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temperature-sensitive host-dependent mutants of sindbis virus. 19817281510
establishment of persistent infection in mouse cells by sindbis virus and its temperature-sensitive mutants.the ability of wild-type (wt) sindbis virus and six temperature-sensitive (ts) mutants to establish persistent infection in mouse l cells and a line of mouse embryo (me) cells was determined. the wt established persistent infection in both me cells and l cells at 39 degrees c. at 30 degrees c the wt established persistent infection in l cells but not me cells, which did not recover from the initial infection. for the ts mutants, both cell lines survived the initial infection at 39 degrees c (the ...19817288408
a sindbis virus mutant temperature-sensitive in the regulation of minus-strand rna synthesis. 19817292988
specific and non-specific immunological changes in epidemic polyarthritis patients.t-lymphocytes from epidemic polyarthritis patients exhibited a virus specific proliferative response when exposed to ross river virus in vitro. the magnitude of this response was greater than that of lymphocytes from asymptomatic seropositive donors. the natural killer cell activity of peripheral blood mononuclear leucocytes from these patients was depressed early in disease but returned to normal levels as the severity of the symptoms decreased. although no in vivo role can yet be assigned to n ...19817325890
design and preparation of affinity columns for the purification of eukaryotic messenger ribonucleic acid cap binding protein.2',3'-o-[1-(2-carboxyethyl) ethylidene]-7-methylguanosine 5'-diphosphate (5) and 7-(5-carboxypentyl) guanosine 5'-diphosphate (13) have been synthesized and immobilized on ah-sepharose 4b to the extent of 17.4 and 36.6 mumol of ligand/g of gel, respectively. the affinity resins thus derives were employed in columns for the purificaton of 24k cap binding protein (cbp) from rabbit reticulocytes. each resin was found to retain the protein of interest; elution of 24k cbp could then be effected by wa ...19817306524
studies on the intracellular distribution of sindbis messenger rna in infected chick-embryo fibroblasts. 1. presence of extrapolyribosomal 26-s rna in the membrane fraction.four hours after infection with sindbis virus, chick embryo fibroblasts were studied by electron microscopy and cell fractionation. electron microscopy of infected and non-infected cells revealed that infection produced a disaggregation of polyribosomes into monomers. apart from this observation most cells appeared well preserved, and no degranulation of the rough endoplasmic reticulum was visible. analysis of postnuclear supernatants by sucrose density gradients showed that no change in the rel ...19807358048
studies on the intracellular distribution of sindbis messenger rna in infected chick embryo fibroblasts. 2. non-parallel distribution of 26-s rna and ribosomes within microsomal subfractions.the submicrosomal distribution of membrane-associated viral 26-s rna in chick embryo fibroblasts infected with sindbis virus was studied. infected chick embryo fibroblasts were homogenized in the presence of low amounts of edta and fractionated by differential centrifugation. analysis of postmitochondrial supernatants by isopycnic flotation on continuous sucrose gradients showed that membrane-associated 26-s rna and ribosomes were not distributed in parallel, with an enrichment in 26-s rna in th ...19807358049
actinomycin d cytotoxicity for mouse peritoneal macrophages and effect on lactate dehydrogenase-elevating virus replication.treatment of lactate dehydrogenase-elevating virus(ldv)-infected mouse peritoneal macrophage cultures with actinomycin d (1 microgram/ml) resulted in a progressive reduction in the formation of ldv-specific rna and mature virus as the time of incubation with actinomycin d increased beyond 2-3 h. this effect, however, seemed to reflect an unusual sensitivity of macrophages to toxic effects of actinomycin d. macrophage cytotoxicity and lysis became apparent 3-4 h after addition of actinomycin d; t ...19807358507
identification of a cell-surface protein involved in the binding site of sindbis virus on human lymphoblastic cell lines using a heterobifunctional cross-linker.a cell-surface protein of molecular weight 90000 was identified as the major protein located at or near the binding site of sindbis virus on the lymphoblastic cell lines jy and daudi. this sindbis-virus-binding protein was detected by cross-linking sindbis virus, modified with 4-[6-formyl-3-azidophenoxy]butyrimidate, to the cell surface. the applicability of this compound in the identification of specific cell-surface proteins was demonstrated using a monoclonal antibody reactive with beta 2-mic ...19817014219
the role of complement in viral infections. iii. activation of the classical and alternative complement pathways by sindbis virus. 19807365263
the role of complement in viral infections. ii. the clearance of sindbis virus from the bloodstream and central nervous system of mice depleted of complement.the following studies were performed to investigate the mechanism(s) by which the complement system limits sindbis virus infection in the central nervous system of mice. after the intracerebral inoculation of sindbis virus, no differences in mortality or viral growth in the central nervous system were observed between normal mice and mice depleted of complement by treatment with cobra venom factor. in addition, animals that had been inoculated subcutaneously with sindbis virus and depleted of co ...19807365277
characterization of a small, nonstructural viral polypeptide present late during infection of bhk cells by semliki forest virus.bhk cells, late in infection with semliki forest virus, were found to contain a small virus-specific polypeptide not found in the mature virion. this polypeptide had an apparent molecular weight of 6,000 and is referred to here as the 6k protein. no [2-3h]mannose was incorporated into 6k, and hence it does not appear to be a glycoprotein. this protein appears to be a primary translation product of the subgenomic 26s mrna, which encodes the viral structural proteins. the genes encoding the viral ...19807365868
inhibition of sindbis virus replication in aedes albopictus cells by virazole (ribavirin) and its reversal by actinomycin: a correction. 19807368572
differential sensitivity of infectious and defective-interfering particles of sindbis virus to ultraviolet irradiation. 19807368573
partial amino acid sequences of sindbis and semliki forest virus-specific core proteins. 19807368574
establishment and maintenance of persistent infection by sindbis virus in bhk cells.we have established a persistent infection of bhk cells with a preparation of sindbis virus heavily enriched in defective interfering (di) particles. the small fraction of cells that survived the initial infection grew out to form a stable population of cells [bhk(sin-1) cells], most of which synthesized viral rna and viral antigens. the presence of di particles in this virus stock was required to establish this persistent state. bhk(sin-1) cells released a small-plaque, temperature-sensitive vi ...19807365871
