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efficient gene delivery into adult cardiomyocytes by recombinant sindbis virus.somatic gene therapy as a potential strategy for the treatment of myocardial diseases relies on an efficient gene transfer into cardiac muscle cells. the difficulty of delivering genes into adult cardiomyocytes exists not only in vivo but also in primary culture systems. therefore, possibilities for ex vivo gene transfer and the in vitro study of physiological processes by reverse genetics are limited. we investigated the potential of an alphavirus-based vector system to transduce adult rat card ...199910682322
alternative splicing results in differential expression, activity, and localization of the two forms of arginyl-trna-protein transferase, a component 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. the underlying ubiquitin-dependent proteolytic system, called the n-end rule pathway, is organized hierarchically: n-terminal aspartate and glutamate (and also cysteine in metazoans) are secondary destabilizing residues, in that they function through their conjugation, by arginyl-trna-protein transferase (r-transferase), to arginine, a primary destabilizing residue. we isolated cdna encoding the ...19999858543
evaluation of recombinant dense granule antigen 7 (gra7) of toxoplasma gondii for detection of immunoglobulin g antibodies and analysis of a major antigenic domain.dense granule protein 7 (gra7) of toxoplasma gondii was expressed in escherichia coli as a fusion protein. the leader peptide contained a 25-amino-acid mouse tumor necrosis factor fragment and six histidyl residues. after purification by metal chelate affinity chromatography, the antigen was evaluated in an enzyme-linked immunosorbent assay for detection of immunoglobulin g (igg). for two sets of igg-positive human serum samples, obtained from routine screening, an overall sensitivity of 81% was ...19999874659
use of vegetable-derived tween 80 for virus inactivation by solvent/detergent treatment. 199910652181
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
[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
active residues and viral substrate cleavage sites of the protease of the birnavirus infectious pancreatic necrosis virus.the polyprotein of infectious pancreatic necrosis virus (ipnv), a birnavirus, is processed by the viral protease vp4 (also named ns) to generate three polypeptides: pvp2, vp4, and vp3. site-directed mutagenesis at 42 positions of the ipnv vp4 protein was performed to determine the active site and the important residues for the protease activity. two residues (serine 633 and lysine 674) were critical for cleavage activity at both the pvp2-vp4 and the vp4-vp3 junctions. wild-type activity at the p ...200010666235
the in vitro-synthesized rna from a cdna clone of hepatitis e virus is infectious.hepatitis e virus (hev) is an important etiological agent of epidemic and sporadic hepatitis, which is endemic to the indian subcontinent and prevalent in most of the developing parts of the world. the infection is often associated with acute liver failure and high mortality, particularly in pregnant women. in order to develop methods of intervention, it is essential to understand the biology of the virus. this is particularly important as no reliable in vitro culture system is available. we hav ...200010666275
pseudotyping of glycoprotein d-deficient herpes simplex virus type 1 with vesicular stomatitis virus glycoprotein g enables mutant virus attachment and entry.the use of herpes simplex virus (hsv) vectors for in vivo gene therapy will require the targeting of vector infection to specific cell types in certain in vivo applications. because hsv glycoprotein d (gd) imparts a broad host range for viral infection through recognition of ubiquitous host cell receptors, vector targeting will require the manipulation of gd to provide new cell recognition specificities in a manner designed to preserve gd's essential role in virus entry. in this study, we have d ...200010666285
development of a sindbis virus expression system that efficiently expresses green fluorescent protein in midguts of aedes aegypti following per os infection.a double subgenomic sindbis (dssin) virus, mre/3'2 j/gfp, was constructed to efficiently express green fluorescent protein (gfp) in the midgut of aedes aegypti following per os infection. the mre/3'2 j/gfp rna genome contained the nonstructural genes and cis-acting sequences of the dssin virus, te/3'2 j/gfp, but had the structural genes of mre16 sin virus. mre/3'2 j/gfp virus, unlike te/3'2 j/gfp virus, efficiently infected mosquitoes orally. at 1-2 days postinfection, gfp was observed as multip ...200010672072
the promise of nucleic acid vaccines.establishing the effective use of 'naked' nucleic acids as vaccines would undoubtedly be one of the most important advances in the history of vaccinology. while nucleic acids show much promise for use as vaccine vectors in experimental animals, not a single naked nucleic acid vector has been approved for use in humans. indeed, data from human clinical trials is scant: nucleic acid vaccines have not been clearly demonstrated to have any convincing efficacy in the prevention or treatment of infect ...200010673713
imaging high-resolution structure of gfp-expressing neurons in neocortex in vivo.to detect subtle changes in neuronal morphology in response to changes in experience, one must image neurons at high resolution in vivo over time scales of minutes to days. we accomplished this by infecting postmitotic neurons in rat and mouse barrel cortex with a sindbis virus carrying the gene for enhanced green fluorescent protein. visualized with 2-photon excitation laser scanning microscopy, infected neurons showed bright fluorescence that was distributed homogeneously throughout the cell, ...200011112802
chronic arthritis after sindbis-related (pogosta) virus infection. 200011132212
a mutant allele of essential, general translation initiation factor ded1 selectively inhibits translation of a viral mrna.positive-strand rna virus genomes are substrates for translation, rna replication, and encapsidation. to identify host factors involved in these functions, we used the ability of brome mosaic virus (bmv) rna to replicate in yeast. we report herein identification of a mutation in the essential yeast gene ded1 that inhibited bmv rna replication but not yeast growth. ded1 encodes a dead (asp-glu-ala-asp)-box rna helicase required for translation initiation of all yeast mrnas. inhibition of bmv rna ...200011069307
the host range phenotype displayed by a sindbis virus glycoprotein variant results from virion aggregation and retention on the surface of mosquito cells.the sindbis virus variant ne2g216 is a pe2-containing host range mutant that is growth restricted in cultured mosquito cells (c6/36) due to inefficient release of virions from this cell type. the maturation defect of ne2g216 has been linked to the structures of n-linked oligosaccharides synthesized by arthropod cells. analysis of c6/36 cells infected with ne2g216 by transmission electron microscopy revealed the presence of dense virus aggregates within cytoplasmic vacuoles and virus aggregates a ...200011070041
