fusion-Related Domain of the Glycoprotein of VHSV, a Fish Rhabdovirus

Cited by 26 publications

References 53 publications

Viral membrane fusion: is glycoprotein G of rhabdoviruses a representative of a new class of viral fusion proteins?

Viral membrane fusion: is glycoprotein G of rhabdoviruses a representative of a new class of viral fusion proteins?

Membrane Fusion Induced by Vesicular Stomatitis Virus Depends on Histidine Protonation

Conformation- and Fusion-Defective Mutations in the Hypothetical Phospholipid-Binding and Fusion Peptides of Viral Hemorrhagic Septicemia Salmonid Rhabdovirus Protein G

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