expression of foreign proteins by poliovirus polyprotein fusion: analysis of genetic stability reveals rapid deletions and formation of cardioviruslike open reading frames.using a strategy developed by r. andino, d. silvera, s. d. suggett, p. i. achacoso, c. j. miller, d. baltimore, and m. b. feinberg (science 265:1448-1451, 1994), we constructed recombinant polioviruses by fusing the open reading frame (orf) of the green fluorescent protein gene (gfp) of aequorea victoria or the gag gene (encoding p17-p24) of human immunodeficiency virus type 1 (hiv-1) to the n terminus of the poliovirus polyprotein. all poliovirus expression vectors constructed by us and those o ...19989420196
the papillomavirus minor capsid protein, l2, induces localization of the major capsid protein, l1, and the viral transcription/replication protein, e2, to pml oncogenic domains.we have used immunofluorescent staining and confocal microscopy to examine the subcellular localization of structural and nonstructural bovine papillomavirus (bpv) proteins in cultured cells that produce infectious virions. when expressed separately, l1, the major capsid protein, showed a diffuse nuclear distribution while l2, the minor capsid protein, was found to localize to punctate nuclear regions identified as promonocytic leukemia protein (pml) oncogenic domains (pods). coexpression of l1 ...19989420209
dna immunization with japanese encephalitis virus nonstructural protein ns1 elicits protective immunity in mice.japanese encephalitis virus (jev), a mosquito-borne flavivirus, is a zoonotic pathogen that is prevalent in some southeast asian countries and causes acute encephalitis in humans. to evaluate the potential application of gene immunization to jev infection, we characterized the immune responses from mice intramuscularly injected with plasmid dna encoding jev glycoproteins, including the precursor membrane (prm) plus envelope (e) proteins and the nonstructural protein ns1. when injected with the p ...19989420215
infectious transcripts from cloned genome-length cdna of porcine reproductive and respiratory syndrome virus.the 5'-terminal end of the genomic rna of the lelystad virus isolate (lv) of porcine reproductive and respiratory syndrome virus was determined. to construct full-length cdna clones, the 5'-terminal sequence was ligated to cdna clones covering the complete genome of lv. when rna that was transcribed in vitro from these full-length cdna clones was transfected into bhk-21 cells, infectious lv was produced and secreted. the virus was rescued by passage to porcine alveolar lung macrophages or cl2621 ...19989420236
sindbis virus induces apoptosis through a caspase-dependent, crma-sensitive pathway.sindbis virus infection of cultured cells and of neurons in mouse brains leads to programmed cell death exhibiting the classical characteristics of apoptosis. although the mechanism by which sindbis virus activates the cell suicide program is not known, we demonstrate here that sindbis virus activates caspases, a family of death-inducing proteases, resulting in cleavage of several cellular substrates. to study the role of caspases in virus-induced apoptosis, we determined the effects of specific ...19989420245
a defective interference-like phenomenon of human hepatitis b virus in chronic carriers.defective interfering (di) particles have been found in many rna and dna viruses of bacteria, plants, and animals since their first discovery in influenza virus. however, this fundamental phenomenon has not been demonstrated in human natural infections. using a new approach, here we provide the first experimental evidence for the existence of di-like viruses in human chronic carriers of hepatitis b virus (hbv). functional characterization of naturally occurring core internal deletion (cid) varia ...19989420261
effects of defined mutations in the 5' nontranslated region of rubella virus genomic rna on virus viability and macromolecule synthesis.the 5' end of the genomic rna of rubella virus (rub) contains a 14-nucleotide (nt) single-stranded leader (ss-leader) followed by a stem-and-loop structure [5'(+)sl] (nt 15 to 65), the complement of which at the 3' end of the minus-strand rna [3'(-)sl] has been proposed to function as a promoter for synthesis of genomic plus strands. a second intriguing feature of the 5' end of the rub genomic rna is the presence of a short (17 codons) open reading frame (orf) located between nt 3 and 54; the or ...19989420269
apoptosis in the mouse central nervous system in response to infection with mouse-neurovirulent dengue viruses.apoptosis has been suggested as a mechanism by which dengue (den) virus infection may cause neuronal cell death (p. desprès, m. flamand, p.-e. ceccaldi, and v. deubel, j. virol. 70:4090-4096, 1996). in this study, we investigated whether apoptotic cell death occurred in the central nervous system (cns) of neonatal mice inoculated intracerebrally with den virus. we showed that serial passage of a wild-type human isolate of den virus in mouse brains selected highly neurovirulent variants which rep ...19989420294
identification of the leader-body junctions for the viral subgenomic mrnas and organization of the simian hemorrhagic fever virus genome: evidence for gene duplication during arterivirus evolution.simian hemorrhagic fever virus (shfv) was recently reclassified and assigned to the new virus family arteriviridae. during replication, arteriviruses produce a 3' coterminal, nested set of subgenomic mrnas (sgrnas). these sgrnas arise by discontinuous transcription, and each contains a 5' leader sequence which is joined to the body of the mrna through a conserved junction sequence. only the 5'-most open reading frame (orf) is believed to be transcribed from each sgrna. the shfv genome encodes ni ...19989420301
a single point mutation controls the cholesterol dependence of semliki forest virus entry and exit.membrane fusion and budding are key steps in the life cycle of all enveloped viruses. semliki forest virus (sfv) is an enveloped alphavirus that requires cellular membrane cholesterol for both membrane fusion and efficient exit of progeny virus from infected cells. we selected an sfv mutant, srf-3, that was strikingly independent of cholesterol for growth. this phenotype was conferred by a single amino acid change in the e1 spike protein subunit, proline 226 to serine, that increased the cholest ...19989425157
genus-specific detection of alphaviruses by a semi-nested reverse transcription-polymerase chain reaction.a reverse transcription-polymerase chain reaction (rt-pcr) was developed for the genus-specific detection of alphaviruses. based on the available published sequences, degenerate primers were designed to ensure hybridization to a conserved region within the nonstructural protein 1 gene of all alphavirus species. the expected 434-basepair (bp) cdna fragment was amplified from all 27 alphavirus species by using rna extracted from 200 microl of infected cell culture supernatant. in addition, eight s ...19979430533
modulation of cell death by bcl-xl through caspase interaction.the caspases are cysteine proteases that have been implicated in the execution of programmed cell death in organisms ranging from nematodes to humans. many members of the bcl-2 family, including bcl-xl, are potent inhibitors of programmed cell death and inhibit activation of caspases in cells. here, we report a direct interaction between caspases and bcl-xl. the loop domain of bcl-xl is cleaved by caspases in vitro and in cells induced to undergo apoptotic death after sindbis virus infection or ...19989435230