reversible transport defects of virus membrane glycoproteins in sindbis virus mutant infected cells.surface immunofluorescence with anti-envelope serum was used to study the transport of virus membrane glycoproteins in cells infected with temperature-sensitive mutants of sindbis virus. mutants belonging to complementation group d (ts-10 and ts-23), which have a defect in the envelope protein e1, showed a temperature dependent defect in the transport of the virus glycoproteins to the cell surface. in contrast, cells infected with mutant ts-20, the only representative of complementation group e, ...19807379138
defective mutant of sindbis virus with a smaller-molecular-weight form of the e1 glycoprotein.we have isolated from a single plaque a mutant of sindbis virus characterized by an e1 glycoprotein with higher electrophoretic mobility. this higher mobility is not attributable to a different extent of glycosylation of the protein nor to an altered proteolytic maturation pathway of the polypeptide precursor, but is the result of a deletion occurring during the replication of the viral rna. the 26s rna (the messenger for the sindbis structural proteins) extracted from cells infected with the mu ...19807381993
sindbis virus rna replication. ii. strand composition and metabolic fate of the multi-stranded rna species.double-stranded rna from sb virus-infected cells was denatured and analysed on agarose-methylmercuric hydroxide gels. equimolar amounts of three single-stranded species with mol. wt. of 4 x 10(6), 2.5 x 10(6) and 1.8 x 10(6) were found. pulse and chase experiments in infected cells established a precursor-product relationship between the multi-stranded and single-stranded virus rna species. the present results support the model in which the 49s and 26s species of virus rna are synthesized in inf ...19807381436
alphaviruses in peninsular malaysia: ii. serological evidence of human infection.a serum survey of several characteristic groups of humans in urban, rural, and forested areas of peninsular malaysia for evidence of infection with three alphaviruses (sindbis, getah, and chikungunya) was made on 4384 specimens collected between 1965 and 1969. analysis of the serological results indicated that 1) persons residing in predominantly rural and forested areas have higher frequencies of specific alphavirus antibody of all three viruses than persons residing in urban areas, 2) human in ...19807403943
dna: chapter of accidents at san diego. 19807403872
a case of sindbis virus infection in zimbabwe. 19807407896
separation of alphavirus nucleocapsids from envelope fragments.methods for the preparation of alphavirus nucleocapsids and envelope fragments for biochemical, electron microscopical and serological investigations are described. difficulties which have been encountered previously in attempts to isolate viral nucleocapsids for morphological studies, namely disruption of the fragile particles, have been overcome by treatment with formaldehyde.19807405358
analysis of semliki-forest-virus structural proteins to illustrate polyprotein processing of alpha viruses.the four structural proteins of semliki forest virus were purified in an amount of 30-50 nmol by preparative sodium dodecylsulfate/polyacrylamide gel electrophoresis. each protein was subjected to n-terminal structural analysis by degradation in a liquid-phase sequencer. about 20 residues were determined for each of the two membrane glycoproteins e1 and e2. the amino acid sequence of e1 but not that of e2 showed extensive homology to the corresponding proteins of the closely related sindbis viru ...19807408852
[action of remantadine on virus-specific polypeptide synthesis in a sindbis virus-infected cell culture].effect of remantadine (alpha-methyl-i-adamantane methylamine) on reproduction and synthesis of the virus-specific polypeptides was studied in culture of chicken embryo fibroblasts, infected with virus syndbis. the preparation inhibited the virus reproduction as well as the formation of virus-specific proteins after addition into the cultural medium together with virus, immediately after absorption and within 1 and 2 hrs after absorption. the data obtained suggest a possibility of an immediate ef ...19807423886
application of high-field proton magnetic resonance spectroscopy in the structural determination of membrane-derived sindbis virus glycopeptides.sindbis virus membrane glycopeptides have ben purified in chemical quantities and their oligosaccharide structures analyzed by 1h nmr spectroscopy at 360 mhz. interpretable spectra could be obtained with approximately 100 micrograms of oligosaccharide. spectral analysis of the sialyl glycopeptides s1, s2, and s3 at high and low temperatures confirms their structures to be nana alpha (2,3)gal beta (1,4)-glcnac beta (1,2)man alpha (1,6)-[nana alpha (2,3)gal beta (1,4)-glcnac beta (1,2)man alpha (1 ...19817317384
virion nucleic acid of ebola virus.the virion nucleic acid of ebola virus consists of a single-stranded rna with a molecular weight of approximately 4.0 x 10(6). the virion rna did not bind to oligodeoxythymidylic acid-cellulose under conditions known to bind rnas rich in polyadenylic acid and was not infectious under conditions which yielded infectious rna from sindbis virus, suggesting that ebola virus virion nucleic acid is a negative-stranded rna.19807431486
[combined effect of remantadine (alpha-methyl-1-adamantane methylamine) and ribovirin (1-beta-d-ribofuranosyl-1,2,4-triazole-3-carboxamide) on the reproduction of sindbis virus in cell culture].the influence of remantadine, ribovirine, and combination thereof on sindbis virus reproduction in cell cultures was studied. comparison of remantadine and ribovirine showed the former to be a stronger inhibitor of sindbis virus reproduction. the results of the combined effect of both drugs in chick embryo fibroblast cultures showed undoubtful advantages of this treatment over the use of each of the drugs alone. in studies of the effect of ribovirine on synthesis of virus-specific proteins and r ...19807434742
host cell receptors for two strains of sindbis virus.experiments were performed to determine whether neuronal cells have different numbers of receptors for a neurovirulent and an avirulent strain of the same virus and whether neurotropic strains of different viruses share the same cellular receptors. attempts were also made to characterize the receptors for the two strains of sindbis virus on viable cells by studying their enzyme sensitivity. the number of cellular receptors available for sindbis virus attachment to several cell lines was determin ...19807436739