survival motor neuron protein modulates neuron-specific apoptosis.spinal muscular atrophy (sma) is attributed to mutations in the smn1 gene, leading to loss of spinal cord motor neurons. the neurotropic sindbis virus vector system was used to investigate a role for the survival motor neuron (smn) protein in regulating neuronal apoptosis. here we show that smn protects primary neurons and differentiated neuron-like stem cells, but not cultured cell lines from virus-induced apoptotic death. smn also protects neurons in vivo and increases survival of virus-infect ...200011078511
broad antiviral activity in tissues of crustaceans.innate antiviral substances occur in vertebrates and may function as host defenses. virus infections are common among invertebrates, but little is known about the ability of invertebrates to control viral infections. pre-existing antiviral substances may be particularly important, since invertebrates lack the antiviral defense conferred by specific immunity. in our study, we found that tissue extracts of blue crab (callinectes sapidus), shrimp (penaeus setiferus), and crayfish (procambarus clark ...200011080539
correlation of susceptibility of immature mice to fungal infection (blastomycosis) and effector cell function.immature mice are highly susceptible to blastomycosis, which is similar to other mycoses and has parallels in humans. the murine susceptibility is noteworthy in that it persists beyond the development of resistance to other, nonfungal pathogens and the maturation of most immune functions. as the susceptibility to blastomycosis appeared to be related to an early event after infection, primary effector cell function was studied. we found that peritoneal inflammatory cells, enriched for neutrophils ...200011083802
a universal 'one-tube' rt-pcr protocol for amplifying isolates of bovine viral diarrhoea virus.the increase in the knowledge of the genetic variability of bvdv and the identification of some of the genetic determinants of its pathogenicity require robust and practical tools for rapid molecular characterization of the various genotypes of this virus. this study was undertaken to develop a standard protocol for rt-pcr that allows the amplification of various parts of the genome of bvdv without the need for optimizing each individual reaction. the reaction set-up is very flexible because it ...200011085469
prolonged arthritis associated with sindbis-related (pogosta) virus infection.a follow-up study of musculoskeletal symptoms after pogosta virus infection.200011085809
interaction between complement receptor gc1qr and hepatitis c virus core protein inhibits t-lymphocyte proliferation.hepatitis c virus (hcv) is an important human pathogen that is remarkably efficient at establishing persistent infection. the hcv core protein is the first protein expressed during the early phase of hcv infection. our previous work demonstrated that the hcv core protein suppresses host immune responses, including anti-viral cytotoxic t-lymphocyte responses in a murine model. to investigate the mechanism of hcv core-mediated immunosuppression, we searched for host proteins capable of associating ...200011086025
enhancing b- and t-cell immune response to a hepatitis c virus e2 dna vaccine by intramuscular electrical gene transfer.we describe an improved genetic immunization strategy for eliciting a full spectrum of anti-hepatitis c virus (hcv) envelope 2 (e2) glycoprotein responses in mammals through electrical gene transfer (egt) of plasmid dna into muscle fibers. intramuscular injection of a plasmid encoding a cross-reactive hypervariable region 1 (hvr1) peptide mimic fused at the n terminus of the e2 ectodomain, followed by electrical stimulation treatment in the form of high-frequency, low-voltage electric pulses, in ...200011090158
infection of human dendritic cells by a sindbis virus replicon vector is determined by a single amino acid substitution in the e2 glycoprotein.the ability to target antigen-presenting cells with vectors encoding desired antigens holds the promise of potent prophylactic and therapeutic vaccines for infectious diseases and cancer. toward this goal, we derived variants of the prototype alphavirus, sindbis virus (sin), with differential abilities to infect human dendritic cells. cloning and sequencing of the sin variant genomes revealed that the genetic determinant for human dendritic cell (dc) tropism mapped to a single amino acid substit ...200011090185
efficient enhancement of priming effect by intermittent treatment with interferon.to improve the efficacy of interferon (ifn) therapy for chronic hepatitis c, we proposed a therapy with twice-a-day injection of ifnbeta as the induction. to assess its biological enhancement, we compared antiviral activities in vitro using intermittent treatment schedules simulating the clinical condition. fl cells were treated with ifnbeta twice in 12 h interval (single treatment, 1000 and 0 iu/ml; double treatment, 500 iu/ml each) and challenged with sindbis virus. antiviral activities were d ...200011095955
virus-expressed, recombinant single-chain antibody blocks sporozoite infection of salivary glands in plasmodium gallinaceum-infected aedes aegypti.transgenic mosquitoes resistant to malaria parasites are being developed to test the hypothesis that they may be used to control disease transmission. we have developed an effector portion of an antiparasite gene that can be used to test malaria resistance in transgenic mosquitoes. mouse monoclonal antibodies that recognize the circumsporozoite protein of plasmodium gallinaceum can block sporozoite invasion of aedes aegypti salivary glands. an anti-circumsporozoite monoclonal antibody, n2h6d5, w ...200011220756
adaptive mutations in sindbis virus e2 and ross river virus e1 that allow efficient budding of chimeric viruses.alphavirus glycoproteins e2 and e1 form a heterodimer that is required for virus assembly. we have studied adaptive mutations in e2 of sindbis virus (sin) and e1 of ross river virus (rr) that allow these two glycoproteins to interact more efficiently in a chimeric virus that has sin e2 but rr e1. these mutations include k129e, k131e, and v237f in sin e2 and s310f and c433r in rr e1. although rr e1 and sin e2 will form a chimeric heterodimer, the chimeric virus is almost nonviable, producing abou ...200010684281
genetics of mosquito vector competence.mosquito-borne diseases are responsible for significant human morbidity and mortality throughout the world. efforts to control mosquito-borne diseases have been impeded, in part, by the development of drug-resistant parasites, insecticide-resistant mosquitoes, and environmental concerns over the application of insecticides. therefore, there is a need to develop novel disease control strategies that can complement or replace existing control methods. one such strategy is to generate pathogen-resi ...200010704476
functional correlation of gaba(a) receptor alpha subunits expression with the properties of ipscs in the developing thalamus.gaba(a) receptor alpha1 and alpha2 subunits are expressed differentially with ontogenic period in the brain, but their functional roles are not known. we have recorded gaba(a) receptor-mediated ipscs from laterodorsal (ld) thalamic relay neurons in slices of rat brain at various postnatal ages and found that decay times of evoked ipscs and spontaneous miniature ipscs undergo progressive shortening during the first postnatal month. with a similar time course, expression of transcripts and protein ...200010704495
