PMID(sorted ascending)
mutual interference between genomic rna replication and subgenomic mrna transcription in brome mosaic virus.replication by many positive-strand rna viruses includes genomic rna amplification and subgenomic mrna (sgrna) transcription. for brome mosaic virus (bmv), both processes occur in virus-induced, membrane-associated compartments, require bmv replication factors 1a and 2a, and use negative-strand rna3 as a template for genomic rna3 and sgrna syntheses. to begin elucidating their relations, we examined the interaction of rna3 replication and sgrna transcription in saccharomyces cerevisiae expressin ...200515650170
sindbis virus translation is inhibited by a pkr/rnase l-independent effector induced by alpha/beta interferon priming of dendritic cells.the tropism of sindbis virus (sb) for cells of the dendritic cell (dc) lineage and the virulence of sb in vivo are largely determined by the efficacy of alpha/beta interferon (ifn-alpha/beta)-mediated antiviral responses. these responses are essentially intact in the absence of pkr and/or rnase l (k. d. ryman, l. j. white, r. e. johnston, and w. b. klimstra, viral immunol. 15:53-76, 2002). in the present studies, we investigated the nature of antiviral effects and identity of antiviral effectors ...200515650175
involvement of clathrin-mediated endocytosis in human immunodeficiency virus type 1 entry.productive entry of human immunodeficiency virus (hiv) is believed to occur by direct fusion at the plasma membrane. endocytic uptake of hiv particles has been observed in several studies but is considered to be nonproductive, leading to virus degradation in the lysosome. we show here that endocytosis contributes significantly to productive hiv entry in hela cells by using trans dominant-negative mutants of dynamin and eps15. inducible expression of a dominant-negative mutant of dynamin in a cd4 ...200515650184
human immunodeficiency virus type 1 vectors with alphavirus envelope glycoproteins produced from stable packaging cells.alphavirus glycoproteins have broad host ranges. human immunodeficiency virus type 1 (hiv-1) vectors pseudotyped with their glycoproteins could extend the range of tissues that can be transduced in both humans and animal models. here, we established stable producer cell lines for hiv vectors pseudotyped with alphavirus ross river virus (rrv) and semliki forest virus (sfv) glycoproteins e2e1. rrv e2e1-stable clones could routinely produce high-titer pseudotyped vectors for at least 5 months. sfv ...200515650201
yellow fever 17d as a vaccine vector for microbial ctl epitopes: protection in a rodent malaria model.the yellow fever vaccine 17d (17d) is safe, and after a single immunizing dose, elicits long-lasting, perhaps lifelong protective immunity. one of the major challenges facing delivery of human vaccines in underdeveloped countries is the need for multiple injections to achieve full efficacy. to examine 17d as a vector for microbial t cell epitopes, we inserted the h-2k(d)-restricted ctl epitope of the circumsporozoite protein (cs) of plasmodium yoelii between 17d nonstructural proteins ns2b and n ...200515657290
requirement of the vesicular system for membrane permeabilization by sindbis virus.the vast majority of animal viruses enhance membrane permeability at two moments of infection. herein, we describe that the entry of sindbis virus (sv) in bhk cells promotes the co-entry of the macromolecule alpha-sarcin into the cytoplasm, thereby blocking translation. at a later stage, this protein toxin cannot enter the cell, while low molecular weight compounds, such as hygromycin b, readily pass through the plasma membrane of sindbis virus-infected bhk cells. to unveil the participation of ...200515661162
invariant valpha14 chain nkt cells promote plasmodium berghei circumsporozoite protein-specific gamma interferon- and tumor necrosis factor alpha-producing cd8+ t cells in the liver after poxvirus vaccination of mice.understanding the protective mechanism in the liver induced by recombinant vaccines against the pre-erythrocytic stages of malaria is important for vaccine development. most studies in mice have focused on splenic and peripheral blood t cells and identified gamma interferon (ifn-gamma)-producing cd8+ t cells as correlates of protection, which can be induced by prime-boost vaccination with recombinant poxviruses. invariant natural killer t (valpha14inkt) cells can also protect against liver stage ...200515664925
aedes aegypti salivary gland extracts modulate anti-viral and th1/th2 cytokine responses to sindbis virus infection.vector-borne viruses are naturally transmitted when a vector salivates during feeding on a vertebrate host. most laboratory studies of infection disregard the role that the vector plays in the pathogenesis of the virus. in this study, intradermal inoculations of aedes aegypti salivary gland extract (sge) and sindbis virus (sinv) were used to investigate the effect of mosquito feeding on the vertebrate immune response to infection with an arthropod-borne virus. murine cytokine expression in the s ...200415671753
heparin binding sites on ross river virus revealed by electron cryo-microscopy.cell surface glycosaminoglycans play important roles in cell adhesion and viral entry. laboratory strains of two alphaviruses, sindbis and semliki forest virus, have been shown to utilize heparan sulfate as an attachment receptor, whereas ross river virus (rrv) does not significantly interact with it. however, a single amino acid substitution at residue 218 in the rrv e2 glycoprotein adapts the virus to heparan sulfate binding and expands the host range of the virus into chicken embryo fibroblas ...200515680416
in vivo processing and isolation of furin protease-sensitive alphavirus glycoproteins: a new technique for producing mutations in virus assembly.sindbis virus particles are composed of three structural proteins (capsid/e2/e1). in the mature virion the e1 glycoprotein is organized in a highly constrained, energy-rich conformation. it is hypothesized that this energy is utilized to drive events that deliver the viral genome to the cytoplasm of a host cell. the extraction of the e1 glycoprotein from virus membranes with detergent results in disulfide-bridge rearrangement and the collapse of the protein to a number of low-energy, non-native ...200515680428
characterization of helper virus-independent cytopathogenic classical swine fever virus generated by an in vivo rna recombination system.molecular analyses revealed that most cytopathogenic (cp) pestivirus strains evolve from noncytopathogenic (noncp) viruses by nonhomologous rna recombination. in contrast to bovine viral diarrhea virus (bvdv), cp classical swine fever virus (csfv) field isolates were rarely detected and always represented helper virus-dependent subgenomes. to investigate rna recombination in more detail, we recently established an in vivo system allowing the efficient generation of recombinant cp bvdv strains in ...200515681445
proteasome inhibition protects ht22 neuronal cells from oxidative glutamate toxicity.oxidative stress caused by glutathione depletion after prolonged exposure to extracellular glutamate leads to a form of neuronal cell death that exhibits morphologically mixed features of both apoptosis and necrosis. however, specific downstream executioners involved in this form of cell death have yet to be identified. we report here that glutamate exposure does not activate caspase-3 in the ht22 neuronal cell line. furthermore, no cytoprotection was achieved with either the pan-caspase inhibit ...200515686484
