PubMed Health⌕ Search

SEARCH · PubMed Health

Results for “SEMLIKI FOREST VIRUS”

Explore indexed PubMed citations for clinical trials, systematic reviews and public health research. Read source abstracts and follow each citation to its original PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 37 records · Page 2Linked to original sources
PubMed1998

The role of low pH and disulfide shuffling in the entry and fusion of Semliki Forest virus and Sindbis virus.

Semliki Forest virus (SFV), an enveloped alphavirus, infects cells via a membrane fusion reaction that is induced by the low pH in endocytic vesicles. The role of low pH in the entry of the alphavirus Sindbis virus (SIN) is unclear, and an alternative fusion mechanism involving receptor-induced disulfide bond rearrangements at neutral pH has been proposed. The entry properties of SFV and SIN were here compared in parallel using treatment with the weak base NH4Cl or the vacuolar ATPase inhibitors bafilomycin A-1 or concanamycin to neutralize endosome pH. Three membrane impermeant thiol modifying reagents, 5,5'-dithio-bis(2-nitrobenzoic acid) (DTNB), p-chloromercuriphenylsulfonic acid (pCMBS), and monobromotrimethylammoniobimane (Thiolyte MQ), were used to inhibit thiol-disulfide exchange reactions. Primary infection by both SFV and SIN was inhibited by neutralization of endosome pH using NH4Cl, bafilomycin, or concanamycin. The concentration of NH4Cl or bafilomycin required for inhibition correlated with the pH dependence of membrane fusion for SFV, SIN, and a pH-shift mutant of SFV. SFV and SIN infection were partially inhibited by the thiol blocker DTNB, but not by pCMBS or Thiolyte MQ. Our data suggest that acidic endosomal pH induces the fusion activity of both SFV and SIN during virus infection.

4-Chloromercuribenzenesulfonate↗
PubMed1975

Virus development in enucleate cells: echovirus, poliovirus, pseudorabies virus, reovirus, respiratory syncytial virus and Semliki Forest virus.

A group of RNA viruses, echovirus, poliovirus, reovirus, respiratory syncytial virus and Semliki Forest virus have been examined for ability to grow in enucleate African green monkey kidney (BSCi) cells. Semliki Forest virus produced an almost normal yield of virus but poliovirus, echovirus, reovirus and respiratory syncytial virus, although showing clear evidence of virus replication when compared with a nuclear DNA virus (pseudorabies virus) gave much lower yields than those from nucleate cells. Analysis of enucleate cells infected with echovirus and reovirus showed no evidence of a specific block in the synthesis of any virus-specified polypeptide. Infection with vesicular stomatitis virus at intervals after enucleation demonstrated a diminishing ability to support virus growth with increasing time. It is suggested that the yield of virus obtained from an enucleate cell is related to the length of the growth cycle of the virus, the reduced yield obtained with some viruses reflecting the declining ability of the enucleate cell to support virus growth.

Autoradiography↗
PubMed2004

Human immunodeficiency virus type 1-derived lentivirus vectors pseudotyped with envelope glycoproteins derived from Ross River virus and Semliki Forest virus.

Ross River virus (RRV) and Semliki Forest virus (SFV) are two alphaviruses that have a high degree of amino acid homology, as well as a very broad host range. We show here that envelope glycoproteins derived from both viruses can pseudotype human immunodeficiency virus type 1 (HIV-1)-derived lentivirus vectors. Both RRV and SFV glycoproteins considerably expand the host range of the lentivirus vector, and vectors can be efficiently concentrated by ultracentrifugation. A systematic analysis comparing the alphaviral glycoproteins to the vesicular stomatitis virus glycoprotein (VSV-G) revealed that lentivirus vectors incorporate RRV glycoproteins with an efficiency comparable to that of VSV-G. Both pseudotypes have comparable physical titers, but infectious titers with the RRV pseudotype are lower than with VSV-G. Incorporation of SFV glycoproteins into lentivirus vector is less efficient, leading to decreased physical and infectious titers. The transduction rates with VSV-G-, RRV-, and SFV-pseudotyped lentivirus vectors into adherent cell lines can be significantly increased by using a combination of Polybrene and plates coated with CH-296 recombinant fibronectin fragments. Together, our data suggest that RRV and SFV glycoproteins might be suitable as alternatives to VSV-G for pseudotyping lentivirus vectors.