human immunodeficiency virus type 1 vpr induces apoptosis through caspase activation.human immunodeficiency virus type 1 (hiv-1) vpr is a 96-amino-acid protein that is found associated with the hiv-1 virion. vpr induces cell cycle arrest at the g(2)/m phase of the cell cycle, and this arrest is followed by apoptosis. we examined the mechanism of vpr-induced apoptosis and found that hiv-1 vpr-induced apoptosis requires the activation of a number of cellular cysteinyl aspartate-specific proteases (caspases). we demonstrate that ectopic expression of anti-apoptotic viral proteins, ...200010708425
evolutionarily related sindbis-like plant viruses maintain different levels of population diversity in a common host.the levels of population diversity of three related sindbis-like plant viruses, tobacco mosaic virus (tmv), cucumber mosaic virus (cmv), and cowpea chlorotic mottle virus (ccmv), in infections of a common host, nicotiana benthamiana, established from genetically identical viral rna were examined. despite probably having a common evolutionary ancestor, the three viruses maintained different levels of population diversity. cmv had the highest levels of diversity, tmv had an intermediate level of d ...200010708428
isolation and characterization of an arterivirus defective interfering rna genome.equine arteritis virus (eav), the type member of the family arteriviridae, is a single-stranded rna virus with a positive-stranded genome of approximately 13 kb. eav uses a discontinuous transcription mechanism to produce a nested set of six subgenomic mrnas from which its structural genes are expressed. we have generated the first documented arterivirus defective interfering (di) rnas by serial undiluted passaging of a wild-type eav stock in bhk-21 cells. a cdna copy of the smallest di rna (5.6 ...200010708432
cis- and trans-acting elements in flavivirus rna replication.most of the seven flavivirus nonstructural proteins (ns1 to ns5) encoded in the distal two-thirds of the rna positive-sense genome are believed to be essential components of rna replication complexes. to explore the functional relationships of these components in rna replication, we used trans-complementation analysis of full-length infectious rnas of kunjin (kun) virus with a range of lethal in-frame deletions in the nonstructural coding region, using as helper a repbhk cell line stably produci ...200010708442
rotavirus spike protein vp4 is present at the plasma membrane and is associated with microtubules in infected cells.vp4 is an unglycosylated protein of the outer layer of the capsid of rotavirus. it forms spikes that project from the outer layer of mature virions, which is mainly constituted by glycoprotein vp7. vp4 has been implicated in several important functions, such as cell attachment, penetration, hemagglutination, neutralization, virulence, and host range. previous studies indicated that vp4 is located in the space between the periphery of the viroplasm and the outside of the endoplasmic reticulum in ...200010708448
alpha/beta interferon protects adult mice from fatal sindbis virus infection and is an important determinant of cell and tissue tropism.infection of adult 129 sv/ev mice with consensus sindbis virus strain tr339 is subclinical due to an inherent restriction in early virus replication and viremic dissemination. by comparing the pathogenesis of tr339 in 129 sv/ev mice and alpha/beta interferon receptor null (ifn-alpha/betar(-/-)) mice, we have assessed the contribution of ifn-alpha/beta in restricting virus replication and spread and in determining cell and tissue tropism. in adult 129 sv/ev mice, subcutaneous inoculation with 100 ...200010708454
[comparative study of the reproduction of sidnbis virus sensitive and resistant to adamantane derivatives]. 200010710638
inhibitor binding induces active site stabilization of the hcv ns3 protein serine protease domain.few structures of viral serine proteases, those encoded by the sindbis and semliki forest viruses, hepatitis c virus (hcv) and cytomegalovirus, have been reported. in the life cycle of hcv a crucial role is played by a chymotrypsin-like serine protease encoded at the n-terminus of the viral ns3 protein, the solution structure of which we present here complexed with a covalently bound reversible inhibitor. unexpectedly, the residue in the p2 position of the inhibitor induces an effective stabiliz ...200010716920
morphological features of murray valley encephalitis virus infection in the central nervous system of swiss mice.we have examined the histological and ultrastructural features of cns infection with murray valley encephalitis (mve) virus in mice inoculated with a virulent parental strain (bh3479). light microscopic examination revealed neuronal necrosis in the olfactory bulb and hippocampus of mve-infected brains by 5 days post-infection (pi). electron microscopy of these regions showed endoplasmic reticulum membrane proliferation, and tubular and spherical structures in the cisternae of the endoplasmic ret ...200010718862
broad-spectrum antiviral activity of the imp dehydrogenase inhibitor vx-497: a comparison with ribavirin and demonstration of antiviral additivity with alpha interferon.the enzyme imp dehydrogenase (impdh) catalyzes an essential step in the de novo biosynthesis of guanine nucleotides, namely, the conversion of imp to xmp. the major event occurring in cells exposed to competitive impdh inhibitors such as ribavirin or uncompetitive inhibitors such as mycophenolic acid (mpa) is a depletion of the intracellular gtp and dgtp pools. ribavirin is approved as an inhaled antiviral agent for treatment of respiratory syncytial virus (rsv) infection and orally, in combinat ...200010722482
specific phosphorylated forms of glyceraldehyde 3-phosphate dehydrogenase associate with human parainfluenza virus type 3 and inhibit viral transcription in vitro.we previously reported specific interaction of cellular glyceraldehyde 3-phosphate dehydrogenase (gapdh), the key glycolytic enzyme, and la protein, the rna polymerase iii transcription factor, with the cis-acting rnas of human parainfluenza virus type 3 (hpiv3) and packaging of these proteins within purified virions (b. p. de, s. gupta, h. zhao, j. z. drazba, and a. k. banerjee, j. biol. chem. 271:24728-24735, 1996). to gain further insight into these molecular interactions, we analyzed the vir ...200010729139
control of sindbis virus infection by antibody in interferon-deficient mice.antibodies clear sindbis virus from infected animals through an unknown mechanism. to determine whether interferon-induced pathways are required for this clearance, we examined mice which are unable to respond to alpha/beta interferon or gamma interferon. although extremely susceptible to infection, such mice survived and completely cleared virus if antibodies against sindbis virus were given.200010729167