genomic divergence among sindbis virus strains. 19807414959
cell and virus propagation on cylindrical cellulose based microcarriers.microgranular deae-cellulose anion exchanger with exchange capacities ranging between 0-2.7 meq/gr dry materials were tested for their suitability to be used as microcarriers for culturing several anchorage-dependent-cells. it was found that cells from established cell lines (bhk, mdck), grew as confluent monolayer on deae-cellulose microcarriers with exchange capacity ranging between 1.00-2.0 meq/gr dry materials. primary cells (chick embryo fibroblast) grow as a cell-microcarriers aggregate on ...19817341288
unsaturated free fatty acids inactivate animal enveloped viruses.unsaturated free fatty acids such as oleic, arachidonic or linoleic at concentrations of 5-25 microgram/ml inactivate enveloped viruses such as herpes, influenza, sendai, sindbis within minutes of contact. at these concentrations the fatty acids are inocuous to animal host cells in vitro. naked viruses, such as polio, sv40 or emc are not affected by these acids. saturated stearic acid does not inactivate any viruses at concentrations tested. though the mode of action of unsaturated fatty acids i ...19807447706
relationship between enzymatic modifications of serum low density lipoproteins and their haemagglutination inhibiting activity towards sindbis virus.the chemical structure of human serum low density lipoproteins (ldl) involved in the inhibition of the haemagglutination by sindbis virus was studied. separation of lipoproteins into their protein and lipid components demonstrated that the inhibiting activity is almost completely due to the lipid moiety. the treatment of ldl and extracted lipoprotein lipids by phospholipase a2, c and d produced different effects and in particular a marked increase in the hi titre by the action of phospholipase a ...19807425849
solid-phase immune electron microscopy (spiem) for rapid viral diagnosis.immune electron microscopy (iem) is one of the fastest and most sensitive methods for the detection and diagnosis of viruses. this technique is based on formation of immune complexes of the virus with its corresponding antibody. in iem optimal precipitation depends on a correct ratio, and there is a prozone effect. these problems can be overcome by using the solid-phase immune electron microscopic (spiem) technique. in this technique the antibody is attached to a particle which is used for 'fish ...19807452001
sindbis virus. i. gene order of translation in vivo. 19807456333
restricted initiation of protein synthesis on the potentially polycistronic sindbis virus 42 s rna.sindbis 42 s genome rna was isolated from virions and translated in vitro before and after purification by oligo(dt)-cellulose chromatography and sucrose density gradient centrifugation under denaturing conditions. in intact 42 s rna, only the 5'-proximal initiation site for the synthesis of nonstructural proteins was used. the internally located start site for viral structural protein formation was active in broken genome rna molecules where, as a consequence of fragmentation, it was closer to ...19807440550
evidence that sindbis virus infects bhk-21 cells via a lysosomal route.chloroquine and nh4cl, potent inhibitors of lysosomal function, decreased the production of infectious sindbis virus particles in bhk-21 cells by 10- and 12-fold, respectively. there were no apparent toxic effects on cells exposed to these lysosomotropic agents. these chemicals did not alter the rate of cellular protein synthesis, with the exception of a reversible twofold inhibition by chloroquine. no additive effects of chloroquine and nh4cl were observed when the cells were saturated with the ...19807459676
intracellular distribution of sindbis virus membrane proteins in bhk-21 cells infected with wild-type virus and maturation-defective mutants.the association of sindbis virus proteins with cellular membranes during virus maturation was examined by utilizing a technique for fractionating the membranes of bhk-21 cells into three subcellular classes, which were enriched for rough endoplasmic reticulum, smooth endoplasmic reticulum, and plasma membrane. pulse-chase experiments with wild-type (strain svhr) virus-infected cells showed that virus envelope proteins were incorporated initially into membranes of the rough endoplasmic reticulum ...19807463555
effects of anti-e2 monoclonal antibody on sindbis virus replication in at3 cells expressing bcl-2.antibodies directed to sindbis virus (sv) envelope protein e2 are able to control virus replication in vivo and in persistently infected cultures of neurons in vitro. we investigated the mechanisms by which anti-e2 monoclonal antibody (mab) alters virus replication by using at3 rat prostatic carcinoma cells expressing the inhibitor of apoptosis bcl-2. treatment of sv-infected at3-bcl-2 cells with anti-e2 mab g5 for 2 h decreased the rate of virus release for 6 to 8 h after removal of the antibod ...19957474120
antiviral activity of alpha interferon in sindbis virus-infected cells is restored by anti-e2 monoclonal antibody treatment.pretreatment of at3 rat prostatic carcinoma cells expressing the inhibitor of apoptosis bcl-2 (at3-bcl-2 cells) with alpha interferon (ifn-alpha) affected replication of a virulent strain of sindbis virus (sv) but did not protect against virus-induced cell death. treatment of cells with ifn-alpha late during infection affected ongoing sv replication very little. previous studies have shown that cross-linking of the viral glycoprotein e2 with antibody delays the inhibition of k+ influx by improvi ...19957474167
formation of the semliki forest virus membrane glycoprotein complexes in the infected cell.in semliki forest virus (sfv)-infected cells, all structural proteins are translated from a 26s mrna using a single initiation site. the capsid protein which is made first is released into the cytoplasm whereas the two membrane proteins, p62 (the precursor for e2 and e3) and e1, are inserted into the rough endoplasmic reticulum membrane. based on gradient centrifugation and cross-linking studies, it can be seen that the p62 and e1 polypeptides form a complex immediately after synthesis and migra ...19807441208
[the distribution of sbv nonstructural protein 2 (nsp2) in host cell].the distribution of sbv nsp2 has been observed by means of immunolabelling electron microscopy. the results showed that sbv nsp2 existed both in the cytoplasm and in the nucleus of host cell. when the cdna of sbv nsp2 was transfected in host cell, the nsp2 also accumulates in the nucleus.19957483582
immunoglobulins in the cerebrospinal fluid: changes during acute viral encephalitis in mice.the development and pathogenesis of immunoglobulin changes in the csf in acute encephalitis were studied in mice infected with sindbis virus. three phases of immunoglobulin changes were noted. 1) during the first 2 days when virus replication in the brain is maximal, few changes were seen in the csf. 2) at the time of maximal meningeal inflammation, 3 to 5 days after infection, there was a transudation of protein from serum into csf. 3) during the period of recovery, there was evidence of a loca ...19817451970