comparison of various expression plasmids for the induction of immune response by dna immunization.intramuscular injection of plasmid dna is an efficient method to introduce a foreign gene into a live animal. we investigated several factors affecting the gene transfer efficiency and the following immune response by intramuscular injection of plasmid dna. when the strength of several highly efficient viral promoters was compared in muscle by using the chloramphenicol acetyltransferase (cat) gene as an indicator, cytomegalovirus (cmv) immediate early promoter was found to be stronger than any o ...19979339893
molecular genetic study of the interaction of sindbis virus e2 with ross river virus e1 for virus budding.glycoprotein pe2 of sindbis virus will form a heterodimer with glycoprotein e1 of ross river virus that is cleaved to an e2/e1 heterodimer and transported to the cell plasma membrane, but this chimeric heterodimer fails to interact with sindbis virus nucleocapsids, and very little budding to produce mature virus occurs upon infection with chimeric viruses. we have isolated in both sindbis virus e2 and in ross river virus e1 a series of suppressing mutations that adapt these two proteins to one a ...19989445043
structural localization of the e3 glycoprotein in attenuated sindbis virus mutants.we have determined the three-dimensional structures of the wild-type sindbis virus and two of its mutants that retain the e3 sequence within pe2. using difference imaging between these mutants and the wild-type virus, we have assigned a location for the 64-amino-acid sequence corresponding to e3 in the mutant spike complex. in the wild-type virus, the spike is composed of an e1-e2 heterotrimer. the e3 protein was found to protrude midway between the center of the spike complex and the tips. base ...19989445057
the evolution of virus-induced apoptosis.viruses from several different families are able to exploit their host's cell death programmes so as to maximize viral fitness. consideration of the evolution of such strategies has lead to the suggestion that the virus should inhibit apoptosis, in order to prolong the life of the cell and thereby maximize the number of progeny virions. the host, on the other hand, should stimulate apoptosis thereby inhibiting viral growth and blocking viral spread. for example, the function of the latent membra ...19979447732
the alphavirus 3'-nontranslated region: size heterogeneity and arrangement of repeated sequence elements.the 3'-nontranslated region (ntr) of representative strains of all known alphavirus species was amplified by reverse transcription-polymerase chain reaction. for 23 of them, the 3'-ntr sequence was determined. together with previously published data, this allowed an analysis of the 3'-ntr of the viruses in the genus alphavirus. the length of the 3'-ntrs varied from 77 nt for pixuna virus to 609 nt for bebaru virus. the 19-nt conserved sequence element directly adjacent to the poly(a) tract was f ...19989448694
identification and genetic analysis of panama-genotype venezuelan equine encephalitis virus subtype id in peru.venezuelan equine encephalitis (vee) virus was isolated in 1993, 1994, and 1995 from human cases of acute, undifferentiated, febrile illness in the peruvian amazon basin. two virus isolates were recovered in 1994 from peruvian soldiers at a jungle outpost near pantoja in northern peru, and 10 isolates were obtained from military personnel and civilians in 1993-1995 in iquitos, an urban center in northeastern peru. the genetic relationship of these isolates to other vee virus strains was determin ...19989452290
comparison of sindbis virus-induced pathology in mosquito and vertebrate cell cultures.we have compared sindbis virus-induced cytopathology in vertebrate and mosquito (aedes albopictus) cell cultures. it has been shown that vertebrate cells undergo apoptosis when infected by sindbis virus and this was confirmed here using hamster cells (bhk). the occurrence of cell death in sindbis virus-infected a. albopictus cells is a cell clone-specific phenomenon and, unlike in bhk cell cultures, mosquito cell death does not correlate with a large induction of apoptosis, as determined by assa ...19989454692
cyclin dependent kinase inhibitors and dominant negative cyclin dependent kinase 4 and 6 promote survival of ngf-deprived sympathetic neurons.neuronal apoptosis plays a critical role in both normal development and disease. however, the precise molecular events controlling neuronal apoptosis are not well understood. previously, we hypothesized that cell cycle regulatory molecules function in controlling the apoptotic pathways of trophic factor-deprived neurons. to test this hypothesis, we used the rna alphavirus sindbis to express three known cyclin dependent kinase inhibitors (ckis), p16(ink4), p21(waf/cip), and p27(kip1), and dominan ...19979364045
regulation of alphavirus 26s mrna transcription by replicase component nsp2.semliki forest virus (sfv) mutant ts4 has a reversible temperature-sensitive defect in the synthesis the subgenomic 26s mrna. the viral nonstructural protein nsp2 was identified as a regulator of 26s synthesis by transferring nsp2 coding sequences from ts4 into the infectious sfv cdna clone (sfoto) to create sfots4. sequencing identified the causal mutation as c4038u, predicting the amino acid change m781t in nsp2. a revertant was isolated in which a back mutation of u to c restored the wild-typ ...19989472615
eps15 and eps15r are essential components of the endocytic pathway.eps15 and eps1sr are substrates of the epidermal growth factor (egf) receptor kinase that are characterized by the presence of a protein:protein interaction domain, the eh domain, and by their ability to bind to the clathrin adaptor protein complex adaptor protein 2. indirect evidence suggests that eps15 and eps15r are involved in endocytosis. here we show that microinjection of antibodies against eps15 and eps15r inhibits internalization of egf and transferrin. in addition, fragments of eps15 ( ...19979407958
alphaviruses induce apoptosis in bcl-2-overexpressing cells: evidence for a caspase-mediated, proteolytic inactivation of bcl-2.bcl-2 oncogene expression plays a role in the establishment of persistent viral infection by blocking virus-induced apoptosis. this might be achieved by preventing virus-induced activation of caspase-3, an il-1beta-converting enzyme (ice)-like cysteine protease that has been implicated in the death effector phase of apoptosis. contrary to this model, we show that three cell types highly overexpressing functional bcl-2 displayed caspase-3 activation and underwent apoptosis in response to infectio ...19989482724
signal peptidase cleavage at the flavivirus c-prm junction: dependence on the viral ns2b-3 protease for efficient processing requires determinants in c, the signal peptide, and prm.signal peptidase cleavage at the c-prm junction in the flavivirus structural polyprotein is inefficient in the absence of the cytoplasmic viral protease, which catalyzes cleavage at the cooh terminus of the c protein. the signal peptidase cleavage occurs efficiently in circumstances where the c protein is deleted or if the viral protease complex is present. in this study, we used cdna of murray valley encephalitis virus (mve) to examine features of the structural polyprotein which allow this reg ...19989499070