subcellular localisation of recombinant alpha- and gamma-synuclein.alpha-synuclein, a protein implicated in neurodegenerative diseases and of elusive physiological function owes its name to an observed presence in presynaptic and nuclear compartments. however, its nuclear localisation has remained controversial. we expressed synuclein-egfp fusion proteins in organotypic rat hippocampal slice cultures and murine hippocampal primary neurons using a sindbis virus expression system. recombinant full-length alpha-synuclein accumulated in presynaptic locations, mimic ...200515691713
novel replicon-based reporter gene assay for detection of rubella virus in clinical specimens.proof of concept for a novel diagnostic assay for rubella virus (rub) based on rub replicons expressing reporter genes was demonstrated. rub replicons have the structural protein coding region replaced with a reporter gene such as green fluorescent protein or chloramphenicol acetyltransferase. previously, it was shown that a replicon construct with a specific in-frame deletion in the nonstructural protein coding region (noti, approximately nucleotides 1500 to 2100 of the genome) failed to replic ...200515695695
mosquitoborne viruses, czech republic, 2002.specimens from residents (n = 497) of an area affected by the 2002 flood were examined serologically for mosquitoborne viruses. antibodies were detected against tahyna (16%), sindbis (1%), and batai (0.2%) viruses, but not west nile virus. an examination of paired serum samples showed 1 tahyna bunyavirus (california group) infection.200515705333
lentiviral vector retargeting to p-glycoprotein on metastatic melanoma through intravenous injection.targeted gene transduction to specific tissues and organs through intravenous injection would be the ultimate preferred method of gene delivery. here, we report successful targeting in a living animal through intravenous injection of a lentiviral vector pseudotyped with a modified chimeric sindbis virus envelope (termed m168). m168 pseudotypes have high titer and high targeting specificity and, unlike other retroviral pseudotypes, have low nonspecific infectivity in liver and spleen. a mouse can ...200515711560
phospholipase d1-promoted release of tissue plasminogen activator facilitates neurite outgrowth.temporal lobe epilepsy (tle) is the most common form of epilepsy, affecting approximately 1-2% of the population. seizure events resulting from tle are characterized by aberrant hippocampal mossy fiber sprouting and plastic responses that affect brain function. seizure susceptibility is modulated by the enzyme tissue plasminogen activator (tpa), the normal physiological role of which includes promotion of synaptic reorganization in the mossy fiber pathway by initiating a proteolytic cascade that ...200515716416
antiviral activity of human lactoferrin: inhibition of alphavirus interaction with heparan sulfate.human lactoferrin is a component of the non-specific immune system with distinct antiviral properties. we used alphaviruses, adapted to interaction with heparan sulfate (hs), as a tool to investigate the mechanism of lactoferrin's antiviral activity. lactoferrin inhibited infection of bhk-21 cells by hs-adapted, but not by non-adapted, sindbis virus (sin) or semliki forest virus (sfv). lactoferrin also inhibited binding of radiolabeled hs-adapted viruses to bhk-21 cells or liposomes containing l ...200515721362
association of sindbis virus capsid protein with phospholipid membranes and the e2 glycoprotein: implications for alphavirus assembly.a late stage in assembly of alphaviruses within infected cells is thought to be directed by interactions between the nucleocapsid and the cytoplasmic domain of the e2 protein, a component of the viral e1/e2 glycoprotein complex that is embedded in the plasma membrane. recognition between the nucleocapsid protein and the e2 protein was explored in solution using nmr spectroscopy, as well as in binding assays using a model phospholipid membrane system that incorporated a variety of sindbis virus e ...200515723524
early expression of ampa receptors and lack of nmda receptors in developing rat climbing fibre synapses.whether nascent glutamatergic synapses acquire their ampa receptors constitutively or via a regulated pathway triggered by pre-existing nmda receptor activation is still an open issue. here, we provide evidence that some glutamatergic synapses develop without expressing nmda receptors. using immunocytochemistry, we showed that synapses between developing rat climbing fibres and purkinje cells expressed glur2-containing ampa receptors as soon as they were formed (i.e. on embryonic day 19) but nev ...200515731186
dual mechanisms of pestiviral superinfection exclusion at entry and rna replication.for many viruses, primary infection has been shown to prevent superinfection by a homologous second virus. in this study, we investigated superinfection exclusion of bovine viral diarrhea virus (bvdv), a positive-sense rna pestivirus. cells acutely infected with bvdv were protected from superinfection by homologous bvdv but not with heterologous vesicular stomatitis virus. superinfection exclusion was established within 30 to 60 min but was lost upon passaging of persistently infected cells. sup ...200515731218
characterization of an in vitro model of alphavirus infection of immature and mature neurons.terminally differentiated, mature neurons are essential cells that are not easily regenerated. neurotropic viruses, such as sindbis virus (sv), cause encephalomyelitis through their ability to replicate in neurons. sv causes the death of immature neurons, while mature neurons can often survive infection. the lack of a reproducible and convenient neuronal cell culture system has hindered a detailed study of the differences in levels of virus replication between immature and mature neurons and the ...200515731238
3'-terminal sequence in poliovirus negative-strand templates is the primary cis-acting element required for vpgpupu-primed positive-strand initiation.the 5' cloverleaf in poliovirus rna has a direct role in regulating the stability, translation, and replication of viral rna. in this study, we investigated the role of stem a in the 5' cloverleaf in regulating the stability and replication of poliovirus rna in hela s10 translation-replication reactions. our results showed that disrupting the duplex structure of stem a destabilized viral rna and inhibited efficient negative-strand synthesis. surprisingly, the duplex structure of stem a was not r ...200515731251
inhibition of viral replication by ribozyme: mutational analysis of the site and mechanism of antiviral activity.a controlled mutational study was used to determine the site and mechanism of the antiviral action of ribozymes that inhibit sindbis virus replication. a hairpin ribozyme targeting g575 of the sindbis virus genomic rna was designed and cloned into a minimized alphavirus amplicon vector. cells that were stably transfected with this construct expressed low levels of a constitutive transcript containing the ribozyme plus recognition sequences for sindbis rna replicase. upon infection, the ribozyme ...200515731266
