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sindbis virus variant with a deletion in the 6k gene shows defects in glycoprotein processing and trafficking: lack of complementation by a wild-type 6k gene in trans.a sindbis virus (sv) variant with a 6k gene partially deleted has been obtained. this sv del6k virus is defective in the proteolytic processing of virus glycoprotein precursor, transport of glycoproteins to the plasma membrane, and plaque phenotype. a revertant virus (sv del6k-revq21l) containing a point mutation in the deleted 6k gene was isolated and characterized. sv del6k-revq21l has corrected the defects of proteolytic processing and transport of virus glycoproteins to the plasma membrane, ...200111462055
gene transfer: manipulating and monitoring function in cells and tissues.1. the ectopic expression of genes has proven to be an extremely valuable tool for biologists. the most widely used systems involve electrically or chemically mediated transfer of genes to immortalized cell lines and, at the other end of the spectrum, transgenic animal models. as would be expected, there are compromises to be made when using either of these broad approaches. immortalized cell lines have limited "physiological relevance" and transgenic approaches are costly and out of the reach o ...200111473537
genetic analysis of the pestivirus nonstructural coding region: defects in the ns5a unit can be complemented in trans.the functional analysis of molecular determinants which control the replication of pestiviruses was considerably facilitated by the finding that subgenomic forms of the positive-strand rna genome of bvdv (bovine viral diarrhea virus) are capable of autonomous replication in transfected host cells. the prototype replicon, bvdv di9c, consists of the genomic 5' and 3' untranslated regions and a truncated open reading frame (orf) encoding mainly the nonstructural proteins ns3, ns4a, ns4b, ns5a, and ...200111483722
salicylates inhibit flavivirus replication independently of blocking nuclear factor kappa b activation.flaviviruses comprise a positive-sense rna genome that replicates exclusively in the cytoplasm of infected cells. whether flaviviruses require an activated nuclear factor(s) to complete their life cycle and trigger apoptosis in infected cells remains elusive. flavivirus infections quickly activate nuclear factor kappa b (nf-kappab), and salicylates have been shown to inhibit nf-kappab activation. in this study, we investigated whether salicylates suppress flavivirus replication and virus-induced ...200111483726
antibody-directed targeting of retroviral vectors via cell surface antigens.targeted stable transduction of specific cells is a highly desirable goal for gene therapy applications. we report an efficient and broadly applicable approach for targeting retroviral vectors to specific cells. we find that the envelope of the alphavirus sindbis virus can pseudotype human immunodeficiency virus type 1- and murine leukemia virus-based retroviral vectors. when modified to contain the fc-binding domain of protein a, this envelope gives a significant enhancement in specificity in c ...200111483746
in vivo generation and characterization of a soluble form of the semliki forest virus fusion protein.during infection of host cells, a number of enveloped animal viruses are known to produce soluble forms of viral membrane glycoproteins lacking the transmembrane domain. the roles of such soluble glycoproteins in viral life cycles are incompletely understood, but in several cases they are believed to modulate host immune response and viral pathogenesis. semliki forest virus (sfv) is an enveloped alphavirus that infects cells through low-ph-dependent fusion and buds from the plasma membrane. fusi ...200111483778
abundant gfp expression and ltp in hippocampal acute slices by in vivo injection of sindbis virus.virus-mediated gene transfer into neurons is a powerful tool for the analysis of neuronal structure and function. recombinant sindbis virus has been previously used to study protein function in hippocampal neuron cultures as well as in hippocampal organotypic slice cultures. nevertheless, some concern still exists about the physiological relevance of these cultured preparations. acute hippocampal slices are a widely used preparation for the study of synaptic transmission, but currently recombina ...200111495971
n-methyl-d-aspartate-induced alpha-amino-3-hydroxy-5-methyl-4-isoxazoleproprionic acid (ampa) receptor down-regulation involves interaction of the carboxyl terminus of glur2/3 with pick1. ligand-binding studies using sindbis vectors carrying ampa receptor decoys.the dynamics of alpha-amino-3-hydroxy-5-methyl-4-isoxazoleproprionic acid (ampa)-type glutamate receptors, as represented by their exocytosis, endocytosis and cytoskeletal linkage, has often been implicated in n-methyl-d-aspartate (nmda)-dependent synaptic plasticity. to explore the molecular mechanisms underlying the ampa receptor dynamics, cultured hippocampal neurons were stimulated with 100 microm nmda, and the biochemical and pharmacological changes in the ligand binding activity of ampa re ...200111498531
[effect of sindbis xj-160 virus structural proteins on bhk-21 cell].to study the effect of sindbis xj-160 virus structural proteins on bhk-21 cell.200011498682
candidate vaccine against botulinum neurotoxin serotype a derived from a venezuelan equine encephalitis virus vector system.a candidate vaccine against botulinum neurotoxin serotype a (bont/a) was developed by using a venezuelan equine encephalitis (vee) virus replicon vector. this vaccine vector is composed of a self-replicating rna containing all of the vee nonstructural genes and cis-acting elements and also a heterologous immunogen gene placed downstream of the subgenomic 26s promoter in place of the viral structural genes. in this study, the nontoxic 50-kda carboxy-terminal fragment (h(c)) of the bont/a heavy ch ...200111500447
isolation and characterization of a mutant chinese hamster ovary cell line that is resistant to chlamydia trachomatis infection at a novel step in the attachment process.host factors involved in chlamydia trachomatis pathogenesis were investigated by random chemical mutagenesis of chinese hamster ovary (cho-k1) cells followed by selection for clones resistant to chlamydial infection. a clonal mutant cell line, d4.1-3, refractory to infection by the c. trachomatis l2 serovar was isolated. the d4.1-3 cell line appears to be lacking in a previously undescribed temperature-dependent and heparin-resistant binding step that occurs subsequent to engagement of cell surf ...200111500469
expression and functional characterization of bluetongue virus vp5 protein: role in cellular permeabilization.segment 5 of bluetongue virus (btv) serotype 10, which encodes the outer capsid protein vp5, was tagged with glutathione s-transferase and expressed by a recombinant baculovirus. the recombinant protein was subsequently purified to homogeneity, and its possible biological role in virus infection was investigated. purified vp5 was able to bind mammalian cells but was not internalized, which indicates it is not involved in receptor-mediated endocytosis. the purified vp5 protein was shown to be abl ...200111507181