chimeric measles viruses with a foreign envelope.measles virus (mv) and vesicular stomatitis virus (vsv) are both members of the mononegavirales but are only distantly related. we generated two genetically stable chimeric viruses. in mgv, the reading frames of the mv envelope glycoproteins h and f were substituted by a single reading frame encoding the vsv g glycoprotein; mg/fv is similar but encodes a g/f hybrid in which the vsv g cytoplasmic tail was replaced by that of mv f. in contrast to mg/fv, mgv virions do not contain the mv matrix (m) ...19989499071
a retention signal necessary and sufficient for endoplasmic reticulum localization maps to the transmembrane domain of hepatitis c virus glycoprotein e2.the hepatitis c virus (hcv) genome encodes two envelope glycoproteins (e1 and e2). these glycoproteins interact to formin a noncovalent heterodimeric complex which is retained in the endoplasmic reticulum (er). to identify whether e1 and/or e2 contains an er-targeting signal potentially involved in er retention of the e1-e2 complex, these proteins were expressed alone and their intracellular localization was studied. due to misfolding of e1 in the absence of e2, no conclusion on the localization ...19989499075
dna immunization against herpes simplex virus: enhanced efficacy using a sindbis virus-based vector.previously we reported the development of a plasmid dna expression vector system derived from sindbis virus (t. w. dubensky, jr., et al., j. virol. 70:508-519, 1996). in vitro, such vectors exhibit high-level heterologous gene expression via self-amplifying cytoplasmic rna replication. in the present study, we demonstrated the in vivo efficacy of the sindbis virus-based psin vectors as dna vaccines. a single intramuscular immunization of balb/c mice with psin vectors expressing the glycoprotein ...19989444987
characterization of an autonomous subgenomic pestivirus rna an initial approach to define the requirements for the replication of bovine viral diarrhea virus (bvdv), a member of the flaviviridae family with a positive-strand rna genome, full-length genomic and subgenomic rnas were originated by in vitro transcription of diverse bvdv cdna constructs and transfected into eucaryotic host cells. rna replication was measured either directly by an rnase protection method or by monitoring the synthesis of viral protein. when full-length bvdv crna was initial ...19989499097
characterization of defective viral rna produced during persistent infection of vero cells with murray valley encephalitis virus.defective interfering viral particles are readily produced in cell culture after a high multiplicity of infection with many animal rna viruses. due to defects that they carry in their genomes, their life cycle needs to be complemented by the helper functions provided by a parental virus which makes them both dependent on and competitive with the parental virus. in many instances, this may cause the abrogation of a lytic cycle of the parental virus, leading to a persistent infection. in this pape ...19989499109
effect of nitric oxide on poliovirus infection of two human cell lines.the role of nitric oxide after poliovirus infection of the human hela (carcinoma) and u937 (promonocytic) cell lines has been analyzed. both types of cells produced detectable levels of nitric oxide after poliovirus infection. however, this production was not sufficient to limit viral productivity. on the other hand, pretreatment with the nitric oxide donor glycerine trinitrate lengthened the course of poliovirus infection.19989499120
n-terminal protease of pestiviruses: identification of putative catalytic residues by site-directed mutagenesis.pestiviruses are the only members of the flaviviridae that encode a nonstructural protease at the n terminus of their polyproteins. this n-terminal protease (npro) cleaves itself off of the nascent polyprotein autocatalytically and thereby generates the n terminus of the adjacent viral capsid protein c. in previous reports, sequence similarities between npro and the catalytic residues of papain-like cysteine proteases were put forward. to test this hypothesis, substitutions of cysteine and histi ...19989499122
suppressors of cleavage-site mutations in the p62 envelope protein of semliki forest virus reveal dynamics in spike structure and function.the e2 spike glycoprotein of semliki forest virus is produced as a p62 precursor protein, which is cleaved by host proteases to its mature form, e2. cleavage is not necessary for particle formation or release but is necessary for infectivity. previous results had shown that phenotypic revertants of cleavage-deficient p62 mutants are generated, and here we show that these may contain second-site suppressor mutations in the vicinity of the cleavage site. these hot-spot sites were mutated to abolis ...19989525602
specific interactions between retrovirus env and gag proteins in rat this work we have studied the intracellular localization properties of the gag and env proteins of moloney murine leukemia virus (mlv) and human immunodeficiency virus (hiv) in dorsal root ganglion (drg) neurons of rat. these neurons form thick bundles of axons, which facilitates protein localization studies by immunofluorescence analyses. when such neuron cultures were infected with recombinant semliki forest virus particles carrying the gag genes of either retrovirus, the expressed gag prot ...19989525603
role of actin microfilaments in black creek canal virus morphogenesis.we have investigated the involvement of cytoskeletal proteins in the morphogenesis of black creek canal virus (bccv), a new world hantavirus. immunofluorescent staining of bccv-infected cells revealed a filamentous pattern of virus antigen, the appearance of which was sensitive to treatment with cytochalasin d, an actin microfilament-depolymerizing drug. double immunofluorescence staining of bccv-infected vero cells with anti-bccv nucleocapsid (n) monoclonal antibody and phalloidin revealed a co ...19989525606
development of a chimeric sindbis virus with enhanced per os infection of aedes aegypti.the te/3'2j double subgenomic sindbis (dssin) viruses have been used to stably express genes in aedes aegypti nerve and salivary gland tissues. however, because these viruses inefficiently infect ae. aegypti when administered by the per os route, te/3'2j viruses must be intrathoracically inoculated into the mosquitoes to infect these tissues. a malaysian sindbis (sin) virus isolate (mre16) does efficiently infect ae. aegypti midgut tissues after ingestion, and approximately 95% of these mosquito ...19989527919
functional role of hepatitis c virus chimeric glycoproteins in the infectivity of pseudotyped virus.the putative envelope glycoproteins of hepatitis c virus (hcv) likely play an important role in the initiation of viral infection. available information suggests that the genomic regions encoding the putative envelope glycoproteins, when expressed as recombinant proteins in mammalian cells, largely accumulate in the endoplasmic reticulum. in this study, genomic regions which include the putative ectodomain of the e1 (amino acids 174 to 359) and e2 (amino acids 371 to 742) glycoproteins were appe ...19989557633
coding sequences enhance internal initiation of translation by hepatitis a virus rna in vitro.hepatitis a virus (hav), unlike other picornaviruses, has a slow-growth phenotype in permissive cell lines and in general does not induce host cell cytopathology. although there are no published reports of productive infection of hela cells by hav, hav rna appears to be readily translated in hela cells when transcribed by t7 rna polymerase provided by a recombinant vaccinia virus. the 5' noncoding region of hav was fused to poliovirus (pv) coding sequences to determine the effect on translation ...19989557637