cell type-specific targeting with sindbis pseudotyped lentiviral vectors displaying anti-ccr5 single-chain antibodies.lentiviral vectors are among the most efficient tools for gene delivery into mammalian cells. a major goal of lentiviral gene delivery systems is to develop vectors that can efficiently target specific cell types. in the present work, we attempt to generate viral particles for targeting gene delivery. we have used ccr5-positive cells as the target for our strategy. therefore, we developed a novel sindbis pseudotyped lentiviral vector where the sindbis receptor binding envelope protein was modifi ...200515761262
acid-resistant bovine pestivirus requires activation for ph-triggered fusion during entry.the route of internalization of the pestivirus bovine viral diarrhea virus (bvdv) was studied by using different chemical and biophysical inhibitors of endocytosis. expression of the dominant-negative mutant dyn(k44a) of the gtpase dynamin in mdbk cells, as well as the treatment of the cells with chlorpromazine and beta-methyl-cyclodextrin inhibited bvdv entry. bvdv infection was also abolished by potassium (k+) depletion, hyperosmolarity, and different inhibitors of endosomal acidification. we ...200515767420
identification of adult mouse neurovirulence determinants of the sindbis virus strain ar86.sindbis virus infection of mice has provided valuable insight into viral and host factors that contribute to virus-induced neurologic disease. in an effort to further define the viral genetic elements that contribute to adult mouse neurovirulence, the neurovirulent sindbis virus strain ar86 was compared to the closely related (22 single amino acid coding changes and the presence or absence of an 18-amino-acid sequence in nsp3 [positions 386 to 403]) but avirulent girdwood strain. initial studies ...200515767423
two dimensional vopba reveals laminin receptor (lamr1) interaction with dengue virus serotypes 1, 2 and 3.the search for the dengue virus receptor has generated many candidates often identified only by molecular mass. the wide host range of the viruses in vitro combined with multiple approaches to identifying the receptor(s) has led to the notion that many receptors or attachment proteins may be involved and that the different dengue virus serotypes may utilize different receptors on the same cells as well as on different cell types.200515790424
recoding elements located adjacent to a subset of eukaryal selenocysteine-specifying uga codons.incorporation of the 21st amino acid, selenocysteine, into proteins is specified in all three domains of life by dynamic translational redefinition of uga codons. in eukarya and archaea, selenocysteine insertion requires a cis-acting selenocysteine insertion sequence (secis) usually located in the 3'utr of selenoprotein mrnas. here we present comparative sequence analysis and experimental data supporting the presence of a second stop codon redefinition element located adjacent to a selenocystein ...200515791204
neuronal cell death in alphavirus encephalomyelitis.alphaviruses are mosquito-borne, enveloped, plus-strand rna viruses that cause a spectrum of diseases in humans that include fever, rash, arthritis, meningitis, and encephalomyelitis. sindbis virus (sinv) is the prototype alphavirus, causes encephalomyelitis in mice, and provides a model system for studying the pathogenesis of alphavirus-induced neurological disease. major target cells for sinv infection in the central nervous system (cns) are neurons, and both host and viral factors determine t ...200515791951
development of cell-based assays for in vitro characterization of hepatitis c virus ns3/4a protease inhibitors.a recombinant vaccinia virus, expressing the ns3-to-ns5 region of the n clone of hepatitis c virus (hcv), was generated and utilized both in a gel-based assay and in an enzyme-linked immunosorbent assay (elisa) to evaluate the pyrrolidine-5,5-trans-lactams, a series of inhibitors of the hcv ns3/4a protease. the absolute levels of processed, mature hcv nonstructural proteins in this system were found to decrease in the presence of the trans-lactams. monitoring of this reduction enabled end points ...200515793116
inhibition of flavivirus infections by antisense oligomers specifically suppressing viral translation and rna replication.rna elements within flavivirus genomes are potential targets for antiviral therapy. a panel of phosphorodiamidate morpholino oligomers (pmos), whose sequences are complementary to rna elements located in the 5'- and 3'-termini of the west nile (wn) virus genome, were designed to anneal to important cis-acting elements and potentially to inhibit wn infection. a novel arg-rich peptide was conjugated to each pmo for efficient cellular delivery. these pmos exhibited various degrees of antiviral acti ...200515795246
requirements at the 3' end of the sindbis virus genome for efficient synthesis of minus-strand rna.the 3'-untranslated region of the sindbis virus genome is 0.3 kb in length with a 19-nucleotide conserved sequence element (3' cse) immediately preceding the 3'-poly(a) tail. the 3' cse and poly(a) tail have been assumed to constitute the core promoter for minus-strand rna synthesis during genome replication; however, their involvement in this process has not been formally demonstrated. utilizing both in vitro and in vivo analyses, we have examined the role of these elements in the initiation of ...200515795249
virus-specific and bystander cd8 t cells recruited during virus-induced encephalomyelitis.neurotropic coronavirus-induced encephalitis was used to evaluate recruitment, functional activation, and retention of peripheral bystander memory cd8+ t cells. mice were first infected with recombinant vaccinia virus expressing a non-cross-reactive human immunodeficiency virus (hiv) epitope, designated p18. following establishment of an endogenous p18-specific memory cd8+ t-cell population, mice were challenged with coronavirus to directly compare recruitment, longevity, and activation characte ...200515795256
identification of structural domains involved in astrovirus capsid biology.coat proteins of non-enveloped, icosahedral viruses must perform a variety of functions during their life cycle such as assembly of the coat protein subunits into a closed shell, specific encapsidation of the viral nucleic acid, maturation of the capsid, interaction with host receptors, and disassembly to deliver the genetic information into the newly infected cell. a thorough understanding of the multiple capsid properties at the molecular level is required in order to identify potential target ...200515802951
mechanisms of reovirus-induced cell death and tissue injury: role of apoptosis and virus-induced perturbation of host-cell signaling and transcription factor activation.reoviruses have provided insight into the roles played by specific viral genes and the proteins they encode in virus-induced cell death and tissue injury. apoptosis is a major mechanism of cell death induced by reoviruses. reovirus-induced apoptosis involves both death-receptor and mitochondrial cell death pathways. reovirus infection is associated with selective activation of mitogen activated protein kinase (mapk) cascades including jnk/sapk. infection also perturbs transcription factor signal ...200515802955