genesis of sindbis virus by in vivo recombination of nonreplicative rna precursors.genetically engineered rna transcripts coding for various sindbis virus (sin) genes were used to study structure and sequence requirements of rna recombination in bhk cells. three different groups of rna transcripts were made: (i) rnas which retain the ability to replicate and which carry sequences coding for either viral polymerase or viral structural proteins; (ii) rnas which lack the complete 3' end of the sin genome and thus are incapable of replicating; and (iii) rnas which lack the complet ...19957494243
evolution of the sindbis virus subgenomic mrna promoter in cultured cells.transcription of the subgenomic mrna of alphaviruses initiates at an internal site, called the promoter, which is highly conserved. to determine the functional significance of this conservation, we used an approach that randomizes positions -13 to -9 of the promoter to generate a library containing all possible sequences within this region, including the wild-type sequence. viruses in the mixed population with more-efficient promoters were selected for during passaging in mammalian (bhk-21) cell ...19957494287
host-dependent evolution of the sindbis virus promoter for subgenomic mrna synthesis.alphaviruses are alternately transmitted between arthropod and vertebrate hosts. in each host, the virus transcribes a subgenomic mrna that encodes the viral structural proteins which encapsidate the genome to form progeny virions. transcription initiates at an internal site called the promoter. to determine if promoter utilization varies in mammalian versus mosquito cells, we used these cells as hosts to select for active promoters among a library of different mutant promoters. compared with th ...19957494288
the expression of chloramphenicol acetyltransferase in mosquitoes and mosquito cells using a packaged sindbis replicon system.sindbis (sin) replicon virus was used to express chloramphenicol acetyltransferase (cat) in aedes albopictus (c6/36) cells and aedes triseriatus mosquitoes. rna transcribed in vitro from a sin replicon plasmid (psinrep5/cat) and from sin virus helper constructs (pdh-eb or pdh(26s)5'sin) was coelectroporated into bhk-21 cells to generate replicon viruses, designated rep5/cat/eb and rep5/cat/26s. c6/36 cells infected with rep5/cat/eb or rep5/cat/26s virus at a multiplicity of infection of 3, expre ...19957498436
expression of the rubella virus structural proteins by an infectious sindbis virus vector.to assess the potential of an infectious sindbis virus vector (dssin) as a eukaryotic expression vector, dssin recombinants which contain each of the rubella virus (rub) structural proteins (c, e2, and e1) individually were constructed. expression of the rub proteins by each dssin recombinant was robust and processing was similar to that observed when these proteins were expressed using other vectors. the c and e2 recombinants, each of which contains a cassette of less than 1,000 nts, were stabl ...19957503703
growth-dependent alterations in oligomannosyl glycopeptides expressed in sindbis virus glycoproteins.sindbis virus was used to examine host cell growth-dependent expression of oligomannosyl glycopeptides within single species of viral glycoproteins. four mannosyl oligosaccharides ranging from 5 to 8 mannose residues were separated on bio-gel p4 column chromatography. virus derived from rapidly growing chicken embryo fibroblasts displayed greater quantities of larger sized oligomannosyl glycopeptides in intact virus and in purified sindbis virus glycoproteins e1 and e2 compared with virus from n ...19807459334
putative receptor binding sites on alphaviruses as visualized by cryoelectron microscopy.the structures of sindbis virus and ross river virus complexed with fab fragments from monoclonal antibodies have been determined from cryoelectron micrographs. both antibodies chosen for this study bind to regions of the virions that have been implicated in cell-receptor recognition and recognize epitopes on the e2 glycoprotein. the two structures show that the fab fragments bind to the outermost tip of the trimeric envelope spike protein. hence, the same region of both the sindbis virus and ro ...19957479858
recombination between sindbis virus rnas.the sindbis virus rna genome is divided into two modules--one coding for the nonstructural protein genes and the other coding for the structural protein genes. in our studies of recombination, the two parental rnas were defective in different modules. analysis of the recombinant rnas demonstrated that the parental rnas each contributed its intact module and that the crossovers occurred within the defective modules. the recombinational events giving rise to infectious virion rnas could create del ...19947518272
recombinant sindbis virus as an expression system for cell biology. 19947529867
recognition of endogenous ecotropic murine leukaemia viruses by anti-akr/gross virus cytotoxic t lymphocytes (ctl): epitope variation in a ctl-resistant virus.akr/gross virus-specific cytotoxic t lymphocytes (ctl) from c57bl/6 (b6) mice are h-2kb-restricted and recognize epitopes encoded by the prototype endogenous ecotropic murine leukaemia virus (emv) akr623. four ctl epitopes have been identified by the use of synthetic peptides corresponding to akr623-encoded amino acid sequences. here we present both functional and nucleotide sequence data indicating that three closely related emv share all of these ctl epitopes. we also found that one other muri ...19957534814
hepatitis c and arboviral antibodies in the island populations of mauritius and rodrigues.a serological survey for antibodies to hepatitis c virus (hcv), dengue viruses (den), west nile virus (wn), and sindbis virus (sin) was carried out in sera of selected groups of the population of the islands of mauritius (n = 449) and rodrigues (n = 115), indian ocean. 8.3% of 564 sera were positive for anti-hcv. in mauritius, 2.1% of sera of healthy individuals were found with anti-hcv. the highest prevalence was found in sexually transmitted disease (std) patients and prison inmates with 46.2% ...19947534820
studies on the antiviral activity of protein kinase inhibitors against the replication of vesicular stomatitis virus.several derivatives of k-252a, a protein kinase inhibitor isolated from nocardiopsis sp., were investigated for their effects on the replication of vesicular stomatitis virus (vsv) in bhk-21 cell cultures. among those we tested, kt5926, which preferentially inhibits the myosin light chain kinase (mlck), suppressed the viral replication by 95-99% at 15 microm. k-252a, which inhibits a broad spectrum of cellular protein kinase, similarly affected the viral replication. other derivatives, kt5720 an ...19957550128