foot-and-mouth disease virus virulent for cattle utilizes the integrin alpha(v)beta3 as its receptor.adsorption and plaque formation of foot-and-mouth disease virus (fmdv) serotype a12 are inhibited by antibodies to the integrin alpha(v)beta3 (a. berinstein et al., j. virol. 69:2664-2666, 1995). a human cell line, k562, which does not normally express alpha(v)beta3 cannot replicate this serotype unless cells are transfected with cdnas encoding this integrin (k562-alpha(v)beta3 cells). in contrast, we found that a tissue culture-propagated fmdv, type o1bfs, was able to replicate in nontransfecte ...19989557639
mutations in rotavirus nonstructural glycoprotein nsp4 are associated with altered virus virulence.rotaviruses are major pathogens causing life-threatening dehydrating gastroenteritis in children and animals. one of the nonstructural proteins, nsp4 (encoded by gene 10), is a transmembrane, endoplasmic reticulum-specific glycoprotein. recently, our laboratory has shown that nsp4 causes diarrhea in 6- to 10-day-old mice by functioning as an enterotoxin. to confirm the role of nsp4 in rotavirus pathogenesis, we sequenced gene 10 from two pairs of virulent and attenuated porcine rotaviruses, the ...19989557647
collaboration of antibody and inflammation in clearance of rabies virus from the central nervous investigate the involvement of various cellular and humoral aspects of immunity in the clearance of rabies virus from the central nervous system, (cns), we studied the development of clinical signs and virus clearance from the cns in knockout mice lacking either b and t cells, cd8+ cytotoxic t cells, b cells, alpha/beta interferon (ifn-alpha/beta) receptors, ifn-gamma receptors, or complement components c3 and c4. following intranasal infection with the attenuated rabies virus cvs-f3, normal ...19989557653
the transmembrane domains of sindbis virus envelope glycoproteins induce cell death.sindbis virus, the prototype alphavirus, kills cells by inducing apoptosis. to investigate potential mechanisms by which sindbis virus induces apoptosis, we examined whether specific viral gene products were able to induce cell death. genes encoding the three structural proteins--capsid, the precursor e1 (6k plus e1), and the precursor e2 (p62 or e3 plus e2)--were cotransfected with a beta-galactosidase reporter plasmid in transient-transfection assays in rat prostate adenocarcinoma at3 cells. c ...19989557679
two types of virus-related particles are found during transmissible gastroenteritis virus morphogenesis.the intracellular assembly of the transmissible gastroenteritis coronavirus (tgev) was studied in infected swine testis (st) cells at different postinfection times by using ultrathin sections of conventionally embedded infected cells, freeze-substitution, and methods for detecting viral proteins and rna at the electron microscopy level. this ultrastructural analysis was focused on the identification of the different viral components that assemble in infected cells, in particular the spherical, p ...19989557690
fus-1, a ph shift mutant of semliki forest virus, acts by altering spike subunit interactions via a mutation in the e2 subunit.semliki forest virus (sfv), an enveloped alphavirus, is a well-characterized paradigm for viruses that infect cells via endocytic uptake and low-ph-triggered fusion. the sfv spike protein is composed of a dimer of e1 and e2 transmembrane subunits, which dissociate upon exposure to low ph, liberating e2 and the fusogenic e1 subunit to undergo independent conformational changes. sfv fusion and infection are blocked by agents such as ammonium chloride, which act by raising the ph in the endosome an ...19989557718
deletion analysis of a defective interfering semliki forest virus rna genome defines a region in the nsp2 sequence that is required for efficient packaging of the genome into virus particles.the 1,244-nucleotide genome of semliki forest virus (sfv) defective interfering (di) rna 19 (di-19) is coterminal with the infectious genome and contains two major deletions. one deletion removes the end of the nsp1 gene and the beginning of the nsp2 gene, and the other removes the end of the nsp2 gene, the nsp3 and nsp4 genes, and all of the structural protein genes (m. thomson and n. j. dimmock, virology 199:354-365, 1994). like all di sfv rnas, di-19 contains three regions that are conserved. ...19989557722
a 68-nucleotide sequence within the 3' noncoding region of simian hemorrhagic fever virus negative-strand rna binds to four ma104 cell proteins.the 3' noncoding region (ncr) of the negative-strand rna [3'(-)ncr rna] of the arterivirus simian hemorrhagic fever virus (shfv) is 209 nucleotides (nt) in length. since this 3' region, designated 3'(-)209, is the site of initiation of full-length positive-strand rna and is the template for the synthesis of the 5' leader sequence, which is found on both full-length and subgenomic mrnas, it is likely to contain cis-acting signals for rna synthesis and to interact with cellular and viral proteins ...19989557724
functions of conserved motifs in the rna-dependent rna polymerase of a yeast double-stranded rna least eight conserved motifs are visible in the totivirus rna-dependent rna polymerase (rdrp). we have systematically altered each of these in the saccharomyces cerevisiae double-stranded rna virus scvl1 by substituting the conserved motifs from a giardiavirus. the results help define the conserved regions of the rdrp involved in polymerase function and those essential for other reasons.19989557735
the rubella virus nonstructural protease requires divalent cations for activity and functions in trans.the rubella virus (rub) nonstructural (ns) protease is a papain-like cysteine protease (pcp) located in the ns-protein open reading frame (nsp-orf) that cleaves the nsp-orf translation product at a single site to produce two products, p150 (the n-terminal product) and p90 (the c-terminal product). the rub ns protease was found not to function following translation in vitro in a standard rabbit reticulocyte lysate system, although all of the other viral pcps do so. however, in the presence of div ...19989557742
[regulation of expression of human fibroblast interferon gene by human leukocyte interferon and cytopathogenic alpha-virus]. 19989559136
expression of a biologically active antiviral antibody using a sindbis virus vector system.monoclonal antibodies to the sindbis virus e2 envelope glycoprotein protect mice against lethal encephalitis and mediate viral clearance from neurons. to facilitate structure-function analyses of anti-e2 mabs, we developed an expression system that can be used for the construction of genetically engineered anti-e2 mabs. we constructed recombinant sindbis/immunoglobulin gene chimeric viruses that express heavy and light chains of an anti-e2 monoclonal antibody, r6. we used a pcr-based strategy to ...19979464526
antibody-mediated neutralization of human rhinovirus 14 explored by means of cryoelectron microscopy and x-ray crystallography of virus-fab complexes.the structures of three different human rhinovirus 14 (hrv14)-fab complexes have been explored with x-ray crystallography and cryoelectron microscopy procedures. all three antibodies bind to the nim-ia site of hrv14, which is the beta-b-beta-c loop of the viral capsid protein vp1. two antibodies, fab17-ia (fab17) and fab12-ia (fab12), bind bivalently to the virion surface and strongly neutralize viral infectivity whereas fab1-ia (fab1) strongly aggregates and weakly neutralizes virions. the stru ...19989573224