class ii fusion protein of alphaviruses drives membrane fusion through the same pathway as class i proteins.viral fusion proteins of classes i and ii differ radically in their initial structures but refold toward similar conformations upon activation. do fusion pathways mediated by alphavirus e1 and influenza virus hemagglutinin (ha) that exemplify classes ii and i differ to reflect the difference in their initial conformations, or concur to reflect the similarity in the final conformations? here, we dissected the pathway of low ph-triggered e1-mediated cell-cell fusion by reducing the numbers of acti ...200515809312
induction of human papillomavirus type 16-specific immunologic responses in a normal and an human papillomavirus-infected populations.human papillomavirus (hpv) infection, especially with the oncogenic genotypes, is the most important risk factor for developing cervical cancer. we focused on generating hpv16 e7-specific cytotoxic cd8(+) t lymphocytes and evaluating hpv16 e7-specific immune responses in hpv16-infected and uninfected populations. peripheral blood mononuclear cells (pbmcs) were first collected from an uninfected group with an human lymphocyte antigen (hla) a2 haplotype (four volunteers). mature monocyte-derived d ...200515819706
gamma interferon-dependent, noncytolytic clearance of sindbis virus infection from neurons in vitro.due to the nonrenewable nature of neurons, recovery from viral infection of the central nervous system requires noncytopathic mechanisms for control of virus replication. recovery from alphavirus encephalitis can occur without apparent neurological damage through the effects of antibody and gamma interferon (ifn-gamma). to establish an in vitro cell culture system that will allow the study of mechanisms of ifn-gamma-mediated control of sindbis virus (sinv) replication in neurons, we have charact ...200515827152
rift valley fever virus nonstructural protein nss promotes viral rna replication and transcription in a minigenome system.rift valley fever virus (rvfv), which belongs to the genus phlebovirus, family bunyaviridae, has a tripartite negative-strand genome (s, m, and l segments) and is an important mosquito-borne pathogen for domestic animals and humans. we established an rvfv t7 rna polymerase-driven minigenome system in which t7 rna polymerase from an expression plasmid drove expression of rna transcripts for viral proteins and minigenome rna transcripts carrying a reporter gene between both termini of the m rna se ...200515827175
ross river virus disease reemergence, fiji, 2003-2004.we report 2 clinically characteristic and serologically positive cases of ross river virus infection in canadian tourists who visited fiji in late 2003 and early 2004. this report suggests that ross river virus is once again circulating in fiji, where it apparently disappeared after causing an epidemic in 1979 to 1980.200515829203
effective protection of pigs against lethal pseudorabies virus infection after a single injection of low-dose sindbis-derived plasmids encoding prv gb, gc and gd glycoproteins.this study compared protection of pigs against lethal pseudorabies virus (prv) infection induced by a single injection of various quantities of sindbis virus-derived plasmids encoding prv glycoproteins gb, gc and gd. pigs were injected with 340, 68 or 13 microg of each plasmid. few immune differences were observed after dna injection and more importantly the pigs of the three groups were equally protected against virulent prv infection. single-shot injection of 13 microg of each prv glycoprotein ...200515837372
bcl-2-related protein a1 is an endogenous and cytokine-stimulated mediator of cytoprotection in hyperoxic acute lung injury.hyperoxic acute lung injury (hali) is characterized by a cell death response with features of apoptosis and necrosis that is inhibited by il-11 and other interventions. we hypothesized that bfl-1/a1, an antiapoptotic bcl-2 protein, is a critical regulator of hali and a mediator of il-11-induced cytoprotection. to test this, we characterized the expression of a1 and the oxygen susceptibility of wt and il-11 tg(+) mice with normal and null a1 loci. in wt mice, 100% o(2) caused tunel(+) cell death, ...200515841185
stereophysicochemical variability plots highlight conserved antigenic areas in flaviviruses.flaviviruses, which include dengue (dv) and west nile (wn), mutate in response to immune system pressure. identifying escape mutants, variant progeny that replicate in the presence of neutralizing antibodies, is a common way to identify functionally important residues of viral proteins. however, the mutations typically occur at variable positions on the viral surface that are not essential for viral replication. methods are needed to determine the true targets of the neutralizing antibodies.200515845145
intranasal vaccination induces protective immunity against intranasal infection with virulent francisella tularensis biovar a.the inhalation of francisella tularensis biovar a causes pneumonic tularemia associated with high morbidity and mortality rates in humans. exposure to f. tularensis usually occurs by accident, but there is increasing awareness that f. tularensis may be deliberately released in an act of bioterrorism or war. the development of a vaccine against pneumonic tularemia has been limited by a lack of information regarding the mechanisms required to protect against this disease. vaccine models for f. tul ...200515845466
virus-inducible reporter genes as a tool for detecting and quantifying influenza a virus replication.the use of influenza a virus-inducible reporter gene segments in detecting influenza a virus replication was investigated. the rna polymerase i promoter/terminator cassette was used to express rna transcripts encoding green fluorescence protein or firefly luciferase flanked by the untranslated regions of the influenza a/wsn/33 nucleoprotein (np) segment. reporter gene activity was detected after reconstitution of the influenza a virus polymerase complex from cdna or after virus infection, and wa ...200515847914
high-level expression of marek's disease virus glycoprotein c is detrimental to virus growth in vitro.expression levels of marek's disease virus (mdv) glycoprotein c (gc) are significantly reduced after serial virus passage in cell culture. reduced gc expression coincides with enhanced mdv growth in vitro and attenuation. to analyze this phenomenon in detail, a full-length infectious mdv clone was modified by red-based and shuttle mutagenesis in escherichia coli. besides a gc-negative deletion mutant harboring a kanamycin resistance gene, a markerless mutant with the u(l)44 gene deleted was cons ...200515857974
roles of cd4+ t-cell-independent and -dependent antibody responses in the control of influenza virus infection: evidence for noncognate cd4+ t-cell activities that enhance the therapeutic activity of antiviral antibodies.previous studies have indicated that b cells make a significant contribution to the resolution of influenza virus infection. to determine how b cells participate in the control of the infection, we transferred intact, major histocompatibility complex class ii (mhc-ii)-negative or b-cell receptor (bcr)-transgenic spleen cells into b-cell-deficient and cd8(+) t-cell-depleted mumt mice, termed mumt(-8), and tested them for ability to recover from infection. mumt(-8) mice that received no spleen cel ...200515857980