pa subunit from influenza virus polymerase complex interacts with a cellular protein with homology to a family of transcriptional activators.the pa subunit of the influenza virus polymerase complex is a phosphoprotein that induces proteolytic degradation of coexpressed proteins. point mutants with reduced proteolysis induction reconstitute viral ribonucleoproteins defective in replication but not in transcriptional activity. to look for cellular factors that could associate with pa protein, we have carried out a yeast two-hybrid screen. using a human kidney cdna library, we identified two different interacting clones. one of them was ...200111507205
genetic control of neuroadapted sindbis virus replication in female mice maps to chromosome 2 and associates with paralysis and mortality.neuroadapted sindbis virus (nsv) infection of mice causes hindlimb paralysis and 100% mortality in the c57bl/6 mouse strain, while adults of the balb/cby mouse strain are resistant to fatal encephalomyelitis. levels of viral rna are higher in the brains of infected c57bl/6 mice than in balb/cby mice (d. c. thach et al., j. virol. 74:6156-6161, 2000). these phenotypic differences between the two strains allowed us to map genetic loci involved in mouse susceptibility to nsv and to find relationshi ...200111507212
gene targeting in mosquito cells: a demonstration of 'knockout' technology in extrachromosomal gene arrays.gene targeting would offer a number of advantages over current transposon-based strategies for insect transformation. these include freedom from both position effects associated with quasi-random integration and concerns over transgene instability mediated by endogenous transposases, independence from phylogenetic restrictions on transposon mobility and the ability to generate gene knockouts.200111513755
effect of prophenoloxidase expression knockout on the melanization of microfilariae in the mosquito armigeres subalbatus.melanization is an effective defence reaction used by mosquito hosts to kill malarial and filarial worm parasites. although phenoloxidase (po) has long been considered to be the key enzyme in the biosynthesis of melanotic material in insects, there is no direct evidence verifying its role in parasite melanization. to elucidate the role of po in the melanization of microfilariae (mf) by mosquitoes, a double subgenomic sindbis (dssin) recombinant virus was used to transduce armigeres subalbatus mo ...200111520354
5'-nor carbocyclic 5'-deoxy-5'-(isobutylthio)adenosine and a 2',3'-dideoxy-2',3'-didehydro derivative.the inhibition of biochemical processes requiring s-adenosylmethionine as a co-factor have led to many nucleoside-based medicinal agents. included in this group are 5'-deoxy-5'-(isobutylthio)adenosine (siba), a nucleoside with antiparasitic, antiviral and antiproliferative effects, and 5'-noraristeromycin, a carbocyclic-derived nucleoside with potent antiviral properties. this report brings together the structural components of these two compounds by describing both enantiomers of carbocyclic 5- ...200111527043
mutant prion proteins are partially retained in the endoplasmic reticulum.familial prion diseases are linked to point and insertional mutations in the prion protein (prp) gene that are presumed to favor conversion of the cellular isoform of prp to the infectious isoform. in this report, we have investigated the subcellular localization of prp molecules carrying pathogenic mutations using immunofluorescence staining, immunogold labeling, and prp-green fluorescent protein chimeras. to facilitate visualization of the mutant proteins, we have utilized a novel sindbis vira ...200111527974
receptors and entry cofactors for retroviruses include single and multiple transmembrane-spanning proteins as well as newly described glycophosphatidylinositol-anchored and secreted proteins.in the past few years, many retrovirus receptors, coreceptors, and cofactors have been identified. these molecules are important for some aspects of viral entry, although in some cases it remains to be determined whether they are required for binding or postbinding stages in entry, such as fusion. there are certain common features to the molecules that many retroviruses use to gain entry into the cell. for example, the receptors for most mammalian oncoretroviruses are multiple membrane-spanning ...200111528001
cooperation of an rna packaging signal and a viral envelope protein in coronavirus rna packaging.murine coronavirus mouse hepatitis virus (mhv) produces a genome-length mrna, mrna 1, and six or seven species of subgenomic mrnas in infected cells. among these mrnas, only mrna 1 is efficiently packaged into mhv particles. mhv n protein binds to all mhv mrnas, whereas envelope m protein interacts only with mrna 1. this m protein-mrna 1 interaction most probably determines the selective packaging of mrna 1 into mhv particles. a short cis-acting mhv rna packaging signal is necessary and sufficie ...200111533169
high resistance of human parainfluenza type 2 virus protein-expressing cells to the antiviral and anti-cell proliferative activities of alpha/beta interferons: cysteine-rich v-specific domain is required for high resistance to the interferons.human parainfluenza type 2 virus (hpiv-2)-infected hela (hela-ca) cells and hpiv-2 v-expressing hela (hela-v) cells show high resistance to alpha/beta interferons (ifn-alpha/beta) irrespective of whether vesicular stomatitis virus or sindbis virus is used as a challenge virus. when sindbis virus is used, these cells show high susceptibility to human ifn-gamma. furthermore, the multiplication of hela-v cells is not inhibited by ifn-alpha/beta. hela cells expressing the n-terminally truncated v pr ...200111533180
additive effects characterize the interaction of antibodies involved in neutralization of the primary dualtropic human immunodeficiency virus type 1 isolate 89.6.human immunodeficiency virus-type i (hiv-1) infection elicits antibodies (abs) directed against several regions of the gp120 and gp41 envelope glycoproteins. many of these abs are able to neutralize t-cell-line-adapted strains (tcla) of hiv-1, but only a few effectively neutralize primary hiv-1 isolates. the nature of hiv-1 neutralization has been carefully studied using human monoclonal abs (mabs), and the ability of such mabs to act in synergy to neutralize hiv-1 has also been extensively stud ...200111533181
chimeric bovine respiratory syncytial virus with attachment and fusion glycoproteins replaced by bovine parainfluenza virus type 3 hemagglutinin-neuraminidase and fusion proteins.chimeric bovine respiratory syncytial viruses (brsv) expressing glycoproteins of bovine parainfluenza virus type 3 (bpiv-3) instead of brsv glycoproteins were generated from cdna. in the brsv antigenome cdna, the open reading frames of the major brsv glycoproteins, attachment protein g and fusion protein f, were replaced individually or together by those of the bpiv-3 hemagglutinin-neuraminidase (hn) and/or fusion (f) glycoproteins. recombinant virus could not be recovered from cdna when the brs ...200111533200
venezuelan equine encephalomyelitis virus structure and its divergence from old world alphaviruses.although alphaviruses have been extensively studied as model systems for the structural organization of enveloped viruses, no structures exist for the phylogenetically distinct eastern equine encephalomyelitis (eee)-venezuelan equine encephalomyelitis (vee) lineage of new world alphaviruses. here we report the 25-a structure of vee virus, obtained from electron cryomicroscopy and image reconstruction. the envelope spike glycoproteins of vee virus have a t=4 icosahedral arrangement, similar to th ...200111533216
genetic selection and the lure of sin. 200111533637
