67 research outputs found
Structural and functional studies of the retrovirus envelope proteins: The role of the cytoplasmic tail.
No full textPalmitoylation of the Murine Leukemia Virus Envelope Glycoprotein Transmembrane Subunits
No full textAbstractThe envelope protein of Friend murine leukemia virus is modified by fatty acylation of the transmembrane (TM) protein subunit. The labeling by [3H]palmitic acid was found to be sensitive to treatment with the reducing reagents 2-mercaptoethanol and hydroxylamine, indicating the presence of a thioester linkage. Pulse-chase experiments showed that the precursor protein can be labeled by [3H]palmitic acid prior to its cleavage into the surface and TM subunits. By using site-directed mutagenesis, we determined that palmitoylation occurs on a cysteine residue, Cys 606, located in the transmembrane domain. A thin-layer chromatography assay after acid hydrolysis showed that incorporated label comigrated with palmitic acid. When another cysteine residue was introduced into the cytoplasmic tail 22 amino acids from the transmembrane domain, no palmitoylation was observed to occur on this cysteine residue, demonstrating the importance of the position of the cysteine residue for palmitoylation. Sequence comparison revealed that most retrovirus envelope proteins have one or two conserved cysteine residues in their transmembrane domain. Mutations that change the palmitoylation state of the murine leukemia virus envelope protein did not affect its transport, processing, surface expression, or cell fusion activity. The palmitate-deficient viral envelope proteins were incorporated into virus particles, and replication of the virusin vitrowas not affected significantly by the mutation of the palmitoylation site
Coreceptor-Dependent Inhibition of the Cell Fusion Activity of Simian Immunodeficiency Virus Env Proteins
No full textABSTRACT
The cytoplasmic tail (R peptide) sequence is able to regulate the fusion activity of the murine leukemia virus (MuLV) envelope (Env) protein. We have previously shown that this sequence exerts a profound inhibitory effect on the fusion activity of simian immunodeficiency virus (SIV)-MuLV chimeric Env proteins which contain the extracellular and transmembrane domains of the SIV Env protein. Recent studies have shown that SIV can utilize several alternative cellular coreceptors for its fusion and entry into the cell. We have investigated the fusion activity of SIV and SIV-MuLV chimeric Env proteins using cells that express different coreceptors. HeLa cells were transfected with plasmid constructs that carry the SIV or SIV-MuLV chimeric Env protein genes and were overlaid with either CEMx174 cells or Ghost Gpr15 cells, which express the Gpr15 coreceptor for SIV, or Ghost CCR5 cells, which express CCR5, an alternate coreceptor for SIV. The R-peptide sequence in the SIV-MuLV chimeric proteins was found to inhibit the fusion with CEMx174 cells or Ghost Gpr15 cells. However, a significant level of fusion was still observed when HeLa cells expressing the chimeric Env proteins were cocultivated with Ghost CCR5 cells. These results show that the R-peptide sequence exerts differential effects on the fusion activity of SIV Env proteins using target cells that express alternative coreceptors.</jats:p
Enhanced cellular immune response against SIV Gag induced by immunization with DNA vaccines expressing assembly and release-defective SIV Gag proteins
No full textAbstractCodon-optimized genes were synthesized for the SIVmac239 Gag, a mutant Gag with mutations in the major homology region, and a chimeric Gag containing a protein destruction signal at the N-terminus of Gag. The mutant and chimeric Gag were expressed at levels comparable to that observed for the wild-type Gag protein but their stability and release into the medium were found to be significantly reduced. Immunization of mice with DNA vectors encoding the mutant or chimeric Gag induced fourfold higher levels of anti-SIV Gag CD4 T cell responses than the DNA vector encoding the wild-type SIV Gag. Moreover, anti-SIV Gag CD8 T cell responses induced by DNA vectors encoding the mutant or chimeric Gag were found to be 5- to 10-fold higher than those induced by the DNA construct for the wild-type Gag. These results indicate that mutations disrupting assembly and/or stability of the SIV Gag protein effectively enhance its immunogenicity when expressed from DNA vaccines
Phenotypic and Genetic Characterization of Avian Influenza H5N2 Viruses with Intra- and Inter-Duck Variations in Taiwan
No full textBackground
Human infections with avian influenza viruses (AIVs) have frequently raised global concerns of emerging, interspecies-transmissible viruses with pandemic potential. Waterfowl, the predominant reservoir of influenza viruses in nature, harbor precursors of different genetic lineages that have contributed to novel pandemic influenza viruses in the past.