mosquito sensitivity to a scorpion neurotoxin expressed using an infectious sindbis virus vector.the scorpion, androctonus australis hector, produces an insect-specific toxin (aahit) encoded by the scotox gene. to assess the toxicity of aahit for mosquitoes, we have taken a novel approach to express the scotox gene in vivo. we have engineered a double subgenomic sindbis (dssin) virus that contains the scotox gene in the viral genome and intrathoracically inoculated the virus (te/3'2j/scotox) into mosquitoes (aedes aegypti, ae. triseriatus and culex pipiens), houseflies (musca domestica) and ...19957551198
double-subgenomic sindbis virus recombinants expressing immunogenic proteins of japanese encephalitis virus induce significant protection in mice against lethal jev infection.a series of double-subgenomic sindbis virus (dssin) recombinants that express cassettes encoding the immunogenic proteins of japanese encephalitis virus (jev) [prm-e, prm-e-ns1, ns1-ns2a, 80%e (encodes the amino-terminal 80% part of e), and ns1] were constructed and analyzed for their ability to confer protective immunity in mice against lethal challenge with neurovirulent jev. the cassettes were introduced into both 5' [second subgenomic promoter of the vector precedes the sin structural open r ...19957571428
equine arteritis virus subgenomic rna transcription: uv inactivation and translation inhibition studies.the expression of the genetic information of equine arteritis virus (eav), an arterivirus, involves the synthesis of six subgenomic (sg) mrnas. these are 5' and 3' coterminal since they are composed of a leader and a body sequence, which are identical to the 5' and 3' ends of the genome, respectively. previously, it has been suggested that cis-splicing of a genome-length precursor rna is involved in their synthesis. this was reevaluated in a comparative analysis of the sg rna synthesis of eav, t ...19957491761
nucleocapsid-glycoprotein interactions required for assembly of alphaviruses.we have studied interactions between nucleocapsids and glycoproteins required for budding of alphaviruses, using ross river virus-sindbis virus chimeras in which the nucleocapsid protein is derived from one virus and the envelope glycoproteins are derived from the second virus. a virus containing the ross river virus genome in which the capsid protein had been replaced with that from sindbis virus was almost nonviable. nucleocapsids formed in normal numbers in the infected cell, but very little ...19947508993
a mutant of sindbis virus which is released efficiently from cells maintained in low ionic strength medium.experiments many years ago showed that reducing the ionic strength of the medium sindbis virus-infected chick embryo fibroblasts were maintained prevented the release of progeny virus into the medium. it was proposed at that time that the susceptibility of virus release to inhibition by low ionic strength (lis) medium might be determined by the properties of the viral envelope proteins. we thought that if such were the case, it might be possible to obtain a viral mutant which is resistant to the ...19957618264
replication and tissue tropism of the alphavirus sindbis in the mosquito aedes albopictus.we have examined the replication and tissue distribution of the alphavirus sindbis in the mosquito aedes albopictus. parenteral inoculation of virus resulted in an acute infection accompanied by rapid virus replication and a persistent infection, during which total virus production was reduced. acute and persistent phase virus rna synthesis, virus production, and organ-specific distribution of infection were determined over an 18-day incubation period. organs were classified as refractory (ovari ...19957676618
iodine-mediated inactivation of lipid- and nonlipid-enveloped viruses in human antithrombin iii concentrate.human plasma-derived protein concentrates intended for clinical use must be treated for viral inactivation to ensure patient safety. this study explored the use of liquid iodine for inactivation of several lipid- and nonlipid-enveloped viruses in an antithrombin iii (at-iii) concentrate. iodine at levels of 0.01% to 0.02% caused between 43% and 94% loss of at-iii activity, as well as degradation of at-iii as shown by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (sds-page) and wester ...19957606009
structural rearrangement of infecting sindbis virions at the cell surface: mapping of newly accessible epitopes.sindbis virus glycoproteins e1 and e2 undergo a conformational alteration during early virus-cell interaction at the cell surface (d. flynn, w. j. meyer, j. m. mackenzie, jr., and r. e. johnston, j. virol. 64:3643-3653, 1990). certain epitopes normally internal on native virus become accessible to monoclonal antibody (mab) binding after attachment but before internalization of virus particles. these newly exposed epitopes, termed transitional epitopes, may be part of functionally important domai ...19937688818
rubella virus-specific cytotoxic t-lymphocyte responses: identification of the capsid as a target of major histocompatibility complex class i-restricted lysis and definition of two epitopes.the role of major histocompatibility complex (mhc) class i-restricted cd8+ cytotoxic t lymphocytes in immunity to rubella virus (rv) infection is unknown. lymphocytes of rv-immune individuals were prestimulated on an rv-infected mhc class i-matched (or partially matched) fibroblast monolayer which generated cd8+ lymphoblasts capable of lysing rv-infected fibroblast targets in a class i-restricted manner. using an infectious sindbis virus (sv) vector which expressed the rv capsid protein (sv/rubc ...19937690412
[the multiplication of sindbis virus in bhk-21 cell].one step growth curve of sindbis virus (sbv) in bhk-21 cells showed that a lot of progeny viruses could be observed at 2 hr. after infection. the titer of virus reached the highest about 10(9)tcid50/ml at 6hr. post infection. the viral morphological characters and morphogenetic process in bhk-21 cells were examined by electron microscopic techniques. the dynamics of viral proteins synthesis and the effect on host cell were studied and discussed in this paper.19937694428
polypeptide requirements for assembly of functional sindbis virus replication complexes: a model for the temporal regulation of minus- and plus-strand rna synthesis.proteolytic processing of the sindbis virus non-structural polyproteins (p123 and p1234) and synthesis of minus- and plus-strand rnas are highly regulated during virus infection. although their precise roles have not been defined, these polyproteins, processing intermediates or mature cleavage products (nsp1-4) are believed to be essential components of viral replication and transcription complexes. in this study, we have shown that nsp4 can function as the polymerase for both minus- and plus-st ...19947517863