a high-throughput alphavirus-based expression cloning system for mammalian cells.we have developed a widely applicable functional genomics strategy based on alphavirus expression vectors. the technology allows for rapid identification of genes encoding a functional activity such as binding of a defined ligand. complementary dna (cdna) libraries were expressed in mammalian cells following infection with recombinant sindbis virus (sin replicon particles), a member of the alphavirus genus. virus-infected cells that specifically bound a ligand of choice were isolated using fluor ...200111533644
membrane topography and topogenesis of prenylated rab acceptor (pra1).the mouse prenylated rab acceptor (mpra1) is associated with the golgi membrane at steady state and interacts with rab proteins. it contains two internal hydrophobic domains (34 residues each) that have enough residues to form four transmembrane (tm) segments. in this study, we have determined the membrane topography of mpra1 in both intact cells and isolated microsomes. the putative tm segments of mpra1 were used to substitute for a known tm segment of a model membrane protein to determine whet ...200111535589
complete, long-lasting protection against malaria of mice primed and boosted with two distinct viral vectors expressing the same plasmodial antigen.we report that complete protection against malaria and total inhibition of liver stage development and parasitemia was obtained in 100% of balb/c mice primed with a replication-defective recombinant adenovirus expressing the circumsporozoite (cs) protein of plasmodium yoelii (adpycs), followed by a booster with an attenuated recombinant vaccinia virus, expressing the same malaria antigen, vacpycs. we found increased levels of activated cs-specific cd8(+) and cd4(+) t cells, higher anti-sporozoit ...200111553779
interaction of classical swine fever virus with membrane-associated heparan sulfate: role for virus replication in vivo and virulence.passage of native classical swine fever virus (csfv) in cultured swine kidney cells (sk6 cells) selects virus variants that attach to the surface of cells by interaction with membrane-associated heparan sulfate (hs). a ser-to-arg change in the c terminus of envelope glycoprotein e(rns) (amino acid 476 in the open reading frame of csfv) is responsible for selection of these hs-binding virus variants (m. m. hulst, h. g. p. van gennip, and r. j. m. moormann, j. virol. 74:9553-9561, 2000). in this i ...200111559790
frequency of spontaneous mutations in an avian hepadnavirus infection.in this study, we measured the frequency of revertants of a cytopathic strain of the duck hepatitis b virus that bears a single nucleotide substitution in the pre-s envelope protein open reading frame, resulting in the amino acid substitution g133e. cytopathic virus mixed with known amounts of a genetically marked wild-type virus was injected into ducklings. virus outgrowth was accompanied by a coselection of wild-type and spontaneous revertants during recovery of the ducklings from the acute li ...200111559794
resistance to nucleoside analog reverse transcriptase inhibitors mediated by human immunodeficiency virus type 1 p6 protein.resistance of human immunodeficiency virus type 1 (hiv-1) to antiretroviral agents results from target gene mutation within the pol gene, which encodes the viral protease, reverse transcriptase (rt), and integrase. we speculated that mutations in genes other that the drug target could lead to drug resistance. for this purpose, the p1-p6(gag)-p6(pol) region of hiv-1, placed immediately upstream of pol, was analyzed. this region has the potential to alter pol through frameshift regulation (p1), th ...200111559796
synthesis and methyltransferase activity of nonstructural protein nsp1 in sindbis virus-infected aedes albopictus cells.to investigate the synthesis and turnover of the nonstructure protein, nsp1, in aedes albopictus cells, we labeled infected cells with [35s]-methionine and immunoprecipitated nsp1 with a polyclonal monospecific rabbit antibody. synthesis of nsp1 in mosquito cells could be detected 2 hr after infection and continued as long as 24 hr post-infection, regardless whether the infected cells were maintained at 28 degrees c , 34.5 degrees c , or 37 degrees c. whereas the time pattern of nsp1 synthesis v ...199911561581
murine gammaherpesvirus-68-induced interleukin-10 increases viral burden, but limits virus-induced splenomegaly and leukocytosis.based on its genomic sequence and its pathogenesis, murine gammaherpesvirus-68 (gammahv-68) has been established as a tractable model for the study of viral infections caused by the human gammaherpesviruses, epstein-barr virus or human herpesvirus-8. despite significant advances, the mechanisms responsible for gammahv-68-induced alterations in the protective host response, and the accompanying virus-induced leukocytosis, are not clear. in the present study, we questioned whether viral infection ...200111576228
phosphatidylinositol-4-phosphate 5-kinase-1beta is essential for epidermal growth factor receptor-mediated endocytosis.phosphatidylinositol-4,5-bisphosphate (pip(2)) is known to play an important role in signal transduction and membrane trafficking. we show that one enzyme responsible for pip(2) production, phosphatidylinositol-4-phosphate 5-kinase type 1beta (pipkbeta), is essential for epidermal growth factor receptor (egfr)-mediated endocytosis. expression of murine pipkbeta in nr6 cells expressing egfr strikingly increased receptor internalization. moreover, the kinase was shown to form an immunoprecipitable ...200111581249
evolutionary relationships and systematics of the alphaviruses.partial e1 envelope glycoprotein gene sequences and complete structural polyprotein sequences were used to compare divergence and construct phylogenetic trees for the genus alphavirus. tree topologies indicated that the mosquito-borne alphaviruses could have arisen in either the old or the new world, with at least two transoceanic introductions to account for their current distribution. the time frame for alphavirus diversification could not be estimated because maximum-likelihood analyses indic ...200111581380
identification of genes involved in the host response to neurovirulent alphavirus infection.single-amino-acid mutations in sindbis virus proteins can convert clinically silent encephalitis into uniformly lethal disease. however, little is known about the host gene response during avirulent and virulent central nervous system (cns) infections. to identify candidate host genes that modulate alphavirus neurovirulence, we utilized genechip expression analysis to compare cns gene expression in mice infected with two strains of sindbis virus that differ by one amino acid in the e2 envelope g ...200111581411
cd8(+) t lymphocytes mediate borna disease virus-induced immunopathology independently of perforin.perforin-mediated lysis of target cells is the major antiviral effector mechanism of cd8(+) t lymphocytes. we have analyzed the role of perforin in a mouse model for cd8(+) t-cell-mediated central nervous system (cns) immunopathology induced by borna disease virus. when a defective perforin gene was introduced into the genetic background of the borna disease-susceptible mouse strain mrl, the resulting perforin-deficient mice developed strong neurological disease in response to infection indistin ...200111581414