naturally occurring tap-dependent specific t-cell tolerance for a variant of an immunodominant retroviral cytotoxic t-lymphocyte epitope.upon immunization and restimulation with tumors induced by the endogenous akr/gross murine leukemia virus (mulv), c57bl/6 mice generate vigorous h-2k(b)-restricted cytotoxic t-lymphocyte (ctl) responses to a determinant (kspwfttl) derived from the p15e transmembrane portion of the viral envelope glycoprotein. by contrast, the highly homologous determinant rspwfttl, expressed by tumor cells induced by friend/moloney/rauscher (fmr) mulv, is not immunogenic, even when presented to the immune system ...200010729170
development of reverse transcription-pcr assays specific for detection of equine encephalitis viruses.specific and sensitive reverse transcription-pcr (rt-pcr) assays were developed for the detection of eastern, western, and venezuelan equine encephalitis viruses (eee, wee, and vee, respectively). tests for specificity included all known alphavirus species. the eee-specific rt-pcr amplified a 464-bp region of the e2 gene exclusively from 10 different eee strains from south and north america with a sensitivity of about 3,000 rna molecules. in a subsequent nested pcr, the specificity was confirmed ...200010747138
identification of a novel function of the alphavirus capping apparatus. rna 5'-triphosphatase activity of nsp2.both genomic and subgenomic rnas of the alphavirus have m(7)g(5')ppp(5')n (cap0 structure) at their 5' end. previously it has been shown that alphavirus-specific nonstructural protein nsp1 has guanine-7n-methyltransferase and guanylyltransferase activities needed in the synthesis of the cap structure. during normal cap synthesis the 5' gamma-phosphate of the nascent viral rna chain is removed by a specific rna 5'-triphosphatase before condensation with gmp, delivered by the guanylyltransferase. ...200010748213
clinical isolates of measles virus use cd46 as a cellular receptor.laboratory strains of measles viruses (mv), such as edmonston and halle, use the complement regulatory protein cd46 as a cell surface receptor. the receptor usage of clinical isolates of mv, however, remains unclear. receptor usage by primary patient isolates of mv was compared to isolates that had been passaged on a variety of tissue culture cell lines. all of the isolates could infect cells in a cd46-dependent manner, but their tropism was restricted according to cell type (e.g., lymphocytes v ...200010756008
subgenomic negative-strand rna function during mouse hepatitis virus infection.mouse hepatitis virus (mhv)-infected cells contain full-length and subgenomic-length positive- and negative-strand rnas. the origin and function of the subgenomic negative-strand rnas is controversial. in this report we demonstrate that the synthesis and molar ratios of subgenomic negative strands are similar in alternative host cells, suggesting that these rnas function as important mediators of positive-strand synthesis. using kinetic labeling experiments, we show that the full-length and subg ...200010756015
a single amino acid change in nsp1 attenuates neurovirulence of the sindbis-group alphavirus s.a.ar86.s.a.ar86, a member of the sindbis group of alphaviruses, is neurovirulent in adult mice and has a unique threonine at position 538 of nsp1; nonneurovirulent members of this group of alphaviruses encode isoleucine. isoleucine was introduced at position 538 in the wild-type s.a.ar86 infectious clone, ps55, and virus derived from this mutant clone, ps51, was significantly attenuated for neurovirulence compared to that derived from ps55. intracranial (i.c. ) s55 infection resulted in severe disease, ...200010756033
frequent homologous recombination events between molecules of one rna component in a multipartite rna virus.brome mosaic bromovirus (bmv), a tripartite plus-sense rna virus, has been used as a model system to study homologous rna recombination among molecules of the same rna component. pairs of bmv rna3 variants carrying marker mutations at different locations were coinoculated on a local lesion host, and the progeny rna3 in a large number of lesions was analyzed. the majority of doubly infected lesions accumulated the rna3 recombinants. the distribution of the recombinant types was relatively even, i ...200010756034
a single deletion in the membrane-proximal region of the sindbis virus glycoprotein e2 endodomain blocks virus assembly.the envelopment of the sindbis virus nucleocapsid in the modified cell plasma membrane involves a highly specific interaction between the capsid (c) protein and the endodomain of the e2 glycoprotein. we have previously identified a domain of the sindbis virus c protein involved in binding to the e2 endodomain (h. lee and d. t. brown, virology 202:390-400, 1994). the c-e2 binding domain resides in a hydrophobic cleft with c y180 and w247 on opposing sides of the cleft. structural modeling studies ...200010756035
the use of chimeric venezuelan equine encephalitis viruses as an approach for the molecular identification of natural virulence determinants.venezuelan equine encephalitis (vee) virus antigenic subtypes and varieties are considered either epidemic/epizootic or enzootic. in addition to epidemiological differences between the epidemic and enzootic viruses, several in vitro and in vivo laboratory markers distinguishing the viruses have been identified, including differential plaque size, sensitivity to interferon (ifn), and virulence for guinea pigs. these observations have been shown to be useful predictors of natural, equine virulence ...200010756040
nucleic acid-dependent cross-linking of the nucleocapsid protein of sindbis virus.the assembly of the alphavirus nucleocapsid core is a multistep event requiring the association of the nucleocapsid protein with nucleic acid and the subsequent oligomerization of capsid proteins into an assembled core particle. although the mechanism of assembly has been investigated extensively both in vivo and in vitro, no intermediates in the core assembly pathway have been identified. through the use of both truncated and mutant sindbis virus nucleocapsid proteins and a variety of cross-lin ...200010756045
stable high-level expression of heterologous genes in vitro and in vivo by noncytopathic dna-based kunjin virus replicon vectors.primary features of the flavivirus kunjin (kun) subgenomic replicons include continuous noncytopathic replication in host cell cytoplasm and the ability to be encapsidated into secreted virus-like particles (vlps). previously we reported preparation of rna-based kun replicon vectors and expression of heterologous genes (hg) in cell culture after rna transfection or after infection with recombinant kun vlps (a. n. varnavski and a. a. khromykh, virology 255:366-375, 1999). in this study we describ ...200010756054
evaluation of recombinant alphaviruses as vectors in gene therapy.alphavirus vectors based on sindbis virus and semliki forest virus (sfv) were characterized as potential gene transfer vectors. initial studies were performed using vectors engineered to transfer either lacz or green fluorescent protein (gfp). high levels of gene transfer were achieved in human primary fibroblasts, bhk and 293t cells, with low levels of transduction observed in more than 20 other target cells. alphavirus-based expression was generally very high, but transient in every cell type. ...200010757020