targeting human glioblastoma cells: comparison of nine viruses with oncolytic potential.brain tumors classified as glioblastomas have proven refractory to treatment and generally result in death within a year of diagnosis. we used seven in vitro tests and one in vivo trial to compare the efficacy of nine different viruses for targeting human glioblastoma. green fluorescent protein (gfp)-expressing vesicular stomatitis (vsv), sindbis virus, pseudorabies virus (prv), adeno-associated virus (aav), and minute virus of mice i-strain (mvmi) and mvmp all infected glioblastoma cells. mouse ...200515857987
suppression of proliferation of poliovirus and porcine parvovirus by novel phenoxazines, 2-amino-4,4 alpha-dihydro-4 alpha-7-dimethyl-3h-phenoxazine-3-one and 3-amino-1,4 alpha-dihydro-4 alpha-8-dimethyl-2h-phenoxazine-2-one.the present study aimed at investigating the antiviral effects of 2-amino-4,4alpha-dihydro-4alpha-7-dimethyl-3h-phenoxazine-3-one (phx-1) and 3-amino-1,4alpha-dihydro-4alpha-8-dimethyl-2h-phenoxazine-2-one (phx-2) on 6 representative viruses: poliovirus, porcine parvovirus, simian virus 40 (sv-40), herpes simplex virus-1 (hsv-1), sindbis virus, and vesicular stomatitis virus (vsv). phx-1 and phx-2 suppressed the proliferation of poliovirus in vero cells and that of porcine parvovirus in esk cell ...200515863903
induction of antigen-specific immune responses against malignant brain tumors by intramuscular injection of sindbis dna encoding gp100 and il-18.we constructed psin-sv40-hdv-sv40pa, an improved sindbis dna expression vector, and evaluated the potential of this vector system for brain tumor therapy. we investigated whether immunizing mice with xenogeneic dna encoding human gp100 and mouse il-18 would enhance the antitumor responses. to study the immune mechanisms involved in tumor regression, we examined tumor growth in b16-gp100-implanted brain tumor models using t-cell subset-depleted and ifn-gamma-neutralized mice. hugp100/mil-18 vacci ...200515869409
identification and phenotypic characterization of sphingomonas wittichii strain rw1 by peptide mass fingerprinting using matrix-assisted laser desorption ionization-time of flight mass spectrometry.mass spectrometry is a potentially attractive means of monitoring the survival and efficacy of bioaugmentation agents, such as the dioxin-mineralizing bacterium sphingomonas wittichii strain rw1. the biotransformation activity of rw1 phenotypes is determined primarily by the presence and concentration of the dioxin dioxygenase, an enzyme initiating the degradation of both dibenzo-p-dioxin and dibenzofuran (df). we explored the possibility of identifying and characterizing putative cultures of rw ...200515870332
properties of sindbis virus vectors produced with a chimeric split helper system.we have evaluated a chimeric, two-component sindbis virus packaging system. as expected, use of this combination of two modified helper rna species prevented formation of infection competent sindbis viruses as analyzed by serial passaging. we observed, however, that vectors produced using this method were able to spread in bhk cell cultures and formed clusters of transgene positive cells that did not display cytopathic effects for up to 3 days post-transduction. formation of spreading sindbis vi ...200515870906
clinical and laboratory manifestations of sindbis virus infection: prospective study, finland, 2002-2003.sindbis virus (sinv) is widespread in europe, africa, australia, and asia, but clinical infection occurs as epidemics in a few geographically restricted areas. we recently proved, by virus isolation from patients, that sinv is the causative agent of pogosta disease, a mosquito-borne rash-arthritis occurring as larger epidemics every seventh year in finland. altogether, 86 patients with serologically verified sinv infection were recruited to the present study during the 2002 outbreak. we now desc ...200515871114
persistent expression of factor viii in vivo following nonprimate lentiviral gene transfer.hemophilia a is a clinically important coagulation disorder caused by the lack or abnormality of plasma coagulation factor viii (fviii). gene transfer of the fviii cdna to hepatocytes using lentiviral vectors is a potential therapeutic approach. we investigated the efficacy of feline immunodeficiency virus (fiv)-based vectors in targeting hepatocytes and correcting fviii deficiency in a hemophilia a mouse model. several viral envelope glycoproteins were screened for efficient fiv vector pseudoty ...200515886327
long-range rna-rna interactions circularize the dengue virus genome.secondary and tertiary rna structures present in viral rna genomes play essential regulatory roles during translation, rna replication, and assembly of new viral particles. in the case of flaviviruses, rna-rna interactions between the 5' and 3' ends of the genome have been proposed to be required for rna replication. we found that two rna elements present at the ends of the dengue virus genome interact in vitro with high affinity. visualization of individual molecules by atomic force microscopy ...200515890901
the 5'-terminal region of the aichi virus genome encodes cis-acting replication elements required for positive- and negative-strand rna synthesis.aichi virus is a member of the family picornaviridae. it has already been shown that three stem-loop structures (sl-a, sl-b, and sl-c, from the 5' end) formed at the 5' end of the genome are critical elements for viral rna replication. in this study, we further characterized the 5'-terminal cis-acting replication elements. we found that an additional structural element, a pseudoknot structure, is formed through base-pairing interaction between the loop segment of sl-b (nucleotides [nt] 57 to 60) ...200515890931
characterization of human immunodeficiency virus gag-specific gamma interferon-expressing cells following protective mucosal immunization with alphavirus replicon particles.a safe, replication-defective viral vector that can induce mucosal and systemic immune responses and confer protection against many infectious pathogens, such as human immunodeficiency virus type 1 (hiv-1), may be an ideal vaccine platform. accordingly, we have generated and tested alphavirus replicon particles encoding hiv-1 gag from sindbis virus (sin-gag) and venezuelan equine encephalitis virus (vee-gag), as well as chimeras between the two (vee/sin-gag). following intramuscular (i.m.), intr ...200515890953
loss of interferon regulatory factor 3 in cells infected with classical swine fever virus involves the n-terminal protease, npro.we show that cells infected with the pestivirus classical swine fever virus (csfv) fail to produce alpha/beta interferon not only following treatment with double-stranded rna but also after superinfection with a heterologous virus, the alphavirus sindbis virus, a virus shown to normally induce interferon. we investigated whether the inhibition of interferon synthesis by csfv involved a block in interferon regulatory factor 3 (irf3) activity. cells infected with csfv exhibited a lack of transloca ...200515890962
secondary dengue virus type 4 infections in vietnam.this study was designated to describe clinical and biological features of patients with a suspected diagnosis of dengue fever/dengue hemorrhagic fever during an outbreak in central vietnam. one hundred and twenty-five consecutive patients hospitalized at khanh hoa and binh thuan provincial hospitals between november 2001 and january 2002 with a diagnosis of suspected dengue infection were included in the present study. viruses were isolated in c6/36 and vero e6 cell cultures or detected by rt-pc ...200515906664