evaluation of the galalpha1-3gal epitope as a host modification factor eliciting natural humoral immunity to enveloped viruses.human sera contain high levels of natural antibody (ab) to galalpha1-3gal, a terminal glycosidic structure expressed on the surface of cells of mammals other than old world primates. incorporation of this determinant onto retroviral membranes by passage of viruses in cells encoding alpha-1-3-galactosyltransferase (gt) renders retroviruses sensitive to lysis by natural ab and complement in normal human serum (nhs). plasma membrane-budding viruses representing four additional virus groups were exa ...19989573228
infectious bovine viral diarrhea virus (strain nadl) rna from stable cdna clones: a cellular insert determines ns3 production and viral cytopathogenicity.bovine viral diarrhea virus (bvdv), strain nadl, was originally isolated from an animal with fatal mucosal disease. this isolate is cytopathic in cell culture and produces two forms of ns3-containing proteins: uncleaved ns2-3 and mature ns3. for bvdv nadl, the production of ns3, a characteristic of cytopathic bvdv strains, is believed to be a consequence of an in-frame insertion of a 270-nucleotide cellular mrna sequence (called cins) in the ns2 coding region. in this study, we constructed a sta ...19989573238
transmissible gastroenteritis coronavirus induces programmed cell death in infected cells through a caspase-dependent this report, we show that apoptosis (or programmed cell death) is induced in different cell lines infected with a coronavirus, the porcine transmissible gastroenteritis virus (tgev). kinetic analysis of internucleosomal dna cleavage by agarose gel electrophoresis and flow cytometry or cytometric monitoring of the mitochondrial transmembrane potential showed that, for st cells infected with tgev, the first overt signs of apoptosis appeared from 10 to 12 h postinfection on. they preceded morpho ...19989573259
induction of protective immunity against japanese encephalitis in mice by immunization with a plasmid encoding japanese encephalitis virus premembrane and envelope genes.a dna vaccine plasmid containing the japanese encephalitis (je) virus premembrane (prm) and envelope (e) genes (designated pcdna3jeme) was evaluated for immunogenicity and protective efficacy in mice. two immunizations of 4-week-old female icr mice with pcdna3jeme by intramuscular or intradermal injections at a dose of 10 or 100 microg per mouse elicited neutralizing (neut) antibodies at titers of 1:10 to 1:20 (90% plaque reduction), and all immunized mice survived a challenge with 10,000 50% le ...19989573260
alphavirus-specific cytotoxic t lymphocytes recognize a cross-reactive epitope from the capsid protein and can eliminate virus from persistently infected macrophages.persistent alphavirus infections in synovial and neural tissues are believed to be associated with chronic arthritis and encephalitis, respectively, and represent likely targets for cd8+ alphabeta cytotoxic t lymphocytes (ctl). here we show that the capsid protein is a dominant target for alphavirus-specific ctl in balb/c mice and that capsid-specific ctl from these mice recognize an h-2kd restricted epitope, qysggrfti. this epitope lies in the highly conserved region of the capsid protein, and ...19989573286
purification of a multipotent antideath activity from bovine liver and its identification as arginase: nitric oxide-independent inhibition of neuronal apoptosis.catalase is an antioxidant enzyme that has been shown to inhibit apoptotic or necrotic neuronal death induced by hydrogen peroxide. we report the purification of a contaminating antiapoptotic activity from a commercial bovine liver catalase preparation by following its ability to inhibit apoptosis when applied extracellularly in multiple death paradigms. the antiapoptotic activity was identified by protein microsequencing as arginase, a urea cycle and nitric oxide synthase-regulating enzyme, and ...19989592089
engineered resistance in aedes aegypti to a west african and a south american strain of yellow fever virus.double subgenomic sindbis (dssin) viruses were engineered to transduce mosquito cells with antisense rna derived either from the premembrane (prm) or polymerase (ns5) coding regions of the 17d vaccine strain of yellow fever virus (yfv). aedes albopictus c6/36 cells were infected at high multiplicities of infection (moi) with each dssin virus. forty-eight hours later, the transduced cells were challenged with an moi of 0.1 of the asibi strain of yfv. at 72-hr postchallenge, the cells were assayed ...19989598458
the serine proteinase inhibitor (serpin) plasminogen activation inhibitor type 2 protects against viral cytopathic effects by constitutive interferon alpha/beta priming.the serine proteinase inhibitor (serpin) plasminogen activator inhibitor type 2 (pai-2) is well characterized as an inhibitor of extracellular urokinase-type plasminogen activator. here we show that intracellular, but not extracellular, pai-2 protected cells from the rapid cytopathic effects of alphavirus infection. this protection did not appear to be related to an effect on apoptosis but was associated with a pai-2-mediated induction of constitutive low-level interferon (ifn)-alpha/beta produc ...19989607921
genes required for replication of the 15.5-kilobase rna genome of a plant closterovirus.a full-length cdna clone of beet yellows closterovirus (byv) was engineered and used to map functions involved in the replication of the viral rna genome and subgenomic rna formation. among 10 open reading frames (orfs) present in byv, orfs 1a and 1b suffice for rna replication and transcription. the proteins encoded in these orfs harbor putative methyltransferase, rna helicase, and rna polymerase domains common to sindbis virus-like viruses and a large interdomain region that is unique to clost ...19989621048
antiapoptotic activity of the herpesvirus saimiri-encoded bcl-2 homolog: stabilization of mitochondria and inhibition of caspase-3-like activity.viruses have evolved different strategies to interfere with host cell apoptosis. herpesvirus saimiri (hvs) and other lymphotropic herpesviruses code for proteins that are homologous to the cellular antiapoptotic bcl-2. in this study hvs-bcl-2 was stably expressed in the human leukemia cell line jurkat and in the murine t-cell hybridoma do to assess its antiapoptotic spectrum and to gain further insight into its mode of action. hvs- bcl-2 prevented apoptosis that occurs as a result of a disturban ...19989621051
encapsidation of the flavivirus kunjin replicon rna by using a complementation system providing kunjin virus structural proteins in trans.kunjin virus (kun) replicon rna was encapsidated by a procedure involving two consecutive electroporations of bhk-21 cells, first with kun replicon rna c20dxrep (with prme and most of c deleted) and about 24 h later with a recombinant semliki forest virus (sfv) replicon rna(s) expressing kun structural proteins. the presence of kun replicon rna in encapsidated particles was demonstrated by its amplification and expression in newly infected bhk-21 cells, detected by northern blotting with a kun-s ...19989621059
his...asp catalytic dyad of ribonuclease a: structure and function of the wild-type, d121n, and d121a enzymes.the side chains of histidine and aspartate residues form a hydrogen bond in the active sites of many enzymes. in serine proteases, the his...asp hydrogen bond of the catalytic triad is known to contribute greatly to catalysis, perhaps via the formation of a low-barrier hydrogen bond. in bovine pancreatic ribonuclease a (rnase a), the his...asp dyad is composed of his119 and asp121. previously, site-directed mutagenesis was used to show that his119 has a fundamental role, to act as an acid during ...19989636030