aura virus is a new world representative of sindbis-like viruses.aura virus is an alphavirus present in brazil and argentina that is serologically related to sindbis virus (present throughout the old world) and to western equine encephalitis (wee) virus (present in the americas). we have previously shown that wee is a recombinant virus whose glycoproteins and part of whose 3' nontranslated region (ntr) are derived from a sindbis-like virus, but the remainder of whose genome is derived from eastern equine encephalitis (eee) virus. we show here that aura virus ...19957747434
sindbis vectors suppress secretion of subviral particles of japanese encephalitis virus from mammalian cells infected with sin-jev recombinants.double-subgenomic sindbis virus (dssin) recombinants that express cassettes encoding prm-e or a c-terminally truncated form of e of japanese encephalitis virus (jev) were constructed. the products were efficiently expressed in both mammalian and mosquito cell lines infected with the dssin recombinants. however, suppression of prm-e secretion from mammalian cells infected with dssin-prm-e recombinants was observed. this suppression was more pronounced late in infection (< 5% of total product was ...19957747465
viral safety of solvent/detergent-treated blood products.laboratory research that began in 1982 led to the licensing in the usa of a solvent/detergent (sd)-treated factor viii concentrate in 1985. the licence was granted on the basis of several factors. first, studies had demonstrated the inactivation of several marker viruses (vesicular stomatitis virus, sindbis virus, sendai virus) and other viruses such as human immunodeficiency virus (hiv), hepatitis b virus (hbv), and non-a, non-b hepatitis virus (nanbhv; now known principally to be hepatitis c v ...19947749045
isolation of the anthropogenic compound fluoranthene in a screening of chinese medicinal plants for antiviral compounds.thirty-one species of medicinal plants used in the treatment of diseases of viral origin in yunnan province of china were assayed for inhibition of sindbis and murine cytomegalovirus in mammalian cell cultures. sixteen species displayed antiviral activity. a compound, which exhibited long wavelength uv-mediated antiviral activity, was isolated from leaves and twigs of elsholtzia ciliata (lamiaceae) using bioassay-guided fractionation and identified as the polycyclic aromatic hydrocarbon, fluoran ...19957753931
myristoylation is required for the intracellular localization and endocytic function of arf6.adp-ribosylation factors (arfs) are modified by a myristate group that is covalently linked to the second glycine residue at the amino terminus. with the recent evidence that arf6 localizes to the cell periphery and plays a regulatory role in endocytic traffic, we have investigated the role of myristoylation on the membrane association, biological activity, and subcellular distribution of arf6 in intact cells. a gly2ala mutation produced a nonmyristoylated protein that failed to associate with m ...19957589240
assessment of fusogenic properties of influenza virus hemagglutinin deacylated by site-directed mutagenesis and hydroxylamine treatment.influenza virus hemagglutinin (ha) subtype h7 expressed from a baculovirus vector in insect cells requires cysteine residues for palmitoylation. mutant ha devoid of fatty acids shows hemagglutinating and hemolytic activities almost identical to those of the acylated wild-type ha (wt). using a membrane mixing assay (r18), neither the kinetics nor the ph dependence of fusion induced by wt or mutant ha was significantly different from virus-induced fusion. ha-induced fusion of insect cells with hum ...19957793071
molecular genetic manipulation of mosquito vectors.despite their central role in disease transmission, relatively little is known of the molecular biology of arthropod vectors. modern molecular approaches will undoubtedly provide considerable information about gene regulation and expression in vectors and consequently a much better understanding of the biology and molecular biology of vectors. such knowledge is essential for developing effective control strategies for vector-borne diseases. in this review, we focus upon techniques and approaches ...19957810990
novel packages of viral and self-antigens are generated during apoptosis.immune context is an essential determinant of the host response to potential autoantigens. the clustering of the autoantigens targeted in systemic lupus erythematosus within surface blebs of apoptotic cells generates high concentrations of autoantigen within discrete subcellular packages. we demonstrate here that when apoptosis is induced by sindbis virus infection, viral antigens and autoantigens cocluster exclusively in small surface blebs of apoptotic cells. the surface of these blebs is rich ...19957699336
reaction in alphavirus mrna capping: formation of a covalent complex of nonstructural protein nsp1 with 7-methyl-gmp.after the start of transcription, the 5' ends of eukaryotic mrna molecules are modified by the addition of a guanylyl residue to form a cap structure, g(5')ppp(5')n. the guanylyltransferase (gtp:mrna guanylyltransferase, ec 2.7.7.50) reaction responsible for cap formation usually proceeds via a covalent enzyme-gmp intermediate. we have studied the alphavirus-specific guanylyltransferase by incubating lysates from semliki forest and sindbis virus-infected cells with [alpha-32p]gtp, using vaccinia ...19957831320
myristoylation cannot functionally replace the isoprenylation of rab5.c-terminal isoprenylation is necessary for the small gtpase rab5 to associate with early endosomes and to exert its regulatory function in endocytosis. in this study, we tested whether rab5 could retain its membrane association and biological function if the isoprenylation were replaced by another type of lipid modification (myristoylation). rab5 mutants were constructed that contained both isoprenylation and myristoylation (gag-rab5), myristoylation only (gag-rab5 delta c4), and neither of the ...19957840662
[genetical and biochemical studies on the metabolisms and biological roles of membrane phospholipids in mammalian cells].the genetical and biochemical approach is used as a powerful tool for the elucidation of the metabolic regulation as well as the biological significance of various constituents in living cells. this technique has been successfully applied to the membrane phospholipids in mammalian cells, actually to phosphatidylserine, sphingomyelin, and cardiolipin, all of which are rather minor constituents in membrane phospholipids. isolation and characterization of the mutant cells defective in biosyntheses ...19947853148