rubella virus capsid protein modulation of viral genomic and subgenomic rna synthesis.the ratio of the subgenomic (sg) to genome rna synthesized by rubella virus (rub) replicons expressing the green fluorescent protein reporter gene (rubrep/gfp) is substantially higher than the ratio of these species synthesized by rub (4.3 for rubrep/gfp vs. 1.3-1.4 for rub). it was hypothesized that this modulation of the viral rna synthesis was by one of the virus structural protein genes and it was found that introduction of the capsid (c) protein gene into the replicons as an in-frame fusion ...200515907967
association of signr1 with tlr4-md-2 enhances signal transduction by recognition of lps in gram-negative bacteria.signr1, a member of a new family of mouse c-type lectins, is expressed at high levels in macrophages (mphi) within the splenic marginal zone, lymph node medulla, and in some strains, in peritoneal cavity. we previously reported that signr1 captures gram-negative bacteria, such as escherichia coli and salmonella typhimurium, as well as candida albicans. we have now investigated the precise ligands and innate responses that involve signr1. the interaction of signr1 with fitc-dextran and e. coli wa ...200515908446
no signalling decodes frequency of neuronal activity and generates synapse-specific plasticity in mouse cerebellum.nitric oxide (no) is an intercellular messenger regulating neuronal functions. to visualize no signalling in the brain, we generated a novel fluorescent no indicator, which consists of the heme-binding region (hbr) of soluble guanylyl cyclase and the green fluorescent protein. the indicator (hbr-gfp) was expressed in the purkinje cells of the mouse cerebellum and we imaged no signals in acute cerebellar slices upon parallel fibre (pf) activation with a train of burst stimulations (bs, each bs co ...200515919714
complement activation is required for induction of a protective antibody response against west nile virus infection.infection with west nile virus (wnv) causes a severe infection of the central nervous system (cns) with higher levels of morbidity and mortality in the elderly and the immunocompromised. experiments with mice have begun to define how the innate and adaptive immune responses function to limit infection. here, we demonstrate that the complement system, a major component of innate immunity, controls wnv infection in vitro primarily in an antibody-dependent manner by neutralizing virus particles in ...200515919902
noncytopathic replication of venezuelan equine encephalitis virus and eastern equine encephalitis virus replicons in mammalian cells.venezuelan equine encephalitis (vee) and eastern equine encephalitis (eee) viruses are important, naturally emerging zoonotic viruses. they are significant human and equine pathogens which still pose a serious public health threat. both vee and eee cause chronic infection in mosquitoes and persistent or chronic infection in mosquito-derived cell lines. in contrast, vertebrate hosts infected with either virus develop an acute infection with high-titer viremia and encephalitis, followed by host de ...200515919912
single amino acid insertions at the junction of the sindbis virus e2 transmembrane domain and endodomain disrupt virus envelopment and alter infectivity.the final steps in the envelopment of sindbis virus involve specific interactions of the e2 endodomain with the virus nucleocapsid. deleting e2 k at position 391 (e2 deltak391) resulted in the disruption of virus assembly in mammalian cells but not insect cells (host range mutant). this suggested unique interactions of the e2 deltak391 endodomain with the different biochemical environments of the mammalian and insect cell lipid bilayers. to further investigate the role of the amino acid residues ...200515919921
indoleamine 2,3-dioxygenase mediates cell type-specific anti-measles virus activity of gamma interferon.gamma interferon (ifn-gamma) has been shown to be increased in sera from patients with acute measles and after vaccination, to exhibit protective functions in brains of patients with subacute sclerosing panencephalitis, and to mediate a noncytolytic clearance of measles virus (mv) from rodent brains. in order to reveal a possible intracellular antiviral activity in the absence of antigen presentation and cytotoxic t cells, we investigated ifn-gamma-induced effects on mv replication in various ti ...200515919929
reversible acid-induced inactivation of the membrane fusion protein of semliki forest virus.previously, it has been shown that the exposure of semliki forest virus (sfv) to a mildly acidic environment induces a rapid and complete loss of the ability of the virus to bind and fuse to target membranes added subsequently. in the present study, incubation of sfv at low ph followed by a specific reneutralization step resulted in a partial reversion of this loss of viral fusion capacity, as assessed in a liposomal model system. also, the ability of the viral e1 fusion protein to undergo lipos ...200515919953
targeting sindbis virus-based vectors to fc receptor-positive cell types.some viruses display enhanced infection for fc receptor (fcr)-positive cell types when complexed with virus-specific immunoglobulin (ig). this process has been termed antibody-dependent enhancement of viral infection (ade). we reasoned that the mechanism of ade could be exploited and adapted to target alphavirus-based vectors to fcr-positive cell types. towards this goal, recombinant sindbis viruses were constructed that express 1 to 4 immunoglobulin-binding domains of protein l (ppl) as n-termi ...200515922395
california serogroup gc (g1) glycoprotein is the principal determinant of ph-dependent cell fusion and entry.members of the california serogroup of orthobunyaviruses, particularly la crosse (lac) and tahyna (tah) viruses, are significant human pathogens in areas where their mosquito vectors are endemic. previous studies using wild-type lac and tah181/57, a highly neurovirulent strain with low neuroinvasiveness (janssen, r., gonzalez-scarano, f., nathanson, n., 1984. mechanisms of bunyavirus virulence. comparative pathogenesis of a virulent strain of la crosse and an avirulent strain of tahyna virus. la ...200515923017
the use of gene silencing to study the role of dopa decarboxylase in mosquito melanization reactions.mosquito melanization involves hydroxylation of tyrosine to dopa, which then is oxidized to dopaquinone by phenoloxidase, or decarboxylated to dopamine by dopa decarboxlase (ddc). an armigeres subalbatus cdna encoding ddc was cloned and real-time pcr analysis revealed increased transcripts in blood-fed and microfilariae (mf)-inoculated mosquitoes. a double subgenomic sindbis virus was used to silence ddc and assess its role in melanization of mf. ddc transcription and activity were significantly ...200515926892
peptide inhibitors of dengue virus and west nile virus infectivity.viral fusion proteins mediate cell entry by undergoing a series of conformational changes that result in virion-target cell membrane fusion. class i viral fusion proteins, such as those encoded by influenza virus and human immunodeficiency virus (hiv), contain two prominent alpha helices. peptides that mimic portions of these alpha helices inhibit structural rearrangements of the fusion proteins and prevent viral infection. the envelope glycoprotein (e) of flaviviruses, such as west nile virus ( ...200515927084