the synthetic peptide p-197 inhibits human t-cell leukemia virus type 1 envelope-mediated syncytium formation by a mechanism that is independent of hsc70.entry of human t-cell leukemia virus type 1 (htlv-1) into cells is mediated by the viral envelope glycoproteins gp46 and gp21. the gp46 surface glycoprotein binds to a poorly characterized cell surface receptor, thereby promoting the gp21-dependent fusion of the viral and cellular membranes. interestingly, a synthetic peptide (p-197) simulating amino acids 197 to 216 of gp46 strongly inhibits envelope-dependent membrane fusion with molt-4 target cells. it has been suggested that this peptide act ...200111581416
antiviral actions of interferons.tremendous progress has been made in understanding the molecular basis of the antiviral actions of interferons (ifns), as well as strategies evolved by viruses to antagonize the actions of ifns. furthermore, advances made while elucidating the ifn system have contributed significantly to our understanding in multiple areas of virology and molecular cell biology, ranging from pathways of signal transduction to the biochemical mechanisms of transcriptional and translational control to the molecula ...011585785
ross river virus transmission, infection, and disease: a cross-disciplinary review.ross river virus (rrv) is a fascinating, important arbovirus that is endemic and enzootic in australia and papua new guinea and was epidemic in the south pacific in 1979 and 1980. infection with rrv may cause disease in humans, typically presenting as peripheral polyarthralgia or arthritis, sometimes with fever and rash. rrv disease notifications in australia average 5,000 per year. the first well-described outbreak occurred in 1928. during world war ii there were more outbreaks, and the name ep ...200111585790
tackling the most difficult diseases. genetics and genomics open new strategies to fight vector-borne diseases. 011600446
poliovirus 5'-terminal cloverleaf rna is required in cis for vpg uridylylation and the initiation of negative-strand rna synthesis.chimeric poliovirus rnas, possessing the 5' nontranslated region (ntr) of hepatitis c virus in place of the 5' ntr of poliovirus, were used to examine the role of the poliovirus 5' ntr in viral replication. the chimeric viral rnas were incubated in cell-free reaction mixtures capable of supporting the sequential translation and replication of poliovirus rna. using preinitiation rna replication complexes formed in these reactions, we demonstrated that the 3' ntr of poliovirus rna was insufficient ...200111602711
c terminus of infectious bursal disease virus major capsid protein vp2 is involved in definition of the t number for capsid assembly.infectious bursal disease virus (ibdv), a member of the birnaviridae family, is a double-stranded rna virus. the ibdv capsid is formed by two major structural proteins, vp2 and vp3, which assemble to form a t=13 markedly nonspherical capsid. during viral infection, vp2 is initially synthesized as a precursor, called vpx, whose c end is proteolytically processed to the mature form during capsid assembly. we have computed three-dimensional maps of ibdv capsid and virus-like particles built up by v ...200111602723
neuroadapted yellow fever virus 17d: genetic and biological characterization of a highly mouse-neurovirulent virus and its infectious molecular clone.a neuroadapted strain of yellow fever virus (yfv) 17d derived from a multiply mouse brain-passaged virus (porterfield yf17d) was additionally passaged in scid and normal mice. the virulence properties of this virus (spyf) could be distinguished from nonneuroadapted virus (yf5.2iv, 17d infectious clone) by decreased average survival time in scid mice after peripheral inoculation, decreased average survival time in normal adult mice after intracerebral inoculation, and occurrence of neuroinvasiven ...200111602731
human immunodeficiency virus type 1 entry into macrophages mediated by macropinocytosis.whereas human immunodeficiency virus (hiv) infects various cell types by fusion at the plasma membrane, we observed a different entry route in human primary macrophages, in which macropinocytosis is active. shortly after exposure of macrophages to hiv-1 and irrespective of viral envelope-receptor interactions, particles were visible in intracellular vesicles, which were identified as macropinosomes. most virions appeared subsequently degraded. however, fusion leading to capsid release in the cyt ...200111602756
pe2 cleavage mutants of sindbis virus: correlation between viral infectivity and ph-dependent membrane fusion activation of the spike heterodimer.the spike glycoprotein e2 of sindbis virus (sin) is synthesized in the infected cell as a pe2 precursor protein, which matures through cleavage by a cellular furin-like protease. previous work has shown that sin mutants impaired in pe2 cleavage are noninfectious on bhk-21 cells, the block in infection being localized at a step after virus-receptor interaction but prior to rna replication. here, we studied the membrane fusion properties of sin pe2 cleavage mutants and observed that these viruses ...200111602759
visualization of ip(3) dynamics reveals a novel ampa receptor-triggered ip(3) production pathway mediated by voltage-dependent ca(2+) influx in purkinje cells.ip(3) signaling in purkinje cells is involved in the regulation of cell functions including ltd. we have used a gfp-tagged pleckstrin homology domain to visualize ip(3) dynamics in purkinje cells. surprisingly, ip(3) production was observed in response not only to mglur activation, but also to ampa receptor activation in purkinje cells in culture. ampa-induced ip(3) production was mediated by depolarization-induced ca(2+) influx because it was mimicked by depolarization and was blocked by inhibi ...200111604143
expression of serum amyloid a genes in mouse brain: unprecedented response to inflammatory mediators.serum amyloid a (saa) proteins were originally identified as prominent acute-phase serum proteins synthesized predominantly by hepatocytes. these small proteins are remarkably lipophilic, and we have sought evidence for their synthesis in mouse brain. rt-pcr showed constitutive expression of the murine saa1 gene in the brains of normal balb/cj mice. after intracerebral inoculation with sindbis virus, these mice predictably increase brain expression of tumor necrosis factor alpha (tnf-alpha), int ...200111641251
the 37-kda/67-kda laminin receptor acts as the cell-surface receptor for the cellular prion protein.recently, we identified the 37-kda laminin receptor precursor (lrp) as an interactor for the prion protein (prp). here, we show the presence of the 37-kda lrp and its mature 67-kda form termed high-affinity laminin receptor (lr) in plasma membrane fractions of n2a cells, whereas only the 37-kda lrp was detected in baby hamster kidney (bhk) cells. prp co-localizes with lrp/lr on the surface of n2a cells and semliki forest virus (sfv) rna transfected bhk cells. cell-binding assays reveal the lrp/l ...200111689427
functional interactions of human immunodeficiency virus type 1 integrase with human and yeast hsp60.integration of human immunodeficiency virus type 1 (hiv-1) proviral dna in the nuclear genome is catalyzed by the retroviral integrase (in). in addition to in, viral and cellular proteins associated in the high-molecular-weight preintegration complex have been suggested to be involved in this process. in an attempt to define host factors interacting with in, we used an in vitro system to identify cellular proteins in interaction with hiv-1 in. the yeast saccharomyces cerevisiae was chosen since ...200111689615