Animals↗
PubMed1981

Evidence for a separate signal sequence for the carboxy-terminal envelope glycoprotein E1 of Semliki forest virus.

When Semliki Forest virus temperature-sensitive mutant ts-3 was grown at the restrictive temperature an aberrant nascent cleavage of the 130,000-dalton structural polyprotein took place relatively frequently. This cleavage yielded an abnormal 86,000-dalton fusion protein (p86) consisting of the amino-terminal capsid protein linked to the amino acid sequences of envelope protein p62 (a precursor of E3 and E2). The other cleavage product was the carboxy-terminal envelope protein E1. p86 was not glycosylated and was sensitive to the action of protease in the microsomal fraction, whereas E1 was glycosylated and protected from proteases, indicating that it had been segregated into the cysternal side of the microsomal vesicles. All attempts to show the E1 protein at the cell surface have failed so far, suggesting that it remains associated with intracellular membranes. When ts-3-infected cells labeled at the restrictive temperature were shifted to the permissive temperature the only labeled protein released with the virus particles was E1, indicating that E1, synthesized at the restrictive temperature, was competent to participate in the virus assembly. These results suggest strongly that there are two separate signal sequences for the envelope proteins of Semliki Forest virus. One follows the capsid protein as shown previously, and the other is for the carboxy-terminal E1. Even if the insertion of the amino-terminal envelope protein (p62) fails due to a cleavage defect, the other signal sequence can operate independently to guide the E1 through the endoplasmic reticulum membrane.

Amino Acid Sequence↗
PubMed1994

Membrane protein lateral interactions control Semliki Forest virus budding.

Semliki Forest virus, SFV, directs the synthesis of two membrane proteins, p62 and E1, which form a p62E1 heterodimer in the endoplasmic reticulum. After being transported to the plasma membrane (PM), they are incorporated into the virus membrane during the process of virus budding. Electronmicroscopic analyses of the envelope in matured virus show that the heterodimers are clustered into trimeric structures (spikes) which further form a regular surface lattice with T = 4. In this work we have used a genetic approach to study the importance of the trimerization event for virus budding. We have coexpressed a budding competent form of the virus heterodimer with another one which cannot be used for particle formation because of a defect in nucleocapsid (NC) binding. We show that the NC binding-deficient heterodimer is able to inhibit the budding of the competent one in a concentration-dependent manner and that the NC binding-competent heterodimers can rescue the incompetent ones into virus particles. This suggests that the heterodimers are complexed together, probably into the trimeric structures (spikes), at the PM to expose a multivalent binding site for the NC and thereby drive efficient virus budding.

Amino Acid Sequence↗
PubMed1995

Generation of cytotoxic and humoral immune responses by nonreplicative recombinant Semliki Forest virus.

The Semliki Forest virus (SFV) expression system can be used to package recombinant RNA into infectious suicide particles. Such RNA encodes only the SFV replicase and the heterologous protein but no structural proteins of SFV, and it is thus deficient in productive replication. We demonstrate here that infection of C57BL/6 (H-2b) and BALB/c (H-2d) mice with recombinant SFV expressing the nucleoprotein (NP) of influenza virus (SFV-NP) resulted in efficient priming of influenza virus-specific CD8+ cytotoxic T-cell (CTL) responses. The generated CTLs lysed both homologous (A/PR/8/34) and heterologous (A/HK/68) influenza virus-infected, or peptide-coated, target cells to a similar degree as CTLs induced by wild-type influenza virus in a major histocompatibility complex class I-restricted fashion. As few as 100 infectious units of virus induced a strong CTL response. Induction of CTL by SFV-NP could also be achieved in CD4 gene-targeted mice, demonstrating the independence of the primary CTL response of CD4+ helper T cells. One immunization generated a CTL response that peaked after 1 week, and an additional booster injection generated a CTL memory, which was still detectable after 40 days. SFV-NP immunizations also generated high-titered IgG humoral responses that remained significant after several months. These results demonstrate that the recombinant SFV suicide system is highly efficient in antigen presentation and suggest that it may have a potential as a recombinant vaccine.

Amino Acid Sequence↗
PubMed1997

Multiple mechanisms for the inhibition of entry and uncoating of superinfecting Semliki Forest virus.