design, synthesis, and evaluation of dioxane-based antiviral agents targeted against the sindbis virus capsid protein.dioxane-based antiviral agents targeted to the hydrophobic binding pocket of sindbis virus capsid protein were designed by computer graphics molecular modeling and synthesized. virus production using sin-ires-luc and capsid assembly were monitored to evaluate antiviral activity. a compound with a three-carbon linker chain connecting two dioxane moieties inhibited virus production by 50% at a concentration of 40 microm, while (r)-hydroxymethyldioxane inhibited virus production by 50% at a concent ...200515927464
interferon and cytomegalovirus in vivo and in vitro. 196715938281
rab7 associates with early endosomes to mediate sorting and transport of semliki forest virus to late endosomes.semliki forest virus (sfv) is internalized by clathrin-mediated endocytosis, and transported via early endosomes to late endosomes and lysosomes. the intracellular pathway taken by individual fluorescently labeled sfv particles was followed using immunofluorescence in untransfected cells, and by video-enhanced, triple-color fluorescence microscopy in live cells transfected with gfp- and rfp-tagged rab5, rab7, rab4, and arf1. the viruses progressed from rab5-positive early endosomes to a populati ...200515954801
enhanced potency of plasmid dna microparticle human immunodeficiency virus vaccines in rhesus macaques by using a priming-boosting regimen with recombinant proteins.dna vaccines have been used widely in experimental primate models of human immunodeficiency virus (hiv), but their effectiveness has been limited. in this study, we evaluated three technologies for increasing the potency of dna vaccines in rhesus macaques. these included dna encoding sindbis virus rna replicons (psincp), cationic poly(lactide-co-glycolide) (plg) microparticles for dna delivery, and recombinant protein boosting. the dna-based psincp replicon vaccines encoding hiv gag and env were ...200515956564
innate stat1-dependent genomic response of neurons to the antiviral cytokine alpha interferon.alpha/beta interferons (ifns-alpha/beta) are cytokines that play an essential role in the host defense against viral infection. our previous studies have shown that the key ifn signaling molecule stat1 is highly elevated and activated in central nervous system neurons during viral infection and in transgenic mice with astrocyte production of ifn-alpha (glial fibrillary acidic protein [gfap]-ifn-alpha), suggesting that neurons are a very responsive target cell population for ifns. to elucidate th ...200515956575
control of translation by the 5'- and 3'-terminal regions of the dengue virus genome.the genomic rnas of flaviviruses such as dengue virus (den) have a 5' m7gpppn cap like those of cellular mrnas but lack a 3' poly(a) tail. we have studied the contributions to translational expression of 5'- and 3'-terminal regions of the den serotype 2 genome by using luciferase reporter mrnas transfected into vero cells. dcld rna contained the entire den 5' and 3' untranslated regions (utrs), as well as the first 36 codons of the capsid coding region fused to the luciferase reporter gene. capp ...200515956576
envelope protein glycosylation status influences mouse neuroinvasion phenotype of genetic lineage 1 west nile virus strains.the introduction of west nile virus (wnv) into north america has been associated with relatively high rates of neurological disease and death in humans, birds, horses, and some other animals. previous studies identified strains in both genetic lineage 1 and genetic lineage 2, including north american isolates of lineage 1, that were highly virulent in a mouse neuroinvasion model, while other strains were avirulent or significantly attenuated (d. w. c. beasley, l. li, m. t. suderman, and a. d. t. ...200515956579
early arthritis and summarize the recent literature on the association of infection with early arthritis, and to discuss the possible role of such infections with respect to the development of chronic rheumatic complications.200515956840
oncolytic viral therapy for cervical and ovarian cancer cells by sindbis virus ar339 strain.recently, the application of replication-competent viruses has been studied as anticancer agents. sindbis virus (sin) is an rna virus that belongs to the alphavirus genus in the togaviridae virus family. the ar339 strain of sin has not been reported to induce any serious disease to humans.200515958641
viroporin activity of murine hepatitis virus e protein.the viroporin activity of the e protein from murine hepatitis virus (mhv), a member of the coronaviruses, was analyzed. viroporins are a growing family of viral proteins able to enhance membrane permeability, promoting virus budding. initially, the mhv e gene was inducibly expressed in escherichia coli cells, leading to the arrest of bacterial growth, cell lysis and permeabilization to different compounds. thus, exit of labeled nucleotides from e. coli cells to the cytoplasm was apparent upon ex ...200515963987
targeted gene delivery by intravenous injection of retroviral vectors.specifically and effectively directing a therapeutic gene to its intended site of action is a critical issue for translation of basic genomics to clinical gene therapy. delivering gene therapy vectors to specific cells or tissues through intravenous injection is the most desirable method for this purpose. in 2001, we reported successful targeted gene transduction in vitro utilizing both oncoretroviral and lentiviral vectors pseudotyped with a chimeric sindbis virus envelope (zz sindbis). however ...200515970695
castanospermine, a potent inhibitor of dengue virus infection in vitro and in vivo.previous studies have suggested that alpha-glucosidase inhibitors such as castanospermine and deoxynojirimycin inhibit dengue virus type 1 infection by disrupting the folding of the structural proteins prm and e, a step crucial to viral secretion. we extend these studies by evaluating the inhibitory activity of castanospermine against a panel of clinically important flaviviruses including all four serotypes of dengue virus, yellow fever virus, and west nile virus. using in vitro assays we demons ...200515994763
infectious pancreatic necrosis virus vp5 is dispensable for virulence and persistence.infectious pancreatic necrosis virus (ipnv) is the causative agent of infectious pancreatic necrosis (ipn) disease in salmonid fish. recent studies have revealed variation in virulence between isolates of the sp serotype, associated with certain residues of the structural protein vp2. the isolates are also highly heterogenic in the coding region of the nonstructural vp5 protein. to study the involvement of this protein in the pathogenesis of disease, we generated three recombinant vp5 mutant vir ...200515994815
respiratory syncytial virus nonstructural proteins ns1 and ns2 mediate inhibition of stat2 expression and alpha/beta interferon responsiveness.respiratory syncytial virus (rsv) subverts the antiviral interferon (ifn) response, but the mechanism for this evasion was unclear. here we show that rsv preferentially inhibits ifn-alpha/beta signaling by expression of viral ns1 and ns2. thus, rsv infection or expression of recombinant ns1 and ns2 in epithelial host cells causes a marked decrease in stat2 levels and the consequent downstream ifn-alpha/beta response. similarly, ns1/ns2-deficient rsv no longer decreases stat2 levels or ifn respon ...200515994826