antiviral effect of nitric oxide during japanese encephalitis virus infection.the ability of japanese encephalitis virus (jev) and jev-induced macrophage derived neutrophil chemotactic factor (mdf) to produce nitric oxide (no), and the possible antiviral effect of no during jev infection, was investigated. splenic macrophages of jev infected mice produced maximum no in vivo at day 7 post infection, and in vitro at 24 h after jev stimulation. mdf-induced no production was dose dependent and maximal at 60 min after mdf treatment. the response was sensitive to anti-mdf antib ...200010762444
isolation and complete nucleotide sequence of a chinese sindbis-like virus.infection with alphaviruses is common in the chinese population. here we report the isolation of a sindbis-like virus from a pool of anopheles mosquitoes collected in xinjiang, china during an arbovirus survey. this virus, designated xj-160, rapidly produced cytopathic effects on mosquito and hamster cells. in addition, it was lethal to neonatal mice if inoculated intracerebrally. serologically, xj-160 reacted with and was neutralized by an anti-sindbis antibody. anti-xj-160 antibodies were foun ...200010769078
maintenance of cd8(+) t-cell memory following infection with recombinant sindbis and vaccinia viruses.cd8(+) t-cell memory is critical for protection against pathogens poorly controlled by humoral immunity. to characterize two distinct vaccine vectors, the acute and memory cd8(+) t-cell responses to an hiv-1 epitope (p18) expressed by recombinant vaccinia (vp18) and sindbis (sinp18) viruses were compared. whereas 9 to 13% of cd8(+) splenocytes were p18 specific during the acute response to vp18, 4% were induced by sinp18 as revealed by class i tetramer staining. increased t-cell activation by vp ...200010772979
replicase complex genes of semliki forest virus confer lethal neurovirulence.semliki forest virus (sfv) is a mosquito-transmitted pathogen of small rodents, and infection of adult mice with sfv4, a neurovirulent strain of sfv, leads to lethal encephalitis in a few days, whereas mice infected with the avirulent a7(74) strain remain asymptomatic. in adult neurons, a7(74) is unable to form virions and hence does not reach a critical threshold of neuronal damage. to elucidate the molecular mechanisms of neurovirulence, we have cloned and sequenced the entire 11,758-nucleotid ...200010775594
alkylation of cysteine-containing peptides to mimic palmitoylation.numerous proteins that are involved in cell signaling and viral replication require post-translational modification by palmitoylation to function properly. the molecular details by which this palmitoyl modification affects protein function remain poorly understood. to facilitate in vitro biochemical and structural studies of the role of palmitoylation on protein function, a method was developed for alkylating peptides with saturated c16 groups at cysteine residues and demonstrated using peptides ...200010784030
standardization of immunoglobulin m capture enzyme-linked immunosorbent assays for routine diagnosis of arboviral infections.immunoglobulin m antibody-capture enzyme-linked immunosorbent assay (mac-elisa) is a rapid and versatile diagnostic method that readily permits the combination of multiple assays. test consolidation is especially important for arthropod-borne viruses (arboviruses) which belong to at least three virus families: the togaviridae, flaviviridae, and bunyaviridae. using prototype viruses from each of these families and a panel of well-characterized human sera, we have evaluated and standardized a comb ...200010790107
modulation of dengue virus infection in human cells by alpha, beta, and gamma interferons.a role for interferon (ifn) in modulating infection by dengue virus (dv) has been suggested by studies in dv-infected patients and ifn receptor-deficient mice. to address how ifn modulates dv type 2 infection, we have assayed ifn-alpha, -beta, and -gamma for the ability to enhance or diminish antibody-independent and antibody-dependent cell infection using a competitive, asymmetric reverse transcriptase-mediated pcr (rt-pcr) assay that quantitates positive and negative strands of viral rna, a fl ...200010799569
assembly of the coronavirus envelope: homotypic interactions between the m proteins.the viral membrane proteins m and e are the minimal requirements for the budding of coronavirus particles. since the e protein occurs in particles only in trace amounts, the lateral interactions between the m proteins apparently generate the major driving force for envelope formation. by using coimmunoprecipitation and envelope incorporation assays, we provide extensive evidence for the existence of such m-m interactions. in addition, we determined which domains of the m protein are involved in ...200010799570
antiapoptotic herpesvirus bcl-2 homologs escape caspase-mediated conversion to proapoptotic proteins.the antiapoptotic bcl-2 and bcl-x(l) proteins of mammals are converted into potent proapoptotic factors when they are cleaved by caspases, a family of apoptosis-inducing proteases (e. h.-y. cheng, d. g. kirsch, r. j. clem, r. ravi, m. b. kastan, a. bedi, k. ueno, and j. m. hardwick, science 278:1966-1968, 1997; r. j. clem, e. h.-y. cheng, c. l. karp, d. g. kirsch, k. ueno, a. takahashi, m. b. kastan, d. e. griffin, w. c. earnshaw, m. a. veliuona, and j. m. hardwick, proc. natl. acad. sci. usa 95 ...200010799576
host protein interactions with the 3' end of bovine coronavirus rna and the requirement of the poly(a) tail for coronavirus defective genome replication.rna viruses have 5' and 3' untranslated regions (utrs) that contain specific signals for rna synthesis. the coronavirus genome is capped at the 5' end and has a 3' utr that consists of 300 to 500 nucleotides (nt) plus a poly(a) tail. to further our understanding of coronavirus replication, we have begun to examine the involvement of host factors in this process for two group ii viruses, bovine coronavirus (bcv) and mouse hepatitis coronavirus (mhv). specific host protein interactions with the bc ...200010799579
cell fusion activity of hepatitis c virus envelope proteins.to examine the cell fusion activity of hepatitis c virus (hcv) envelope proteins (e1 and e2), we have established a sensitive cell fusion assay based on the activation of a reporter gene as described previously (o. nussbaum, c. c. broder, and e. a. berger, j. virol. 68:5411-5422, 1994). the chimeric hcv e1 and e2 proteins, each consisting of the ectodomain of the e1 and e2 envelope protein and the transmembrane and cytoplasmic domains of the vesicular stomatitis virus g glycoprotein, were expres ...200010799580
mov34 protein from mouse brain interacts with the 3' noncoding region of japanese encephalitis virus.the plus-sense rna genome of japanese encephalitis virus (jev) contains noncoding regions (ncrs) of 95 and 585 bases at its 5' and 3' ends, respectively. the last 83 nucleotides of the 3'-ncr are predicted to form stable stem-loop (sl) structures. the shape of this 3'-sl structure is highly conserved among divergent flaviviruses even though only small stretches of nucleotide sequence contained within these structures are conserved. these sl structures have been predicted to function as cis-actin ...200010799585