a novel sialic acid binding site on factor h mediates serum resistance of sialylated neisseria gonorrhoeae.factor h (fh), a key alternative complement pathway regulator, is a cofactor for factor i-mediated cleavage of c3b. fh consists of 20 short consensus repeat (scr) domains. sialic acid binding domains have previously been localized to fh scrs 6-10 and 13. to examine fh binding on a sialylated microbial surface, we grew neisseria gonorrhoeae in the presence of 5'-cytidinemonophospho-n-acetylneuraminic acid, which sialylates lipooligosaccharide and converts to serum resistance gonococci previously ...19989480984
template-dependent initiation of sindbis virus rna replication in vitro.recent insights into the early events in sindbis virus rna replication suggest a requirement for either the p123 or p23 polyprotein, as well as mature nsp4, the rna-dependent rna polymerase, for initiation of minus-strand rna synthesis. based on this observation, we have succeeded in reconstituting an in vitro system for template-dependent initiation of sin rna replication. extracts were isolated from cells infected with vaccinia virus recombinants expressing various sin proteins and assayed by ...19989658098
steady-state plasma membrane expression of human cytomegalovirus gb is determined by the phosphorylation state of ser900.human cytomegalovirus (hcmv) infection of an astrocytoma cell line (u373) or human fibroblast (hf) cells results in a differential cell distribution of the major envelope glycoprotein gb (ul55). this 906-amino-acid type i glycoprotein contains an extracellular domain with a signal sequence, a transmembrane domain, and a 135-amino-acid cytoplasmic tail with a consensus casein kinase ii (ckii) site located at ser900. since phosphorylation of proteins in the secretory pathway is an important determ ...19989658112
coronavirus particle assembly: primary structure requirements of the membrane protein.coronavirus-like particles morphologically similar to normal virions are assembled when genes encoding the viral membrane proteins m and e are coexpressed in eukaryotic cells. using this envelope assembly assay, we have studied the primary sequence requirements for particle formation of the mouse hepatitis virus (mhv) m protein, the major protein of the coronavirion membrane. our results show that each of the different domains of the protein is important. mutations (deletions, insertions, point ...19989658133
recombinant sindbis viruses expressing a cytotoxic t-lymphocyte epitope of a malaria parasite or of influenza virus elicit protection against the corresponding pathogen in mice.subcutaneous administration in mice of recombinant sindbis viruses expressing a class i major histocompatibility complex-restricted 9-mer epitope of the plasmodium yoelii circumsporozoite protein or the nucleoprotein of influenza virus induces a large epitope-specific cd8(+) t-cell response. this immunization also elicits a high degree of protection against infection with malaria or influenza a virus.19989658144
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 gener ...19989660762
inducible nitric oxide synthase and nitrotyrosine are found in monocytes/macrophages and/or astrocytes in acute, but not in chronic, multiple sclerosis.we have examined the localization of inducible nitric oxide synthase (inos) and nitrotyrosine (the product of nitration of tyrosine by peroxynitrite, a highly reactive derivative of nitric oxide [no]) in demyelinating lesions from (i) two young adult patients with acute multiple sclerosis (ms), (ii) a child with ms (consistent with diffuse sclerosis), and (iii) five adult patients with chronic ms. previous reports have suggested a possible correlation between inos, peroxynitrite, related nitroge ...19989665945
infectious bursal disease virus polyprotein processing does not involve cellular proteases.the larger genome segment, segment a, of infectious bursal disease virus (ibdv) encodes vp2, vp3 and vp4 as a precursor polyprotein. the viral protease, vp4, is responsible for self-processing of the polyprotein, however, there are additional secondary precursor products such as vpx whose further processing has not been defined. expression of ibdv cdnas in vitro with rabbit reticulocyte lysates in a coupled transcription-translation system and in the sindbis virus expression system (with bhk-21 ...19979672603
cell-specific transfection of choriocarcinoma cells by using sindbis virus hcg expressing chimeric vector.the development of sindbis virus vectors that can target specific cell types would provide an important gene therapy strategy. we explored the possibility of designing a sindbis virus vector that can target human choriocarcinoma cells via ligand-receptor interaction. the sindbis virus envelope gene was modified by insertion of the alpha- and beta-hcg genes. the chimeric helper rna was then transfected into bhk cells along with a virus-based expression vector, allowing the production of virus par ...19989675133
a novel method of cell-specific mrna this study, we developed a cell-specific mrna transfection system using streptavidin-protein a (st-pa) fusion protein and monoclonal antibodies (mabs). we previously reported that st-pa fusion protein and mab complexes can transfer certain biotinylated proteins into specific cell types. at this time, we combined an in vitro transcribed biotinylated and self-replicating sindbis virus genomic rna with st-pa fusion protein and mabs. in the presence of cationic liposomes, to prevent rna degradati ...19989682217
requirement for an aromatic amino acid or histidine at the n terminus of sindbis virus rna polymerase.the n terminal amino acid of nonstructural protein nsp4, the viral rna polymerase, is a tyrosine in all sequenced alphaviruses; this is a destabilizing amino acid for the n-end rule pathway and results in rapid degradation of nsp4 produced in infected cells or in reticulocyte lysates. we have constructed 11 mutants of sindbis virus bearing phe, ala, thr, cys, leu, met, asn, gln, glu, arg, or pro at the n terminus of nsp4. translation of rnas in reticulocyte lysates showed that cleavage at the ns ...19989499091
mutations in the sindbis virus capsid gene can partially suppress mutations in the cytoplasmic domain of the virus e2 glycoprotein spike.assembly and budding of alphaviruses are postulated to occur by protein-protein interactions between sites on the cytoplasmic domain of the transmembranal envelope e2 glycoprotein and on the surface of the nucleocapsid protein subunits. genetic data to support this model have been obtained by isolating revertants of two slow-growth mutants of sindbis virus and analyzing the sequences of the genes encoding their structural proteins. the slow-growth phenotypes of the mutants were previously shown ...19989568036
interactions between the structural domains of the rna replication proteins of plant-infecting rna viruses.brome mosaic virus (bmv), a positive-strand rna virus, encodes two replication proteins: the 2a protein, which contains polymerase-like sequences, and the 1a protein, with n-terminal putative capping and c-terminal helicase-like sequences. these two proteins are part of a multisubunit complex which is necessary for viral rna replication. we have previously shown that the yeast two-hybrid assay consistently duplicated results obtained from in vivo rna replication assays and biochemical assays of ...19989696810