palmitoylation of the rous sarcoma virus transmembrane glycoprotein is required for protein stability and virus infectivity.the rous sarcoma virus (rsv) transmembrane (tm) glycoprotein is modified by the addition of palmitic acid. to identify whether conserved cysteines within the hydrophobic anchor region are the site(s) of palmitoylation, and to determine the role of acylation in glycoprotein function, cysteines at residues 164 and 167 of the tm protein were mutated to glycine (c164g, c167g, and c164g/c167g). in cv-1 cells, palmitate was added to env gene products containing single mutations but was absent in the d ...200111689636
cell surface heparan sulfate is a receptor for human herpesvirus 8 and interacts with envelope glycoprotein k8.1.an immunodominant envelope glycoprotein is encoded by the human herpesvirus 8 (hhv-8) (also termed kaposi's sarcoma-associated herpesvirus) k8.1 gene. the functional role of glycoprotein k8.1 is unknown, and recognizable sequence homology to k8.1 is not detectable in the genomes of most other closely related gammaherpesviruses, such as herpesvirus saimiri or epstein-barr virus. in search for a possible function for k8.1, we expressed the ectodomain of k8.1 fused to the fc part of human immunoglo ...200111689640
flock house virus rna replicates on outer mitochondrial membranes in drosophila cells.the identification and characterization of host cell membranes essential for positive-strand rna virus replication should provide insight into the mechanisms of viral replication and potentially identify novel targets for broadly effective antiviral agents. the alphanodavirus flock house virus (fhv) is a positive-strand rna virus with one of the smallest known genomes among animal rna viruses, and it can replicate in insect, plant, mammalian, and yeast cells. to investigate the localization of f ...200111689648
construction and analysis of mouse strains lacking the ubiquitin ligase ubr1 (e3alpha) of the n-end rule pathway.the n-end rule relates the in vivo half-life of a protein to the identity of its n-terminal residue. in the yeast saccharomyces cerevisiae, the ubr1-encoded ubiquitin ligase (e3) of the n-end rule pathway mediates the targeting of substrate proteins in part through binding to their destabilizing n-terminal residues. the functions of the yeast n-end rule pathway include fidelity of chromosome segregation and the regulation of peptide import. our previous work described the cloning of cdna and a g ...200111689692
biological activity of guatteria cardoniana fractions.methanolic extract of leaves and twigs of guatteria cardoniana r.e. fries (annonaceae), a plant from the venezuelan rain forest, was separated in alkaloid rich fractions and their biological effect on baby hamster kidney (bhk) cell line was studied. the initial plant extract (fa) induced cell proliferation, cytotoxicity as well as antiviral activity, depending on the concentration used. further separation of this methanolic extract allowed us to separate these biological activities. the fraction ...200111694356
prolonged arthritis associated with sindbis-related (pogosta) virus infection. 200111709622
molecular determinants of virulence, cell tropism, and pathogenic phenotype of infectious bursal disease virus.infectious bursal disease viruses (ibdvs), belonging to the family birnaviridae, exhibit a wide range of immunosuppressive potential, pathogenicity, and virulence for chickens. the genomic segment a encodes all the structural (vp2, vp4, and vp3) and nonstructural proteins, whereas segment b encodes the viral rna-dependent rna polymerase (vp1). to identify the molecular determinants for the virulence, pathogenic phenotype, and cell tropism of ibdv, we prepared full-length cdna clones of a virulen ...200111711587
the helicase-like domain of plant potexvirus replicase participates in formation of rna 5' cap structure by exhibiting rna 5'-triphosphatase activity.open reading frame 1 (orf1) of potexviruses encodes a viral replicase comprising three functional domains: a capping enzyme at the n terminus, a putative helicase in the middle, and a polymerase at the c terminus. to verify the enzymatic activities associated with the putative helicase domain, the corresponding cdna fragment from bamboo mosaic virus (bamv) was cloned into vector pet32 and the protein was expressed in escherichia coli and purified by metal affinity chromatography. an activity ass ...200111711602
functional specialization and evolution of leader proteinases in the family closteroviridae.members of the closteroviridae and potyviridae families of the plant positive-strand rna viruses encode one or two papain-like leader proteinases. in addition to a c-terminal proteolytic domain, each of these proteinases possesses a nonproteolytic n-terminal domain. we compared functions of the several leader proteinases using a gene swapping approach. the leader proteinase (l-pro) of beet yellows virus (byv; a closterovirus) was replaced with l1 or l2 proteinases of citrus tristeza virus (ctv; ...200111711606
matrix protein and another viral component contribute to induction of apoptosis in cells infected with vesicular stomatitis virus.the induction of apoptosis in host cells is a prominent cytopathic effect of vesicular stomatitis virus (vsv) infection. the viral matrix (m) protein is responsible for several important cytopathic effects, including the inhibition of host gene expression and the induction of cell rounding in vsv-infected cells. this raises the question of whether m protein is also involved in the induction of apoptosis. hela or bhk cells were transfected with m mrna to determine whether m protein induces apopto ...200111711608
intracellular trafficking of the ul11 tegument protein of herpes simplex virus type 1.growing evidence indicates that herpes simplex virus type 1 (hsv-1) acquires its final envelope in the trans-golgi network (tgn). during the envelopment process, the viral nucleocapsid as well as the envelope and tegument proteins must arrive at this site in order to be incorporated into assembling virions. to gain a better understanding of how these proteins associate with cellular membranes and target to the correct compartment, we have been studying the intracellular trafficking properties of ...200111711612
identification of sequences in brome mosaic virus replicase protein 1a that mediate association with endoplasmic reticulum membranes.rna replication of all positive-strand rna viruses is closely associated with intracellular membranes. brome mosaic virus (bmv) rna replication occurs on the perinuclear region of the endoplasmic reticulum (er), both in its natural plant host and in the yeast saccharomyces cerevisiae. the only viral component in the bmv rna replication complex that localizes independently to the er is 1a, a multifunctional protein with an n-terminal rna capping domain and a c-terminal helicase-like domain. the o ...200111711627
ribin, a protein encoded by a message complementary to rrna, modulates ribosomal transcription and cell proliferation.the control of rrna transcription, tightly coupled to the cell cycle and growth state of the cell, is a key process for understanding the mechanisms that drive cell proliferation. here we describe a novel protein, ribin, found in rodents, that binds to the rrna promoter and stimulates its activity. the protein also interacts with the basal rrna transcription factor ubf. the open reading frame encoding ribin is 96% complementary to a central region of the large rrna. this demonstrates that riboso ...200111713263