Recombinant Semliki Forest viruses (SFV) that express one or none of the viral structural proteins were used to infect cells and to analyze the fate of incoming superinfecting wild-type viruses. It was found that in addition to the previously described block in replication that superinfecting viruses encounter within 15 min of infection, other mechanisms of superinfection inhibition occurred at later times. Over a 6-hr infection period, inhibition was seen in binding of virus to the cell surface, in acid-activated penetration into the cytoplasm, and in uncoating of nucleocapsids. For each of these processes, the inhibitory mechanism was investigated. In summary, we found that infection evoked several independent mechanisms for blocking the entry and uncoating of superinfecting viruses. The results also offered new insights into the normal processes of penetration and uncoating of SFV.

Animals↗
PubMed1996

Cholesterol-depleted cells that are relatively permissive for Semliki Forest virus infection.

Semliki Forest virus (SFV), an enveloped alphavirus, Infects cells by endocytosis followed by low pH-triggered fusion of the virus and endocytic vesicle membranes. Progeny virus is released by budding from the cell plasma membrane. In vitro, SFV fusion with artificial liposomes is triggered by low pH and is dependent on the presence of cholesterol and sphingolipid in the target liposome membrane. In tissue culture, both SFV fusion and virus exit are strongly cholesterol-dependent when assayed in cholesterol-depleted insect cells. We here describe the preparation of insect cells that while not containing detectable amounts of cholesterol, have adapted to sterol-depleted conditions, resulting in a more permissive phenotype for SFV infection. Although still less efficient at supporting SFV infection than control cholesterol-containing cells, the adapted cells show a 45-fold increase in primary infection by SFV, increased release of progeny virus, and enhanced virus growth kinetics compared to nonadapted cholesterol-depleted cells. The adapted cells are also about 85-fold more permissive for low pH-induced fusion of SFV with the plasma membrane, suggesting that adaptation correlates with a change in the cell membrane.

Adaptation, Physiological↗
PubMed1998

Rapid, high level protein production using DNA-based Semliki Forest virus vectors.

Semliki Forest virus (SFV) vectors can be produced faster, and have a wider host range, than baculovirus vectors. However, the original SFV system requires in vitro manipulation of RNA. We have generated a system that is wholly DNA-based. Both the replicon vector, encoding SFV polymerase and the protein of interest, and the helper vector, encoding viral structural proteins, were modified so that expression was RNA polymerase II-dependent. Transfection of the modified replicon plasmid alone generated 20-30-fold more protein than obtained from a simple expression vector. Expression required the SFV replicase, which amplifies replicon RNA. The SFV-based vector generated 10-20-fold more protein than a plasmid based on Sindbis virus. Cotransfection of SFV replicon and helper vectors generated viral titers of around 10(6) infectious particles/ml. A single electroporation, plated on one 10-cm plate, generated enough virus (10(7) particles) to produce >500 microg of protein. Wild type, replication proficient virus was not detected in three tests utilizing almost 10(8) viral particles, a distinct advantage over a DNA Sindbis-based system in which over half the virus particles generated are fully infectious. The new SFV vectors significantly enhance the utility of this expression system.

Animals↗
PubMed2003

Inhibition of matrix metalloproteinases ameliorates blood-brain barrier disruption and neuropathological lesions caused by avirulent Semliki Forest virus infection.

Semliki Forest virus (SFV) infection of mice is a useful model of viral neuropathogenesis in animals and avirulent strains such as SFV-A7 induce immune-mediated demyelination and death of neurones by necrosis and apoptosis. Matrix metalloproteinases (MMPs) have been implicated in various diseases including arthritis and cancer in many species. In this report, we show that MMP-2 and MMP-9 expression is induced in the brains of mice infected i.n. with SFV-A7. Treatment of mice with the pan MMP inhibitor GM6001 ameliorated the development of SFV-induced neuropathological lesions via an effect on the integrity of the blood-brain barrier. Low levels of neuronal necrosis and demyelination in GM6001-treated mice correlated with localisation of fibrinogen staining to thin-walled blood vessels and less intense staining of the perivascular neuropil.

Alphavirus Infections↗
PubMed2003

Role of interferon-gamma and nitric oxide in the neuropathogenesis of avirulent Semliki Forest virus infection.