rescue of defective poliovirus rna replication by 3ab-containing precursor polyproteins.this study demonstrates the in vitro complementation of an rna replication-defective lesion in poliovirus rna by providing a replicase/polymerase precursor polypeptide [p3(wt) (wild type)] in trans. the replication-defective mutation was a phenylalanine-to-histidine change (f69h) in the hydrophobic domain of the membrane-associated viral protein 3ab. rnas encoding wild-type forms of protein 3ab or the p3 precursor polypeptide were cotranslated with full-length poliovirus rnas containing the f69h ...19989696813
trans-complementation of flavivirus rna polymerase gene ns5 by using kunjin virus replicon-expressing bhk cells.a bhk cell line persistently expressing a kunjin (kun) virus replicon rna (repbhk, similar to our recently described me/76neo bhk cell line [a. a. khromykh and e. g. westaway, j. virol. 71:1497-1505, 1997]) was used for rescue and propagation of kun viruses defective in the rna polymerase gene (ns5). a new infectious full-length kun virus cdna clone, flsdx, prepared from our previously described cdna clone pakun (a. a. khromykh and e. g. westaway, j. virol. 68:4580-4588, 1994) and possessing app ...19989696822
binding of sindbis virus to cell surface heparan sulfate.alphaviruses are arthropod-borne viruses with wide species ranges and diverse tissue tropisms. the cell surface receptors which allow infection of so many different species and cell types are still incompletely characterized. we show here that the widely expressed glycosaminoglycan heparan sulfate can participate in the binding of sindbis virus to cells. enzymatic removal of heparan sulfate or the use of heparan sulfate-deficient cells led to a large reduction in virus binding. sindbis virus bou ...19989696831
adaptation of sindbis virus to bhk cells selects for use of heparan sulfate as an attachment receptor.attachment of sindbis virus to the cell surface glycosaminoglycan heparan sulfate (hs) and the selection of this phenotype by cell culture adaptation were investigated. virus (tr339) was derived from a cdna clone representing the consensus sequence of strain ar339 (k. l. mcknight, d. a. simpson, s. c. lin, t. a. knott, j. m. polo, d. f. pence, d. b. johannsen, h. w. heidner, n. l. davis, and r. e. johnston, j. virol. 70:1981-1989, 1996) and from mutant clones containing either one or two dominan ...19989696832
identification of specific nucleotide sequences within the conserved 3'-sl in the dengue type 2 virus genome required for replication.the flavivirus genome is a positive-stranded approximately 11-kb rna including 5' and 3' noncoding regions (ncr) of approximately 100 and 400 to 600 nucleotides (nt), respectively. the 3' ncr contains adjacent, thermodynamically stable, conserved short and long stem-and-loop structures (the 3'-sl), formed by the 3'-terminal approximately 100 nt. the nucleotide sequences within the 3'-sl are not well conserved among species. we examined the requirement for the 3'-sl in the context of dengue virus ...19989696848
classical swine fever virus leader proteinase npro is not required for viral replication in cell culture.the sequence encoding the viral leader proteinase npro was replaced by the murine ubiquitin gene in a full-length cdna clone of the classical swine fever virus (csfv) strain alfort/187. the recombinant virus va187-ubi showed growth characteristics similar to those of the parent va187-1 virus. at two occasions cells infected with va187-ubi exhibited a cytopathic effect and were found to contain a subgenomic viral rna. this rna lacked the same viral genes as the subgenomic rna which has been found ...19989696875
suppression of steady-state, but not stimulus-induced nf-kappab activity inhibits alphavirus-induced apoptosis.recent studies have established cell type- specific, proapoptotic, or antiapoptotic functions for the transcription factor nf-kappab. in each of these studies, inhibitors of nf-kappab activity have been present before the apoptotic stimulus, and so the role of stimulus- induced nf-kappab activation in enhancing or inhibiting survival could not be directly assessed. sindbis virus, an alphavirus, induces nf-kappab activation and apoptosis in cultured cell lines. to address whether sindbis virus- i ...19989647642
elevated extracellular calcium can prevent apoptosis via the calcium-sensing receptor.the calcium-sensing receptor (car) is a membrane-bound, g-protein-coupled receptor present on parathyroid cells which monitors the level of extracellular calcium (ca2+o) and transduces signals involved in serum calcium regulation. expression of car protein in tissues with functions unrelated to systemic calcium homeostasis, including the brain, suggests that extracellular calcium (ca2+o) may act as a first messenger to regulate diverse cellular functions. to test this hypothesis, we examined the ...19989712695
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 ...19989721245
cleavage of highly structured viral rna molecules by combinatorial libraries of hairpin ribozymes. the most effective ribozymes are not predicted by substrate selection rules.combinatorial libraries of hairpin ribozymes representing all possible cleavage specificities (>10(5)) were used to evaluate all ribozyme cleavage sites within a large (4.2-kilobase) and highly structured viral mrna, the 26 s subgenomic rna of sindbis virus. the combinatorial approach simultaneously accounts for target site structure and dynamics, together with ribozyme folding, and the sequences that result in a ribozyme-substrate complex with maximal activity. primer extension was used to map ...19989722591
use of the sindbis replicon system for expression of lacrosse virus envelope proteins in mosquito cells.the sindbis replicon expression system was used to express la crosse (lac) virus envelope glycoprotein genes in both mammalian and mosquito cell culture. replicon expressed lac proteins had correct molecular mass (mr) and were antigenically similar to wild type lac envelope proteins. in addition, lac g1 and g2 proteins colocalized when expressed from separate constructs in both mammalian and mosquito cells suggesting that they were trafficked through the cell similarly to wild type lac proteins. ...19989722880
analysis of constructed e gene mutants of mouse hepatitis virus confirms a pivotal role for e protein in coronavirus assembly.expression studies have shown that the coronavirus small envelope protein e and the much more abundant membrane glycoprotein m are both necessary and sufficient for the assembly of virus-like particles in cells. as a step toward understanding the function of the mouse hepatitis virus (mhv) e protein, we carried out clustered charged-to-alanine mutagenesis on the e gene and incorporated the resulting mutations into the mhv genome by targeted recombination. of the four possible clustered charged-t ...19989733825
comparison of the neurovirulence of a vaccine and a wild-type mumps virus strain in the developing rat brain.prior to the adoption of widespread vaccination programs, mumps virus was the leading cause of virus-induced central nervous system (cns) disease. mumps virus-associated cns complications in vaccinees continue to be reported; outside the united states, some of these complications have been attributed to vaccination with insufficiently attenuated neurovirulent vaccine strains. the development of potentially neurovirulent, live, attenuated mumps virus vaccines stems largely from the lack of an ani ...19989733843