alphavirus dna and particle replicons for vaccines and gene therapy.alphaviruses have several features that make them attractive as gene delivery platforms, and vectors derived principally from sindbis virus (sin), semliki forest virus (sfv), and venezuelan equine encephalitis virus (vee), are currently being developed as prophylactic and therapeutic vaccines for infectious diseases and cancer. alphavirus vectors, termed "replicons", retain the nonstructural protein genes encoding the viral replicase, that in turn programme high level cytoplasmic amplification o ...200011713818
non-viral amplification systems for gene transfer: vectors based on alphaviruses.non-viral self-replicating vectors based on defective viral genomes have been developed for a number of different alphaviruses including semliki forest virus (sfv), sindbis virus (sin) and venezuelan equine encephalitis virus (vee). these vectors can be used for gene delivery as naked rna or dna. recombinant alphavirus rna can be synthesized in vitro from plasmids containing the alphavirus replicon under the control of a prokaryotic promoter such as sp6 or t7. these self-replicating rnas have be ...199911715947
cis-acting rna elements at the 5' end of sindbis virus genome rna regulate minus- and plus-strand rna synthesis.alphavirus genome replication is a multistep asymmetric process. several lines of evidence suggest that the template preference of the rna replicase is regulated by proteolytic cleavage of the viral nonstructural polyprotein. cis-acting rna elements in the viral genome also play crucial roles in regulating genome replication and subgenomic rna transcription. in this report, a series of rna templates were analyzed in vitro and in vivo to define functional elements in the 5' end of the genome. the ...200111720292
hydrostatic pressure induces the fusion-active state of enveloped viruses.enveloped animal viruses must undergo membrane fusion to deliver their genome into the host cell. we demonstrate that high pressure inactivates two membrane-enveloped viruses, influenza and sindbis, by trapping the particles in a fusion-intermediate state. the pressure-induced conformational changes in sindbis and influenza viruses were followed using intrinsic and extrinsic fluorescence spectroscopy, circular dichroism, and fusion, plaque, and hemagglutination assays. influenza virus subjected ...200211723114
in vivo transduction of central neurons using recombinant sindbis virus: golgi-like labeling of dendrites and axons with membrane-targeted fluorescent proteins.a new recombinant virus which labeled the infected neurons in a golgi stain-like fashion was developed. the virus was based on a replication-defective sindbis virus and was designed to express green fluorescent protein with a palmitoylation signal (palgfp). when the virus was injected into the ventrobasal thalamic nuclei, many neurons were visualized with the fluorescence of palgfp in the injection site. the labeling was enhanced by immunocytochemical staining with an antibody to green fluoresce ...200111724897
clearance of extracellular and cell-associated amyloid beta peptide through viral expression of neprilysin in primary neurons.amyloid beta peptide (abeta), the pathogenic agent of alzheimer's disease (ad), is a physiological metabolite constantly anabolized and catabolized in the brain. we previously demonstrated that neprilysin is the major abeta-degrading enzyme in vivo. to investigate whether or not manipulation of neprilysin activity in the brain would be an effective strategy for regulating abeta levels, we expressed neprilysin in primary cortical neurons using a sindbis viral vector and examined the effect on abe ...200111726269
misreading of termination codons in eukaryotes by natural nonsense suppressor trnas.translational stop codon readthrough provides a regulatory mechanism of gene expression that is extensively utilised by positive-sense ssrna viruses. the misreading of termination codons is achieved by a variety of naturally occurring suppressor trnas whose structure and function is the subject of this survey. all of the nonsense suppressors characterised to date (with the exception of selenocysteine trna) are normal cellular trnas that are primarily needed for reading their cognate sense codons ...011726686
alphavirus vectors: development and potential therapeutic applications.alphaviruses are rna enveloped viruses that are proving their value as expression vectors. they are particularly well-suited for this role as they are easily and quickly engineered and can be used to produce high levels of proteins of interest. a promising and important use is as vaccines against disease-causing agents such as hiv. the three alphaviruses now serving as vectors are sindbis virus, semliki forest virus (sfv) and venezuelan equine encephalitis (vee) virus. sindbis virus and sfv are ...200111727528
antiherpetic activity of dimeric derivatives of netropsin. 200111727562
increased efficacy of the immunoglobulin g2a subclass in antibody-mediated protection against lactate dehydrogenase-elevating virus-induced polioencephalomyelitis revealed with switch mutants.immunoglobulin g1 (igg1), igg2a, and igg2b switch variants were derived from an igg3 monoclonal antibody directed against the vp3 envelope glycoprotein of lactate dehydrogenase-elevating virus (ldv). among the four antibodies, igg2a delayed the onset and progression of ldv-induced polioencephalomyelitis more than did the other subclasses. this suggests that the igg2a predominance observed in many igg antibody responses elicited by live viruses could, at least under some circumstances, correspond ...200211739710
human immunodeficiency virus type 1 gag-specific vaginal immunity and protection after local immunizations with sindbis virus-based replicon particles.the majority of human immunodeficiency virus (hiv) infections occur through vaginal and rectal transmission. in seeking a safe, nonreplicating gene-delivery vector that can induce mucosal and systemic immune responses and protection, sindbis virus-based replicon particles expressing hiv-1 gag (sin-gag) were developed. in mice, after nasal or intramuscular immunization with sin-gag and vaginal challenge with vaccinia virus (vv) expressing hiv-1 gag (vv-gag), cd8(+) t cell-mediated responses were ...200111740739
interferon-stimulated response element (isre)-binding protein complex draf1 is activated in sindbis virus (hr)-infected cells.to elucidate the host cell defense mechanisms in response to sindbis viral infection, we have started to characterize interferon (ifn)-stimulated response element (isre)-binding proteins activated in infected cells that are involved in the transcriptional induction of ifn type i-inducible genes. using electromobility shift assays (emsa), we detected several protein complexes with a human ifn-stimulated gene 15 (isg15) isre in extracts from virus-infected l929 cells that were absent in extracts f ...200111747630
the p23 protein of citrus tristeza virus controls asymmetrical rna accumulation.citrus tristeza virus (ctv), a member of the closteroviridae, has a 19.3-kb positive-stranded rna genome that is organized into 12 open reading frames (orfs) with the 10 3' genes expressed via a nested set of nine or ten 3'-coterminal subgenomic mrnas (sgrnas). relatively large amounts of negative-stranded rnas complementary to both genomic and sgrnas accumulate in infected cells. as is characteristic of rna viruses, wild-type ctv produced more positive than negative strands, with the plus-to-mi ...200211752137