Semliki Forest virus (SFV) infection of mice provides a useful model for the analysis of viral neuropathogenesis. In this study, the roles of interferon (IFN)-gamma and nitric oxide (NO) in the pathogenesis of SFV infection were assessed using mice deficient in inducible nitric oxide synthase (iNOS-/-), an enzyme important in the production of NO, and mice deficient in IFN-gamma receptor (IFN-gammaR-/-). Gene-knockout and wildtype mice were infected intranasally with the avirulent A7 strain of SFV and neuropathological lesions were correlated with levels of IFN-gamma, tumour necrosis factor (TNF)-alpha and interleukin (IL)-10 in the olfactory bulbs and frontal cortex. Lesions in IFN-gammaR-/- mice were characterized by higher levels of neuronal necrosis than in wildtype mice. The higher levels of neuronal necrosis were associated with increased levels of SFV antigen in neurones and increased numbers of macrophages and B cells. Relative differences in the severity of demyelination between IFN-gammaR-/- and wildtype mice were not detected. Similar levels of neuronal necrosis and SFV antigen labelling occurred in iNOS-/- mice and wildtype mice and levels of demyelination and macrophage infiltration in the iNOS-/- mice were lower than those in the wildtype strain. A rapid, but transient increase in the concentration of IFN-gamma was demonstrated in the frontal cortex of all infected mice samples. IL-10 levels in the frontal cortex and olfactory bulbs of SFV-infected iNOS-/- mice exceeded those present in the wildtype mice. This study, taken with our previous reports, provides further evidence that type 1 T cell responses are important in the control of brain viral clearance and the prevention of neuronal necrosis, but not in the development of demyelination.

Alphavirus Infections↗
PubMed1976

Anti-viral activity against encephalomyocarditis virus and Semliki Forest virus and acute toxicity of poly I and poly C administered sequentially to mice.

Mice are protected against lethal intraperitoneal and intravenous infection by encephalomyocarditis virus and Semliki Forest virus by sequential treatment with poly I followed by either polyC or poly5-hydroxyC without production of interferon when the treatments are 4 or more hours apart and by the intraperitoneal or intravenous routes. Maximum protection occurs around 4 hours before infection and is still significant 20 hours after infection. Treatments with combinations of other homoribopolynucleotides were not found to be anti-viral. Protection by sequential polyI, polyC treatment of mice is relatively short-lived and does not 'hypo-reactivate' the protective effect of polyI:C and shows approximately half the protective effect of polyI:C. The toxicity of sequential polyI, polyC treatment is lower than that of polyI:C particularly if poly5-hydroxyC is substituted for polyC. Silica treatment of mice indicates that stationary macrophages are required for protection by polyI followed by polyC but an effect on humoral or cell mediated immune responses does not appear to be involved. The effect appears to be a synergism between the protection conferred by polyI or polyC alone.

Animals↗
PubMed1983

Effect of vesicular stomatitis virus and Semliki Forest Virus on uptake of nutrients and intracellular cation concentration.

BHK-21 cells showed an increased ability to concentrate 2-deoxy-D-glucose (dGlc) 2 to 3 h after infection with vesicular stomatitis virus (VSV) or Semliki Forest virus (SFV), which began to be released at 2 and 3 h post-infection respectively; uptake of other nutrients was not affected in this way. Intracellular Na+ was either unchanged (VSV-infected cells) or increased (SFV-infected cells); K+ content was unchanged. These results do not support the current hypothesis that a non-specific increase in membrane permeability occurs in cells infected with rhabdoviruses or togaviruses.

Animals↗
PubMed1976

Anti-viral activity of single-stranded homopolynucleotides against encephalomyocarditis virus and Semliki Forest virus in adult mice without interferon induction.

Single administrations of poly C or poly I are anti-viral against infections of encephalomyocarditis (EMC) and Semliki Forest virus (SFV) in mice but poly U and poly A are not. The degree of protection is dose-dependent and mice which die do so at a later time when untreated controls even in a strain of mouse in which the time of death is not dependent on the dose of virus given. No circulating interferon is found after treating mice with poly C or poly I even at polynucleotide doses which give the same degree of protection as the interferon inducer, poly I:C. Several additional features distinguish the protection by poly C and poly I from interferon induction: the effect is low 24h before infection and maximal 6 h before infection, the effect is short-lived and mice do not show hypo-reactivation to repeated treatment. Limited treatment of mice with poly I:C, interferon or poly C before infection itself results in additional protection when poly C is also administered after infection, indicating that poly C has an effect after onset of virus replication. After infection poly C and poly I are both more effective by the intravenous route but before infection they are most effective when administered by the same route as the virus. Quantitative comparisons of the protective effects of poly C, poly I and the interferon inducer, poly I:C, are possible from dose response curves of the polynucleotides at different times relative to infection and by different routes of administration. The results are considered in relation to the presence of homopolyribonucleotide tracts in the viral genomes and effects on the reticulo-endothelial system of the mice.