Methods
Two duck influenza H5N2 viruses, DV518 and DV413, isolated through virological surveillance at a live-poultry market in Taiwan, showed phylogenetic relatedness but exhibited different replication capabilities in mammalian Madin-Darby Canine Kidney (MDCK) cells. This study characterizes the replication properties of the two duck H5N2 viruses and the determinants involved.
Results
The DV518 virus replicated more efficiently than DV413 in both MDCK and chicken DF1 cells. Interestingly, the infection of MDCK cells by DV518 formed heterogeneous plaques with great differences in size [large (L) and small (S)], and the two viral strains (p518-L and p518-S) obtained from plaque purification exhibited distinguishable replication kinetics in MDCK cells. Nonetheless, both plaque-purified DV518 strains still maintained their growth advantages over the plaque-purified p413 strain. Moreover, three amino acid substitutions in PA (P224S), PB2 (E72D), and M1 (A128T) were identified in intra-duck variations (p518-L vs p518-S), whereas other changes in HA (N170D), NA (I56T), and NP (Y289H) were present in inter-duck variations (DV518 vs DV413). Both p518-L and p518-S strains had the N170D substitution in HA, which might be related to their greater binding to MDCK cells. Additionally, polymerase activity assays on 293T cells demonstrated the role of vRNP in modulating the replication capability of the duck p518-L viruses in mammalian cells.
Conclusion
These results demonstrate that intra-host phenotypic variation occurs even within an individual duck. In view of recent human infections by low pathogenic AIVs, this study suggests possible determinants involved in the stepwise selection of virus variants from the duck influenza virus population which may facilitate inter-species transmission
Enhancement of immunogenicity of an HIV Env DNA vaccine by mutation of the Tyr-based endocytosis motif in the cytoplasmic domain
No full textAbstractWe investigated the effect of the conserved tyrosine-based endocytosis motif (YXXΦ) in the cytoplasmic domain of the human immunodeficiency viruses (HIV) envelope protein (Env) on its immunogenicity. Genes with codons optimized for mammalian expression were synthesized for the HIV 89.6 Env with a truncated cytoplasmic domain and a mutant Env in which the tyrosine residue in the YXXΦ motif was changed into a serine. Mutation of the Tyr residue enhanced surface expression of the Env protein. Analysis of immune responses induced by DNA immunization of mice showed that the DNA construct for the Tyr mutant Env induced moderately higher levels of T cell responses. More interestingly, the DNA construct for the mutant Env induced significantly higher levels of antibody responses against the Env protein in comparison to the construct for the wild type Env. Our results suggest that the YXXΦ motif in the HIV Env cytoplasmic domain may play a role in virus evasion of host immune responses through affecting its immunogenicity
Surface Stability and Immunogenicity of the Human Immunodeficiency Virus Envelope Glycoprotein: Role of the Cytoplasmic Domain
No full textBlockage of regulatory T cells augments induction of protective immune responses by influenza virus-like particles in aged mice
No full textMutations in the cytoplasmic tail of murine leukemia virus envelope protein suppress fusion inhibition by R peptide
No full textThis article cites 51 articles, 31 of which can be accessed fre
Enhanced immunogenicity of SIV Gag DNA vaccines encoding chimeric proteins containing a C-terminal segment of Listeriolysin O
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