systemic, mucosal, and heterotypic immune induction in mice inoculated with venezuelan equine encephalitis replicons expressing norwalk virus-like particles.norwalk-like viruses (nlvs) are a diverse group of single-stranded, nonenveloped, positive-polarity rna viruses and are the leading cause of epidemic acute gastroenteritis in the united states. in this study, the major capsid gene of norwalk virus, the prototype nlv, has been cloned and expressed in mammalian cells using a venezuelan equine encephalitis (vee) replicon expression system. upon infection of baby hamster kidney (bhk) cells with vee replicon particles (vrps), the norwalk virus capsid ...200211752163
vflip protects pc-12 cells from apoptosis induced by sindbis virus: implications for the role of tnf-alpha.sindbis virus (sv) is an alphavirus used as a model for studying the pathogenesis of viral encephalitis. in this study we examined the effects and the mechanisms involved in the apoptosis induced by sv in pc-12 cells, and the role of a vflip in this process. infection of pc-12 cells with a neurovirulent strain of sv, svni, induced cell apoptosis. overexpression of vflip encoded by the hhv-8 or treatment with a caspase-8 inhibitor inhibited cell apoptosis. svni induced an increase in the expressi ...200111753570
recombinant sindbis virus allows expression and precise targeting of proteins of the contractile apparatus in cultured cardiomyocytes.expression of epitope-tagged sarcomeric proteins in cardiomyocytes is a powerful approach for the characterization of interacting domains. here, we report a new strategy for the study of the targeting of contractile proteins in cardiomyocytes by sindbis virus (sin)-mediated gene transfer. two recombinant sin were generated, one encoding the myosin-light chain mlc3f-egfp fusion protein (sinrep5/mlc3f-egfp), and the other encoding the alpha-actinin-dsred fusion protein (sinrep5/alpha-actinin-dsred ...200111770082
molecular dissection of the semliki forest virus homotrimer reveals two functionally distinct regions of the fusion protein.semliki forest virus (sfv) is an enveloped alphavirus that infects cells via a membrane fusion reaction triggered by the acidic ph of endosomes. in response to low ph, the e1 proteins on the virus membrane undergo a series of conformational changes, resulting in the formation of a stable e1 homotrimer. little is known about the structural basis of either the e1 conformational changes or the resulting homotrimer or about the mechanism of action of the homotrimer in fusion. here, the e1 homotrimer ...200211773395
molecular chaperone hsp90 is important for vaccinia virus growth in cells.molecular chaperones assist protein folding, and some chaperones are induced by heat, nutrient depletion, or pathogen invasion. this study investigates the role played by hsp90 in the life cycle of vaccinia virus. the titer of vaccinia intracellular mature virions (imv) was reduced by 2 orders of magnitude in rk13 cells treated with geldanamycin (ga), which blocks the atpase activity of hsp90. ga does not affect expression from the viral early promoter, but treatment with ga delays dna replicati ...200211773412
heterologous gene expression from transmissible gastroenteritis virus replicon particles.we have recently isolated a transmissible gastroenteritis virus (tgev) infectious construct designated tgev 1000 (b. yount, k. m. curtis, and r. s. baric, j. virol. 74:10600-10611, 2000). using this construct, a recombinant tgev was constructed that replaced open reading frame (orf) 3a with a heterologous gene encoding green fluorescent protein (gfp). following transfection of baby hamster kidney (bhk) cells, a recombinant tgev (tgev-gfp2) was isolated that replicated efficiently and expressed g ...200211773416
mullerian-inhibiting substance regulates nf-kappa b signaling in the prostate in vitro and in vivo.mullerian-inhibiting substance (mis) is a member of the transforming growth factor beta superfamily, a class of molecules that regulates growth, differentiation, and apoptosis in many cells. mis type ii receptor in the mullerian duct is temporally and spatially regulated during development and becomes restricted to the most caudal ends that fuse to form the prostatic utricle. in this article, we have demonstrated mis type ii receptor expression in the normal prostate, human prostate cancer cell ...200211773638
trna elements mediate the assembly of an icosahedral rna virus.trnas, the adapter molecules in protein synthesis, also serve as metabolic cofactors and as primers for viral rna-directed dna synthesis. the genomic and subgenomic rnas of some plant viruses have a 3'-terminal trna-like structure (tls) that can accept a specific amino acid and serve as a site for initiation of replication and as a simple telomere. we report a previously undescribed role for the tls of brome mosaic virus (bmv), and potentially for cellular trna, in mediating the assembly of its ...200211782536
surface accessibility of the 70-kilodalton chlamydia trachomatis heat shock protein following reduction of outer membrane protein disulfide bonds.numerous investigations have shown that 70-kda heat shock protein (hsp70) homologs interact tightly with hydrophobic proteins and functionally assist proteins in membranous organelles and environments. one such protein is the chlamydia trachomatis hsp70 that is associated with isolated outer membrane complexes of infectious elementary bodies (eb). previous observations have indicated that chlamydial hsp70 plays a role in eb attachment to, or entry into, endometrial epithelial cells. in this stud ...200211796580
virion disassembly is required for apoptosis induced by reovirus.reovirus infection leads to apoptosis in cultured cells and in vivo. binding of viral attachment protein final sigma 1 to both sialic acid and junction adhesion molecule is required for induction of apoptosis. however, it is not known whether viral engagement of receptors is sufficient to elicit this cellular response. to determine whether steps in reovirus replication subsequent to viral attachment are required for reovirus-induced apoptosis, we used inhibitors of viral disassembly and rna synt ...200211799158
positively charged amino acid substitutions in the e2 envelope glycoprotein are associated with the emergence of venezuelan equine encephalitis virus.epidemic-epizootic venezuelan equine encephalitis (vee) viruses (veev) have emerged repeatedly via convergent evolution from enzootic predecessors. however, previous sequence analyses have failed to identify common sets of nucleotide or amino acid substitutions associated with all emergence events. during 1993 and 1996, veev subtype ie epizootics occurred on the pacific coast of the states of chiapas and oaxaca in southern mexico. like other epizootic veev strains, when inoculated into guinea pi ...200211799167
behavior of a photoactivatable analog of cholesterol, 6-photocholesterol, in model membranes.6-photocholesterol, a new photoactivatable analog of cholesterol in which a diazirine functionality replaces the 5,6-double bond in the steroid nucleus, was used recently to identify cholesterol-binding proteins in neuroendocrine cells [thiele, c., hannah, m.j., farenholz, f. and huttner, w.b. (2000) nat. cell biol. 2, 42-49], to track the distribution and transport of cholesterol in caenorhabditis elegans [matyash, v., geier, c., henske, a., mukherjee, s., hirsh, d., thiele, c., grant, b., maxf ...200211801250