involvement of the molecular chaperone bip in maturation of sindbis virus envelope glycoproteins.sindbis virus codes for two membrane glycoproteins, e1 and pe2, which assemble into heterodimers within the endoplasmic reticulum. we have examined the role of the molecular chaperone bip (grp78) in the maturation of these two proteins. e1, which folds into its mature conformation via at least three intermediates differing in the configurations of their disulfide bonds, was found to interact strongly and transiently with bip after synthesis. atp depletion mediated by carbonyl cyanide m-chlorophe ...19957853497
roles of immunoglobulin valency and the heavy-chain constant domain in antibody-mediated downregulation of sindbis virus replication in persistently infected neurons.clearance of infectious sindbis virus from neurons is mediated by antibody to the e2 glycoprotein. properties of the antibody important for downregulation of sindbis virus replication are unknown. immunoglobulin isotypes and valency determine many biological properties of antibodies. an immunoglobulin g1 (igg1) isotype switch mutant and f(ab')2 and fab fragments of igg3 monoclonal antibody 209 were prepared and tested for clearance of infectious virus from persistently infected rat dorsal root g ...19957853547
a pseudo-revertant of a sindbis virus 6k protein mutant, which corrects for aberrant particle formation, contains two new mutations that map to the ectodomain of the e2 glycoprotein.most site-directed mutations in the gene encoding the small, membrane-associated 6k protein of sindbis virus interfere selectively with virus assembly and budding. particles are released that are aberrant in structure, with a single membrane enclosing multiple nucleocapsids. a revertant for the mutation that inserted a serine for a cysteine at position 39 in the 6k protein was isolated and found to correct for the defective budding so that normal particles were formed. genetic analysis of this r ...19957856077
nucleocapsid and glycoprotein organization in an enveloped virus.alphaviruses are a group of icosahedral, positive-strand rna, enveloped viruses. the membrane bilayer, which surrounds the approximately 400 a diameter nucleocapsid, is penetrated by 80 spikes arranged in a t = 4 lattice. each spike is a trimer of heterodimers consisting of glycoproteins e1 and e2. cryoelectron microscopy and image reconstruction of ross river virus showed that the t = 4 quaternary structure of the nucleocapsid consists of pentamer and hexamer clusters of the capsid protein, but ...19957867069
[the effect of sindbis virus multiplication on gene expression of host cells].sindbis virus (sbv) infection mediated a rapid shutoff of host cellular gene expression (mrna synthesis and protein synthesis); however the synthesis of cellular rrna remained at the same level as the uninfected cells. meanwhile a cellular protein p105 was shown to be enriched in the nuclear matrix. actionmycin d treatment after virus infection resulted in an apparent reduce in the production of viral structural proteins and infectious virions. the results presented here not only demonstrated th ...19947871779
antigenic comparison of ockelbo virus isolates from sweden and russia with sindbis virus isolates from europe, africa, and australia: further evidence for variation among alphaviruses.the plaque-reduction neutralization test (prnt) was used to compare 15 isolates of ockelbo virus from sweden, one isolate of ockelbo virus from russia, the egyptian prototype sindbis virus, and sindbis-like viruses from slovakia, south africa, cameroon, and australia. strains from northern europe (sweden and russia) were indistinguishable by prnt. we observed some antigenic variation between isolates of sindbis virus from europe, africa, and australia. an australian strain (c-377) was shown to b ...19937902675
subgenomic mrna of aura alphavirus is packaged into virions.purified virions of aura virus, a south american alphavirus related to sindbis virus, were found to contain two rna species, one of 12 kb and the other of 4.2 kb. northern (rna) blot analysis, primer extension analysis, and limited sequencing showed that the 12-kb rna was the viral genomic rna, whereas the 4.2-kb rna present in virus preparations was identical to the 26s subgenomic rna present in infected cells. the subgenomic rna is the messenger for translation of the viral structural proteins ...19947902874
lethality of pe2 incorporation into sindbis virus can be suppressed by second-site mutations in e3 and e2.sindbis virions contain two glycoproteins, e1 and e2. e2 is produced initially as a precursor, pe2, from which the amino-terminal 64 amino acids are cleaved by a cellular protease at a late stage in virion maturation. a mutation at e2 position 1 (arg to asn) was placed into sindbis virus ar339 by site-directed mutagenesis of a full-length ar339 cdna clone, ptrsb, to produce ptrsb-n. the mutation created a signal for n-linked glycosylation immediately adjacent to the pe2 cleavage signal. virions ...19947908062
deletion and duplication mutations in the c-terminal nonconserved region of sindbis virus nsp3: effects on phosphorylation and on virus replication in vertebrate and invertebrate cells.little is known concerning the function of the c-terminal nonconserved region of sindbis virus nsp3. in this report, we created a number of in-frame deletions and duplications (from 12 to 159 residues) in this region and examined their effects on sindbis virus replication. sindbis rna transcripts containing these mutations were infectious and gave rise to viable virus after transfection of chicken embryo fibroblasts (cef). in cef, the rate of virus release and accumulation of viral rnas were sim ...19947912020
mutations in an exposed domain of sindbis virus capsid protein result in the production of noninfectious virions and morphological variants.a critical event in the process of sindbis virus assembly is the interaction of the virus nucleocapsid with the target membrane which has been modified by virus envelope glycoproteins. a specific association between the endodomain of the pe2/e2 glycoprotein and an unidentified domain in the capsid protein is responsible for this association. we have examined the nature of this association by the production of a mutant which has two amino acid substitutions in a domain of the capsid protein known ...19947912022
sindbis virus attachment: isolation and characterization of mutants with impaired binding to vertebrate cells.sindbis virus can infect a broad range of insect and vertebrate cell types. the ability to restrict tissue tropism and target virus infection to specific cell types would expand the usefulness of engineered alphaviruses as gene expression vectors. in this study, virus pools derived from libraries of full-length sindbis virus cdna clones containing random insertion mutations in the pe2 or e1 virion glycoprotein gene were screened for mutants defective for binding to vertebrate cells. binding-comp ...19937684466