mutational analysis of the rubella virus nonstructural polyprotein and its cleavage products in virus replication and rna synthesis.rubella virus nonstructural proteins, translated from input genomic rna as a p200 polyprotein and subsequently processed into p150 and p90 by an intrinsic papain-like thiol protease, are responsible for virus replication. to examine the effect of p200 processing on virus replication and to study the roles of nonstructural proteins in viral rna synthesis, we introduced into a rubella virus infectious cdna clone a panel of mutations that had variable defective effects on p200 processing. the virus ...200010799588
activation of divergent neuronal cell death pathways in different target cell populations during neuroadapted sindbis virus infection of mice.infection of adult mice with neuroadapted sindbis virus (nsv) results in a severe encephalomyelitis accompanied by prominent hindlimb paralysis. we find that the onset of paralysis parallels morphologic changes in motor neuron cell bodies in the lumbar spinal cord and in motor neuron axons in ventral nerve roots, many of which are eventually lost over time. however, unlike nsv-induced neuronal cell death found in the brain of infected animals, the loss of motor neurons does not appear to be apop ...200010799613
altered activity, social behavior, and spatial memory in mice lacking the ntan1p amidase and the asparagine branch 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. n-terminal asparagine and glutamine are tertiary destabilizing residues, in that they are enzymatically deamidated to yield secondary destabilizing residues aspartate and glutamate, which are conjugated to arginine, a primary destabilizing residue. n-terminal arginine of a substrate protein is bound by the ubr1-encoded e3alpha, the e3 component of the ubiquitin-proteasome-dependent n-end rule pat ...200010805755
a novel membrane-anchored rab5 interacting protein required for homotypic endosome fusion.the ras-related gtpase rab5 is rate-limiting for homotypic early endosome fusion. we used a yeast two-hybrid screen to identify a rab5 interacting protein, rab5ip. the cdna sequence encodes a ubiquitous 75-kda protein with an n-terminal transmembrane domain (tm), a central coiled-coil structure, and a c-terminal region homologous to several centrosome-associated proteins. rab5ip lacking the transmembrane domain (rab5iptm(-)) had a greater affinity in vitro for rab5-guanosine 5'-o-2-(thio)diphosp ...200010818110
rubella virus nonstructural protein protease domains involved in trans- and cis-cleavage activities.rubella virus (rv) genomic rna contains two large open reading frames (orfs): a 5'-proximal orf encoding nonstructural proteins (nsps) that function primarily in viral rna replication and a 3'-proximal orf encoding the viral structural proteins. proteolytic processing of the rv nsp orf translation product p200 is essential for viral replication. processing of p200 to two mature products (p150 and p90) in the order nh(2)-p150-p90-cooh is carried out by an rv-encoded protease residing in the c-ter ...200010823845
the surface conformation of sindbis virus glycoproteins e1 and e2 at neutral and low ph, as determined by mass spectrometry-based mapping.sindbis virus contains two membrane glycoproteins, e1 and e2, which are organized into 80 trimers of heterodimers (spikes). these trimers form a precise t=4 icosahedral protein lattice on the surface of the virus. very little is known about the organization of the e1 and e2 glycoproteins within the spike trimer. to gain a better understanding of how the proteins e1 and e2 are arranged in the virus membrane, we have used the techniques of limited proteolysis and amino acid chemical modification i ...200010823875
characterization of the zinc binding activity of the rubella virus nonstructural protease.the rubella virus (rub) nonstructural (ns) protein (nsp) orf encodes a protease that cleaves the nsp precursor (240 kda) at a single site to produce two products. a cleavage site mutation was introduced into a rub infectious cdna clone and found to be lethal, demonstrating that cleavage of the nsp precursor is necessary for rub replication. based on computer alignments, the rub ns protease was predicted to be a papain-like cysteine protease (pcp) with the residues cys1152 and his1273 as the cata ...200010846076
a positive-strand rna virus with three very different subgenomic rna promoters.numerous rna viruses generate subgenomic mrnas (sgrnas) for expression of their 3'-proximal genes. a major step in control of viral gene expression is the regulation of sgrna synthesis by specific promoter elements. we used barley yellow dwarf virus (bydv) as a model system to study transcriptional control in a virus with multiple sgrnas. bydv generates three sgrnas during infection. the sgrna1 promoter has been mapped previously to a 98-nucleotide (nt) region which forms two stem-loop structure ...200010846080
the role of cd8(+) t cells and major histocompatibility complex class i expression in the central nervous system of mice infected with neurovirulent sindbis virus.little is known about the role of cd8(+) t cells infiltrating the neural parenchyma during encephalitis induced by neurovirulent sindbis virus (nsv). nsv preferentially infects neurons in the mouse brain and spinal cord; however, it is generally accepted that neurons can express few if any major histocompatibility complex (mhc) class i molecules. we evaluated the possible roles and interactions of cd8(+) t cells during nsv encephalitis and demonstrated that mhc class i antigen (h2k/d) was expres ...200010846095
differences between c57bl/6 and balb/cby mice in mortality and virus replication after intranasal infection with neuroadapted sindbis virus.neuroadapted sindbis virus (nsv), given intranasally, caused fatal encephalitis in 100% of adult c57bl/6 mice and 0% of balb/cby mice. most c57bl/6 mice developed severe kyphoscoliosis followed by hind-limb paralysis, while balb/cby mice did not. in situ hybridization for detecting nsv rna and immunohistochemistry for detecting nsv antigen indicated that virus delivered by this route infected neurons of the olfactory region and spread caudally without infection of ependymal cells. virus antigen ...200010846099
generation and characterization of a hepatitis c virus ns3 protease-dependent bovine viral diarrhea virus.unique to pestiviruses, the n-terminal protein encoded by the bovine viral diarrhea virus (bvdv) genome is a cysteine protease (npro) responsible for a self-cleavage that releases the n terminus of the core protein (c). this unique protease is dispensable for viral replication, and its coding region can be replaced by a ubiquitin gene directly fused in frame to the core. to develop an antiviral assay that allows the assessment of anti-hepatitis c virus (hcv) ns3 protease inhibitors, a chimeric b ...200010864644
selective membrane permeabilization by the rotavirus vp5* protein is abrogated by mutations in an internal hydrophobic domain.rotavirus infectivity is dependent on the proteolytic cleavage of the vp4 spike protein into vp8* and vp5* proteins. proteolytically activated virus, as well as expressed vp5*, permeabilizes membranes, suggesting that cleavage exposes a membrane-interactive domain of vp5* which effects rapid viral entry. the vp5* protein contains a single long hydrophobic domain (vp5*-hd, residues 385 to 404) at an internal site. in order to address the role of the vp5*-hd in permeabilizing cellular membranes, w ...200010864647