inhibiting apoptosis in mammalian cell culture using the caspase inhibitor xiap and deletion mutants.lower yields and poorer quality of biopharmaceutical products result from cell death in bioreactors. such cell death may occur from necrosis but is more commonly associated with apoptosis. during the process of programmed cell death or apoptosis, caspases become activated and cause a cascade of events that eventually destroy the cell. xiap is the most potent caspase inhibitor encoded in the mammalian genome. the effectiveness of xiap and its deletion mutants was examined in two cell lines common ...200211807766
epstein-barr virus balf1 is a bcl-2-like antagonist of the herpesvirus antiapoptotic bcl-2 proteins.cellular bcl-2 family proteins can inhibit or induce programmed cell death in part by counteracting the activity of other bcl-2 family members. all sequenced gammaherpesviruses encode a bcl-2 homologue that potently inhibits apoptosis and apparently escapes some of the regulatory mechanisms that govern the functions of their cellular counterparts. examples of these protective proteins include bhrf1 of epstein-barr virus (ebv) and ksbcl-2 of kaposi's sarcoma-associated herpesvirus, also known as ...200211836425
rapid activation of poly(adp-ribose) polymerase contributes to sindbis virus and staurosporine-induced apoptotic cell death.poly(adp-ribose) polymerase-1 (parp-1) is a chromatin-associated enzyme that is activated by dna strand breaks and catalyzes the transfer of adp-ribose groups from nad to itself and other nuclear proteins. although caspase-mediated parp-1 cleavage occurs during almost all forms of apoptosis, the contribution of parp-1 activation and cleavage to this cell death process remains unclear. using immortalized fibroblasts from wild-type (parp-1(+/+)) and parp-1 knockout (parp-1(-/-)) mice, and a mouse ...200211853409
expression and self-assembly of norwalk virus capsid protein from venezuelan equine encephalitis virus replicons.the norwalk virus (nv) capsid protein was expressed using venezuelan equine encephalitis virus replicon particles (vrp-nv1). vrp-nv1 infection resulted in large numbers of recombinant nv-like particles that were primarily cell associated and were indistinguishable from nv particles produced from baculoviruses. mutations located in the n-terminal and p1 domains of the nv capsid protein ablated capsid self-assembly in mammalian cells.200211861868
cancer immunotherapy using sindbis virus replicon particles encoding a vp22-antigen fusion.alphavirus vectors have emerged as a strategy for the development of cancer vaccines and gene therapy applications. the availability of a new packaging cell line (pcl), which is capable of generating alphavirus replicon particles without contamination from replication-competent virus, has advanced the field of vaccine development. this replication-defective vaccine vector has potential advantages over naked nucleic acid vaccines, such as increased efficiency of gene delivery and large-scale prod ...200211874633
natural killer t cell ligand alpha-galactosylceramide enhances protective immunity induced by malaria vaccines.the important role played by cd8(+) t lymphocytes in the control of parasitic and viral infections, as well as tumor development, has raised the need for the development of adjuvants capable of enhancing cell-mediated immunity. it is well established that protective immunity against liver stages of malaria parasites is primarily mediated by cd8(+) t cells in mice. activation of natural killer t (nkt) cells by the glycolipid ligand, alpha-galactosylceramide (alpha-galcer), causes bystander activa ...200211877484
restriction of a sindbis virus mutant in bhk cells and relief of the restriction by the addition of adenosine.sv(pzf) is a mutant of sindbis virus (sv) which we selected on the basis of its ability to replicate in mosquito cells treated with pyrazofurin (pzf), a drug which inhibits pyrimidine nucleotide biosynthesis (lin et al., 2000, virology 272, 61-71). three mutations, a6627u, a7543u, and c7593a, were identified in the nsp4 (the viral rna polymerase) coding region, which were required for the pzf-resistant phenotype. we report here that sv(pzf) has a second phenotype. its replication in bhk cells is ...200211878910
induction of bad-mediated apoptosis by sindbis virus infection: involvement of pro-survival members of the bcl-2 family.it is known that infection with sindbis virus (snv) induces apoptosis, which is inhibited by two pro-survival members of the bcl-2 family, bcl-2 and bcl-xl. however, the mechanism of involvement of the other members of the bcl-2 family in snv-induced apoptosis remains unclear. in this study we report that bad protein, one of the pro-apoptotic bcl-2 family members, mediates apoptosis in the mammalian cells infected with snv. expression of bad was shown to promote snv-induced apoptosis in human em ...200211878929
mapping the rubella virus subgenomic promoter.rubella virus (rub), the sole member of the rubivirus genus in the togaviridae family of positive-strand rna viruses, synthesizes a single subgenomic (sg) rna containing sequences from the 3' end of the genomic rna including the open reading frame (orf) that encodes the virion proteins. the synthesis of sg rna is initiated internally on a negative-strand, genome-length template at a site known as the sg promoter (sgp). mapping the rub sgp was initiated by using an infectious cdna vector, dsrobo4 ...200211884543
coxsackievirus b3 replication is reduced by inhibition of the extracellular signal-regulated kinase (erk) signaling pathway.coxsackievirus b3 (cvb3) is the most common human pathogen for viral myocarditis. we have previously shown that the signaling protein p21(ras) gtpase-activating protein (rasgap) is cleaved and that mitogen-activated protein kinases (mapks) erk1/2 are activated in the late phase of cvb3 infection. however, the role of intracellular signaling pathways in cvb3-mediated myocarditis and the relative advantages of such pathways to host or virus remain largely unclear. in this study we extended our pri ...200211884562
glycosylation of the hepatitis c virus envelope protein e1 occurs posttranslationally in a mannosylphosphoryldolichol-deficient cho mutant cell line.the addition of n-linked glycans to a protein is catalyzed by oligosaccharyltransferase, an enzyme closely associated with the translocon. n-glycans are believed to be transferred as the protein is being synthesized and cotranslationally translocated in the lumen of the endoplasmic reticulum. we used a mannosylphosphoryldolichol-deficient chinese hamster ovary mutant cell line (b3f7 cells) to study the temporal regulation of n-linked core glycosylation of hepatitis c virus envelope protein e1. i ...200211886842
antiviral effects of an iminosugar derivative on flavivirus infections.endoplasmic reticulum (er) alpha-glucosidase inhibitors, which block the trimming step of n-linked glycosylation, have been shown to eliminate the production of several er-budding viruses. here we investigated the effects of one such inhibitor, n-nonyl-deoxynojirimycin (nn-dnj), a 9-carbon alkyl iminosugar derivative, on infection by japanese encephalitis virus (jev) and dengue virus serotype 2 (den-2). in the presence of nn-dnj, jev and den-2 infections were suppressed in a dose-dependent manne ...200211907199
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