new phthalocyanines for photodynamic virus inactivation in red blood cell concentrates.cationic phthalocyanines with either aluminum or silicon as the central metal were evaluated for their ability to inactivate viruses in red blood cell concentrates (rbcc) photodynamically. in addition, the virucidal potential of a substituted anionic phthalocyanine, aluminum dibenzodisulfophthalocyanine hydroxide (a1n2sb2poh) was evaluated and compared with that of the much studied anionic aluminum tetrasulfophthalocyanine hydroxide (a1pcs4oh). based on the rate of inactivation of the lipid-enve ...19947938215
antiviral assays on phytochemicals: the influence of reaction parameters.investigations of phytochemicals for antiviral activities are assuming greater importance; but little attention has been given to the influence of various reaction parameters. we found that the activities of several known antiviral phytochemicals were profoundly affected by the presence of serum components, but in different ways. thus, the terthiophene, alpha-terthienyl (alpha-t), was inhibited in a concentration-dependent manner by serum. in the case of a carboxylic acid derivative of alpha-t, ...19947938267
a sindbis virus mrna polynucleotide vector achieves prolonged and high level heterologous gene expression in vivo.the direct intramuscular delivery of naked plasmid dna has been demonstrated to allow expression of encoded heterologous genes in the target myocytes. the method has been employed to elicit immunization based upon delivery of antigen encoding plasmid dna. for application in the context of achieving anti-tumor immunization against antigenic transforming oncoproteins, delivery of plasmid dnas encoding these molecules would create significant potential safety hazards. as an alternative to dna polyn ...19957784202
the amino-terminal residue of sindbis virus glycoprotein e2 influences virus maturation, specific infectivity for bhk cells, and virulence in mice.the e2 glycoprotein of sindbis virus is synthesized as a precursor, pe2, which is cleaved by furin or a furin-like host cell protease at a late stage of maturation. the four-residue pe2 cleavage signal conforms to the basic amino acid-x-basic-basic motif which is present in many other viral and cellular glycoproteins which are processed by the cellular enzyme(s). in this report, we present evidence that the amino acid which immediately follows the signal, the n-terminal residue of e2, can influe ...19947966596
translation of sindbis virus mrna: effects of sequences downstream of the initiating codon.one incentive for developing the alphavirus sindbis virus as a vector for the expression of heterologous proteins is the very high level of viral structural proteins that accumulates in infected cells. although replacement of the structural protein genes by a heterologous gene should lead to an equivalent accumulation of the heterologous protein, the sindbis virus capsid protein is produced at a level 10- to 20-fold higher than that of any foreign protein. chimeric mrnas which contain the first ...19947966601
intracellular interference of tick-borne flavivirus infection by using a single-chain antibody fragment delivered by recombinant sindbis virus.a single-chain antibody fragment that identifies a neutralizing epitope on the envelope protein of louping ill and some other tick-borne flaviviruses was previously expressed in soluble form from bacteria and shown to be functionally active in vitro. to see whether or not the single-chain antibody could bind and inactivate infectious virus in vivo, we have used recombinant sindbis virus as a delivery vehicle for intracellular expression of the antibody fragment. the variable genes and interchain ...19957815482
intracellular immunization of mosquito cells to lacrosse virus using a recombinant sindbis virus vector.a cdna of the small rna genome segment of la crosse (lac) virus was inserted, in an antisense orientation, into a double subgenomic sindbis (dssin) virus expression vector generating pte/3'2j/anti-s (15,000bp). in vitro transcription of the pte/3'2j/anti-s template generated genomic rna that was electrotransfected into bhk-21 cells to produce virus. northern blot analysis of rna isolated from infected aedes albopictus (c6/36) cells showed that the te/3'2j/anti-s virus produced a subgenomic mrna ...19948030366
differential sensitivities of viruses in red cell suspensions to methylene blue photosensitization.previous studies explored the feasibility of using the photosensitizer methylene blue (mb) as a virucidal agent in red cell suspensions. under treatment conditions (5 microm [5 mumol/l] mb, 3.4 x 10(4) j/m2) that resulted in more than 6 log10 inactivation of vesicular stomatitis virus (vsv) or of the enveloped bacteriophage phi 6, red cell membrane alterations were observed. increased red cell ion permeability and the binding of plasma proteins to the red cell surface were the most sensitive ind ...19948023394
age-dependent susceptibility to fatal encephalitis: alphavirus infection of neurons.sindbis virus encephalitis in mice provides a model for studying age-dependent susceptibility to acute viral encephalitis. the ar339 strain of sv causes fatal encephalitis in newborn mice, but weanling mice recover uneventfully. increased virulence for older mice is associated with a single amino acid change from gln to his at position 55 of the e2 glycoprotein. weanling mice with normal immune systems clear infectious virus from neurons through an antibody-mediated mechanism. this does not happ ...19948032263
common replication strategies emerging from the study of diverse groups of positive-strand rna viruses.studies using brome mosaic virus (bmv), sindbis virus and poliovirus have provided evidence that disparate groups of plant and animal positive strand rna viruses have remarkably similar replication strategies. the conservation of several functional domains within virus-encoded nonstructural proteins implies that, although the precise character of these and interacting host components varies for each virus, they employ similar mechanisms for rna replication. for (+) strand replication, similariti ...19948032249
genetic analysis of the nsp3 region of sindbis virus: evidence for roles in minus-strand and subgenomic rna synthesis.sindbis virus nonstructural polyproteins and their cleavage products are believed to be essential components of viral rna replication and transcription complexes. although numerous studies have investigated the effect of mutations in nsp1-, nsp2-, and nsp4-coding regions on sindbis virus-specific rna synthesis, relatively little is known about the function of the region encoding nsp3. nsp3 is a phosphoprotein comprising two regions: an n-terminal portion which is highly conserved among alphaviru ...19948057460
multivalent rna probes and their use in the quantitation of multiple and non-homologous rna species immobilized onto nylon membranes. 19948065945
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