glycosphingolipids promote entry of a broad range of human immunodeficiency virus type 1 isolates into cell lines expressing cd4, cxcr4, and/or ccr5.treatment of human osteosarcoma cells, expressing cd4 and various chemokine receptors, with the glucosylceramide synthase inhibitor 1-phenyl-2-hexadecanoylamino-3-morpholino-1-propanol (ppmp), blocked target membrane glycosphingolipid (gsl) biosynthesis and reduced the susceptibility of cells to infection and fusion mediated by envelope glycoproteins from a variety of human immunodeficiency virus type 1 (hiv-1) isolates that utilize cxcr4 and/or ccr5. ppmp treatment of the cell lines did not sig ...200010864648
sindbis virus entry into cells triggers apoptosis by activating sphingomyelinase, leading to the release of ceramide.sindbis virus (sv) causes acute encephalomyelitis by infecting and inducing the death of neurons. induction of apoptosis occurs during virus entry and involves acid-induced conformational changes in the viral surface glycoproteins and sphingomyelin (sm)-dependent fusion of the virus envelope with the endosomal membrane. we have studied neuroblastoma cells to determine how this entry process triggers cell death. acidic sphingomyelinase was activated during entry followed by activation of neutral ...200010864654
the fusion glycoprotein of human respiratory syncytial virus facilitates virus attachment and infectivity via an interaction with cellular heparan sulfate.human respiratory syncytial virus (rsv) f glycoprotein (rsv-f) can independently interact with immobilized heparin and facilitate both attachment to and infection of cells via an interaction with cellular heparan sulfate. rsv-glycosaminoglycan (gag) interactions were evaluated using heparin-agarose affinity chromatography. rsv-f from a2- and b1/cp-52 (cp-52)-infected cell lysates, rsv-f derived from a recombinant vaccinia virus, and affinity-purified f protein all bound to and were specifically ...200010864656
trypsin-induced structural transformation in aquareovirus.aquareovirus, a member of the family reoviridae, is a large virus with multiple capsid layers surrounding a genome composed of 11 segments of double-stranded rna. biochemical studies have shown that treatment with the proteolytic agent trypsin significantly alters the infectivity of the virus. the most infectious stage of the virus is produced by a 5-min treatment with trypsin. however, prolonged trypsin treatment almost completely abolishes the infectivity. we have used three-dimensional electr ...200010864668
a cytoplasmic rna vector derived from nontransmissible sendai virus with efficient gene transfer and expression.we have recovered a virion from defective cdna of sendai virus (sev) that is capable of self-replication but incapable of transmissible-virion production. this virion delivers and expresses foreign genes in infected cells, and this is the first report of a gene expression vector derived from a defective viral genome of the paramyxoviridae. first, functional ribonucleoprotein complexes (rnps) were recovered from sev cloned cdna defective in the f (envelope fusion protein) gene, in the presence of ...200010864670
construction and characterization of monoclonal antibodies against western equine encephalitis virus.a repertoire of mouse monoclonal antibodies (mabs) against western equine encephalitis virus (wee) was constructed and characterized. anti-wee antibodies were expressed from hybridomas and purified by protein g chromatography. each of the antibodies was functionally assessed by indirect enzyme-linked immunosorbent assays (elisas), western blotting, and immunoprecipitations. all antibodies bound to wee antigen in elisas, whereas only a subgroup of antibodies was found to be active in western blot ...200010868791
a mutant of sindbis virus that is resistant to pyrazofurin encodes an altered rna polymerase.pyrazofurin (pzf), a cytidine analog and an inhibitor of orotate monophosphate decarboxylase, has been shown to decrease the levels of utp and ctp in treated cells. when sindbis virus (sv)-infected aedes albopictus cells were treated with pzf, the yield of virus was reduced 100- to 1000-fold. by serial passage of our standard sv(std) in ae. albopictus cells in the presence of increasing concentrations of pzf, a mutant, sv(pzf), was derived, which was not inhibited by pzf. sv(pzf) is also resista ...200010873749
effects of palmitoylation of replicase protein nsp1 on alphavirus infection.the membrane-associated alphavirus rna replication complex contains four virus-encoded subunits, the nonstructural proteins nsp1 to nsp4. semliki forest virus (sfv) nsp1 is hydrophobically modified by palmitoylation of cysteines 418 to 420. here we show that sindbis virus nsp1 is also palmitoylated on the same site (cysteine 420). when mutations preventing nsp1 palmitoylation were introduced into the genomes of these two alphaviruses, the mutant viruses remained viable and replicated to high tit ...200010888610
measles virus matrix protein specifies apical virus release and glycoprotein sorting in epithelial cells.in polarized epithelial cells measles virus (mv) is predominantly released at the apical cell surface, irrespective of the sorting of its two envelope glycoproteins f and h. it has been reported previously that the viral matrix (m) protein modulates the fusogenic capacity of the viral envelope glycoproteins. here, extant mv mutants and chimeras were used to determine the role of m protein in the transport of viral glycoproteins and release of progeny virions in polarized epithelial caco2 cells. ...200010899112
sindbis virus vector system for functional analysis of apoptosis regulators. 200010914042
ns3 serine protease of bovine viral diarrhea virus: characterization of active site residues, ns4a cofactor domain, and protease-cofactor interactions.the gene expression of bovine viral diarrhea virus (bvdv), a pestivirus, occurs via translation of a hypothetical polyprotein that is processed cotranslationally and posttranslationally by viral and cellular enzymes. a protease located in the n-terminal region of nonstructural (ns) protein ns3 catalyzes the cleavages, leading to the release of ns4a, ns4b, ns5a, and ns5b. our study provides experimental evidence that histidine at position 1658 and aspartic acid at position 1686 constitute togethe ...200010915606
semliki forest virus budding: assay, mechanisms, and cholesterol requirement.all enveloped viruses must bud through a cellular membrane in order to acquire their lipid bilayer, but little is known about this important stage in virus biogenesis. we have developed a quantitative biochemical assay to monitor the budding of semliki forest virus (sfv), an enveloped alphavirus that buds from the plasma membrane in a reaction requiring both viral spike proteins and nucleocapsid. the assay was based on cell surface biotinylation of newly synthesized virus spike proteins and retr ...200010933676