measles virus fusion protein is palmitoylated on transmembrane-intracytoplasmic cysteine residues which participate in cell fusion.[3h]palmitic acid was metabolically incorporated into the viral fusion protein (f) of edmonston or freshly isolated measles virus (mv) during infection of human lymphoid or vero cells. the uncleaved precursor f0 and the f1 subunit from infected cells and extracellular virus were both labeled, indicating that palmitoylation can take place prior to f0 cleavage and that palmitoylated f protein was incorporated into virus particles. [3h]palmitic acid was released from f protein upon hydroxylamine or ...19989733862
effects of site-directed mutations of transmembrane cysteines in sindbis virus e1 and e2 glycoproteins on palmitylation and virus replication.the two glycoproteins that form the external spikes of the alphaviruses are type 1 membrane proteins whose transmembrane domains of hydrophobic amino acids are close to the carboxyl termini of the polypeptides and anchor the proteins in the lipid bilayer. most of the members of the alphavirus genus contain within this transmembrane sequence one or more highly conserved cysteines, which are positioned close to the cytoplasmic face of the lipid bilayer. cysteines in the cytoplasmic domains of the ...19989740777
characterization of a baculovirus-encoded rna 5'-triphosphatase.autographa californica nuclear polyhedrosis virus (acnpv) encodes a 168-amino-acid polypeptide that contains the signature motif of the superfamily of protein phosphatases that act via a covalent cysteinyl phosphate intermediate. the sequence of the acnpv phosphatase is similar to that of the rna triphosphatase domain of the metazoan cellular mrna capping enzyme. here, we show that the purified recombinant acnpv protein is an rna 5'-triphosphatase that hydrolyzes the gamma-phosphate of triphosph ...19989696798
differential downstream functions of protein kinase ceta and -theta in el4 mouse thymoma cells.sensitive el4 mouse thymoma cells (s-el4) respond to phorbol esters with growth inhibition, adherence to substrate, and production of cytokines including interleukin 2. since these cells express several of the phorbol ester-sensitive protein kinase c (pkc) isozymes, the function of each isozyme remains unclear. previous studies demonstrated that s-el4 cells expressed substantially more pkceta and pkctheta than did el4 cells resistant to phorbol esters (r-el4). to examine potential roles for pkce ...19989765301
protection against fatal sindbis virus encephalitis by beclin, a novel bcl-2-interacting protein.bcl-2, the prototypic cellular antiapoptotic gene, decreases sindbis virus replication and sindbis virus-induced apoptosis in mouse brains, resulting in protection against lethal encephalitis. to investigate potential mechanisms by which bcl-2 protects against central nervous system sindbis virus infection, we performed a yeast two-hybrid screen to identify bcl-2-interacting gene products in an adult mouse brain library. we identified a novel 60-kda coiled-coil protein, beclin, which we confirme ...19989765397
ribosomal s27a coding sequences upstream of ubiquitin coding sequences in the genome of a pestivirus.molecular characterization of cytopathogenic (cp) bovine viral diarrhea virus (bvdv) strain cp rit, a temperature-sensitive strain widely used for vaccination, revealed that the viral genomic rna is about 15.2 kb long, which is about 2.9 kb longer than the one of noncytopathogenic (noncp) bvdv strains. molecular cloning and nucleotide sequencing of parts of the genome resulted in the identification of a duplication of the genomic region encoding nonstructural proteins ns3, ns4a, and part of ns4b ...19989765411
isolation of an arabidopsis thaliana mutant in which the multiplication of both cucumber mosaic virus and turnip crinkle virus is affected.during the systemic infection of plants by viruses, host factors play an important role in supporting virus multiplication. to identify and characterize the host factors involved in this process, we isolated an arabidopsis thaliana mutant named rb663, in which accumulation of the coat protein (cp) of cucumber mosaic virus (cmv) in upper uninoculated leaves was delayed. genetic analyses suggested that the phenotype of delayed accumulation of cmv cp in rb663 plants was controlled by a monogenic, r ...19989765416
neuronal death induced by brain-derived human immunodeficiency virus type 1 envelope genes differs between demented and nondemented aids patients.human immunodeficiency virus type 1 (hiv-1) infection of the brain results in viral replication primarily in macrophages and microglia. despite frequent detection of viral genome and proteins in the brains of aids patients with and without hiv dementia, only 20% of aids patients become demented. to investigate the role of viral envelope gene variation in the occurrence of dementia, we examined regions of variability in the viral envelope gene isolated from brains of aids patients. brain-derived ...19989765449
inhibition of nf-kappab activation in combination with bcl-2 expression allows for persistence of first-generation adenovirus vectors in the mouse is a key regulator of the innate antiviral immune response, due in part to its transcriptional activation of cytokines and adhesion molecules, which, in turn, function in chemotaxis and activation of inflammatory cells. we reported earlier that viral gene expression in hepatocytes transduced with first-generation (e1-deleted) adenoviruses induced nf-kappab activation, elevation of serum cytokines, and hepatocellular apoptosis during the first days postinfusion. these events did not occ ...19989765474
the clathrin endocytic pathway in viral important is the clathrin-dependent endocytic pathway for entry of viruses into host cells? while it is widely accepted that semliki forest virus (sfv), an enveloped virus, requires this pathway there are conflicting data concerning the closely related sindbis virus, as well as varying results with picornaviruses such as human rhinovirus 14 (hrv 14) and poliovirus. we have examined the entry mode of sfv, sindbis virus, hrv 14 and poliovirus using a method that identifies single infected cell ...19989707418
characterization of la crosse virus rna in autopsied central nervous system tissues.a reverse transcription-pcr (rt-pcr) technique was used to detect la crosse (lac) virus rna in the central nervous system (cns) tissues of two patients who died of lac encephalitis in 1960 and 1978. viral rna was readily detected by rt-pcr although the tissues had been stored frozen for up to 37 years. lac virus was detected in the cerebral cortex but not in other cns tissues. rt-pcr allowed detection of replicative forms of the virus, indicating that the virus was actively replicating in the sp ...19989774588
viral inactivation of human bone tissue using supercritical fluid extraction.a new bone tissue process using supercritical carbon dioxide fluid extraction (sfe) has been evaluated for its ability to inactivate or eliminate viruses. four viruses, human immunodeficiency virus type 1 (hiv-1), sindbis virus, polio sabin type i virus, and pseudorabies virus (prv), were exposed to four different processing steps. in addition to supercritical co2, hydrogen peroxide, sodium hydroxide, and ethanol treatments were evaluated. the mean cumulated reduction factors (log10) for the fou ...19989682954
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