Animals↗
PubMed2006

Assembly of trans-encapsidated recombinant viral vectors engineered from Tobacco mosaic virus and Semliki Forest virus and their evaluation as immunogens.

RNA virus vectors are attractive vaccine delivery agents capable of directing high-level gene expression without integration into host cell DNA. However, delivery of non-encapsidated RNA viral vectors into animal cells is relatively inefficient. By introducing the tobacco mosaic virus (TMV) origin of assembly into the RNA genome of Semliki Forest virus (SFV), we generated an SFV expression vector that could be efficiently packaged (trans-encapsidated) in vitro by purified TMV coat protein (CP). Using cellular assays, pseudovirus disassembly, RNA replication and reporter gene expression were demonstrated. We also evaluated the immune response to trans-encapsidated recombinant SFV carrying a model antigen gene (beta-galactosidase) in C57/B6 mice. Relative to RNA alone, vector encapsidation significantly improved the humoral and cellular immune responses. Furthermore, reassembly with recombinant TMV CPs permitted the display of peptide epitopes on the capsid surface as either genetic fusions or through chemical conjugation, to complement the immunoreactivity of the encapsidated RNA genetic payload. The SFV vector/TMV CP system described provides an alternative nucleic acid delivery mechanism that is safe, easy to manufacture in vitro and that also facilitates the generation of unique nucleic acid/protein antigen compositions.

Animals↗
PubMed2002

Expression of open reading frame 5 protein of porcine reproductive and respiratory syndrome virus using semliki forest virus expression system.

The ORF5 gene encodes a major envelope glycoprotein (GP5), which is one of the three major proteins of porcine reproductive and respiratory syndrome virus (PRRSV). The GP5 protein has been known to be a 24.5-26 kDa N-glycosylated envelope protein. The GP5 is involved in inducing neutralizing antibodies. For this reason, the GP5 is primary candidate for the PRRSV subunit vaccine. To produce the native form of GP5 in mammalian cells, we have cloned the ORF5 gene from PRRSV CNV-1 into the Semliki Forest virus (SFV)-based expression vector, resulting in recombinant pSFV-ORF5. By the infection with recombinant pSFV-ORF5 to BHK-21 cells, the GP5 expression was confirmed by immunocytochemistry and immunoblotting assay. The recombinant virus particle harboring ORF5 gene was infectious to BHK-21 and MARC-145. The RNA synthesis and expression of GP5 in the infected cell was also confirmed by RT-PCR.

Animals↗
PubMed2000

Replicon vectors derived from Sindbis virus and Semliki forest virus that establish persistent replication in host cells.

Alphavirus replicon vectors are well suited for applications where transient, high-level expression of a heterologous gene is required. Replicon vector expression in cells leads to inhibition of host macromolecular synthesis, culminating in eventual cell death by an apoptotic mechanism. For many applications, including gene expression studies in cultured cells, a longer duration of transgene expression without resulting cytopathic effects is useful. Recently, noncytopathic Sindbis virus (SIN) variants were isolated in BHK cells, and the mutations responsible were mapped to the protease domain of nonstructural protein 2 (nsP2). We report here the isolation of additional variants of both SIN and Semliki Forest virus (SFV) replicons encoding the neomycin resistance gene that can establish persistent replication in BHK cells. The SIN and SFV variant replicons resulted from previously undescribed mutations within one of three discrete regions of the nsP2 gene. Differences among the panel of variants were observed in processing of the nonstructural polyprotein and in the ratios of subgenomic to genomic RNAs. Importantly, high-level expression of a heterologous gene was retained with most replicons. Finally, in contrast to previous studies, efficient packaging was obtained with several of the variant replicons. This work expands the utility of noncytopathic replicons and the understanding of how alphavirus replicons establish persistent replication in cultured cells.

Genetic Vectors↗