infection of human cells by dengue virus is modulated by different cell types and viral strains.although prior studies have investigated cellular infection by dengue virus (dv), many have used highly passaged strains. we have reassessed cellular infection by dv type 2 (dv2) using prototype and low-passage isolates representing genotypes from different geographic areas. we observed marked variation in the susceptibility to infection among cell types by different dv2 strains. hepg2 hepatoma cells were susceptible to infection by all dv2 strains assayed. although the prototype strain generate ...200010933688
mutations in the 5' nontranslated region of bovine viral diarrhea virus result in altered growth characteristics.the 5' nontranslated region (ntr) of pestiviruses functions as an internal ribosome entry site (ires) that mediates cap-independent translation of the viral polyprotein and probably contains additional cis-acting rna signals involved in crucial processes of the viral life cycle. computer modeling suggests that the 5'-terminal 75 nucleotides preceding the ires element form two stable hairpins, ia and ib. spontaneous and engineered mutations located in the genomic region comprising ia and ib were ...200010933696
identification of mouse hepatitis virus papain-like proteinase 2 activity.mouse hepatitis virus (mhv) is a 31-kb positive-strand rna virus that is replicated in the cytoplasm of infected cells by a viral rna-dependent rna polymerase, termed the replicase. the replicase is encoded in the 5'-most 22 kb of the genomic rna, which is translated to produce a polyprotein of >800 kda. the replicase polyprotein is extensively processed by viral and perhaps cellular proteinases to give rise to a functional replicase complex. to date, two of the mhv replicase-encoded proteinases ...200010933699
membrane interface-interacting sequences within the ectodomain of the human immunodeficiency virus type 1 envelope glycoprotein: putative role during viral fusion.we have identified a region within the ectodomain of the fusogenic human immunodeficiency virus type 1 (hiv-1) gp41, different from the fusion peptide, that interacts strongly with membranes. this conserved sequence, which immediately precedes the transmembrane anchor, is not highly hydrophobic according to the kyte-doolittle hydropathy prediction algorithm, yet it shows a high tendency to partition into the membrane interface, as revealed by the wimley-white interfacial hydrophobicity scale. we ...200010933713
chimeric yellow fever/dengue virus as a candidate dengue vaccine: quantitation of the dengue virus-specific cd8 t-cell response.we have constructed a chimeric yellow fever/dengue (yf/den) virus, which expresses the premembrane (prm) and envelope (e) genes from den type 2 (den-2) virus in a yf virus (yfv-17d) genetic background. immunization of balb/c mice with this chimeric virus induced a cd8 t-cell response specific for the den-2 virus prm and e proteins. this response protected yf/den virus-immunized mice against lethal dengue encephalitis. control mice immunized with the parental yfv-17d were not protected against de ...200010933719
characterization of the coronavirus m protein and nucleocapsid interaction in infected cells.coronavirus contains three envelope proteins, m, e and s, and a nucleocapsid, which consists of genomic rna and n protein, within the viral envelope. we studied the macromolecular interactions involved in coronavirus assembly in cells infected with a murine coronavirus, mouse hepatitis virus (mhv). coimmunoprecipitation analyses demonstrated an interaction between n protein and m protein in infected cells. pulse-labeling experiments showed that newly synthesized, unglycosylated m protein interac ...200010933723
sex differences in seoul virus infection are not related to adult sex steroid concentrations in norway rats.field studies of hantavirus infection in rodents report that a higher percentage of infected individuals are males than females. to determine whether males were more susceptible to hantavirus infection than females, adult male and female long evans rats (rattus norvegicus) were inoculated with doses of seoul virus ranging from 10(-4) to 10(6) pfu. the 50% infective doses (id(50)) were not significantly different for male and female rats (10(0.05) and 10(0.8) pfu, respectively). to determine whet ...200010933735
densovirinae as gene transfer vehicles.densoviruses present an attractive opportunity to develop expression vectors for insects. they exhibit several features that are beneficial for such vectors. the genomes of densoviruses are among the smallest of animal dna viruses, and, therefore are easier to use for cloning and transfection procedures. the fact that cloned densovirus genomes are infectious greatly simplifies their applications. it seems that both densovirus promoters have high constitutive activity and they can be further tran ...200010941569
specific ablation of antiviral gene expression in macrophages by antibody-dependent enhancement of ross river virus infection.ross river virus (rrv) is an indigenous australian arthropod-borne alphavirus responsible for epidemic polyarthritis (epa), myalgia, and lethargy in humans. macrophages and monocytes have been associated with human rrv disease, and previous studies have shown that rrv is capable of infecting macrophages via both a natural virus receptor and by fc receptor-mediated antibody-dependent enhancement (ade). similar to other viruses, such as human immunodeficiency virus and dengue virus, ade infection ...200010954537
potential dengue virus-triggered apoptotic pathway in human neuroblastoma cells: arachidonic acid, superoxide anion, and nf-kappab are sequentially involved.direct in vivo evidence for the susceptibility of human neuronal cells to dengue virus has not been reported. in this study, we demonstrated that type 2 dengue (den-2) virus infection induced extensive apoptosis in the human neuroblastoma cell line sk-n-sh. phospholipase a(2) (pla(2)) was activated by den-2 infection, which led to the generation of arachidonic acid (aa). inhibition of pla(2) activity by the pla(2) inhibitors, aacocf(3) and ono-rs-082, diminished den-2 virus-induced apoptosis. in ...200010954569
influenza virus assembly: effect of influenza virus glycoproteins on the membrane association of m1 protein.influenza virus matrix protein (m1), a critical protein required for virus assembly and budding, is presumed to interact with viral glycoproteins on the outer side and viral ribonucleoprotein on the inner side. however, because of the inherent membrane-binding ability of m1 protein, it has been difficult to demonstrate the specific interaction of m1 protein with hemagglutinin (ha) or neuraminidase (na), the influenza virus envelope glycoproteins. using triton x-100 (tx-100) detergent treatment o ...200010954572
inactivation and clearance of viruses during the manufacture of high purity factor ix.haemophilia is a bleeding disorder characterised by a deficiency in factor ix. replacement therapy in the form of a factor ix concentrate is a widely accepted practice. in this paper we describe a double virus inactivated chromatographic process for producing a high purity factor ix product, monofix((r))-vf. the process involves separation of the prothrombin complex by cryoprecipitation, fraction i precipitation and deae-cellulose adsorption, further ion-exchange chromatography of crude factor i ...200010964439
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