aif suppresses chemical stress-induced apoptosis and maintains the transformed state of tumor cells.apoptosis-inducing factor (aif) exhibits reactive oxygen species (ros)-generating nadh oxidase activity of unknown significance, which is dispensable for apoptosis. we knocked out the aif gene in two human colon carcinoma cell lines that displayed lower mitochondrial complex i oxidoreductase activity and produced less ros, but showed increased sensitivity to peroxide- or drug-induced apoptosis. aif knockout cells failed to form tumors in athymic mice or grow in soft agar. only aif with intact na ...200516001080
bax-like protein drob-1 protects neurons from expanded polyglutamine-induced toxicity in drosophila.bcl-2 family proteins regulate cell death through the mitochondrial apoptotic pathway. here, we show that the drosophila bax-like bcl-2 family protein drob-1 maintains mitochondrial function to protect cells from neurodegeneration. a pan-neuronal knockdown of drob-1 results in lower locomotor activity and a shorter lifespan in adult flies. either the rnai-mediated downregulation of drob-1 or overexpression of drob-1 antagonist buffy strongly enhances the polyglutamine-induced accumulation of ubi ...200516001086
reovirus infection of the cns enhances inos expression in areas of virus-induced injury.nitric oxide (no) has been implicated as a contributor to the host's innate defense against viral infections including those affecting the cns. reovirus infection of the cns is a classic experimental system for understanding the pathogenesis of neurotropic viral infection. infection with serotype 3 strains is associated with perturbations in various cellular signaling pathways including nf-kappab and no plays a regulatory role in many of these same pathways. we therefore examined whether no prod ...200516004984
inhibition of transcription and translation in sindbis virus-infected cells.alphaviruses are arthropod-borne viruses (arboviruses) that include a number of important human and animal pathogens. the natural transmission cycle of alphaviruses requires their presence at high concentrations in the blood of amplification hosts for efficient infection of mosquito vectors. the high-titer viremia development implies multiple rounds of infection that proceed in the background of the developing antiviral cell response aimed at blocking virus spread on an organismal level. therefo ...200516014903
characterization of immune responses induced by intramuscular vaccination with dna vaccines encoding measles virus hemagglutinin and/or fusion proteins.measles virus (mv) hemagglutinin (mv-h) and fusion (mv-f) proteins induce plaque reduction neutralizing (prn) antibodies and cell-mediated immune responses that protect against clinical measles. dna vaccines that encode mv-h and mv-f are being investigated as a new generation of measles vaccine to protect infants too young to receive currently licensed attenuated measles vaccines. however, it is unclear whether dna vaccines encoding both mv-h and mv-f act synergistically to induce stronger immun ...200516014946
essential amino acids of the hantaan virus n protein in its interaction with rna.the nucleocapsid (n) protein of hantavirus encapsidates viral genomic and antigenomic rnas. previously, deletion mapping identified a central, conserved region (amino acids 175 to 217) within the hantaan virus (htnv) n protein that interacts with a high affinity with these viral rnas (vrnas). to further define the boundaries of the rna binding domain (rbd), several peptides were synthesized and examined for the ability to bind full-length s-segment vrna. peptide 195-217 retained 94% of the vrna ...200516014963
modulation of disease, t cell responses, and measles virus clearance in monkeys vaccinated with h-encoding alphavirus replicon particles.measles remains a major worldwide problem partly because of difficulties with vaccination of young infants. new vaccine strategies need to be safe and to provide sustained protective immunity. we have developed sindbis virus replicon particles that express the measles virus (mv) hemagglutinin (sin-h) or fusion (sin-f) proteins. in mice, sin-h induced high-titered, dose-dependent, mv-neutralizing antibody after a single vaccination. sin-f, or sin-h and sin-f combined, induced somewhat lower respo ...200516037211
bcl-xl inhibits t-cell apoptosis induced by expression of sars coronavirus e protein in the absence of growth of the hallmark findings in patients suffering from sars (severe acute respiratory syndrome) is lymphopenia, which is the result of massive lymphocyte death. sars-cov (sars coronavirus), a novel coronavirus that has been etiologically associated with sars cases, is homologous with mhv (murine hepatitis coronavirus), and mhv small envelope e protein is capable of inducing apoptosis. we hypothesized that sars-cov encodes a small envelope e protein that is homologous with mhv e protein, thus in ...200516048439
early alpha/beta interferon production by myeloid dendritic cells in response to uv-inactivated virus requires viral entry and interferon regulatory factor 3 but not myd88.alpha/beta interferons (ifn-alpha/beta) are key mediators of innate immunity and important modulators of adaptive immunity. the mechanisms by which ifn-alpha/beta are induced are becoming increasingly well understood. recent studies showed that toll-like receptors 7 and 8 expressed by plasmacytoid dendritic cells (pdcs) mediate the endosomal recognition of incoming viral rna genomes, a process which requires myeloid differentiation factor 88 (myd88). here we investigate the requirements for viru ...200516051830
bovine viral diarrhea virus entry is dependent on clathrin-mediated endocytosis.cellular mechanisms of bovine viral diarrhea virus (bvdv) entry in mdbk cells were investigated. chloroquine, bafilomycin a1, or ammonium chloride inhibited bvdv infection, indicating that an acidic endosomal ph is required for bvdv entry. the tyrosine kinase inhibitor genistein partially inhibited bvdv infection at a postentry step, whereas bvdv entry was strongly inhibited by chlorpromazine or by the overexpression of a dominant-negative form of eps15, a protein essential for the formation of ...200516051874
pkcdelta-mediated regulation of flip expression in human colon cancer cells.flice-like inhibitory protein (flip), a naturally occurring caspase-inhibitory protein that lacks the critical cysteine domain necessary for catalytic activity, is a negative regulator of fas-induced apoptosis. decreased flip levels sensitize tumor cells to fas- and trail-mediated apoptosis; however, the cellular mechanisms regulating flip expression have not been defined. here, we examined the roles of the pkc and nf-kappab pathway in the regulation of flip in human colon cancers. flip mrna lev ...200616052516
a family of mammalian e3 ubiquitin ligases that contain the ubr box motif and recognize n-degrons.a subset of proteins targeted by the n-end rule pathway bear degradation signals called n-degrons, whose determinants include destabilizing n-terminal residues. our previous work identified mouse ubr1 and ubr2 as e3 ubiquitin ligases that recognize n-degrons. such e3s are called n-recognins. we report here that while double-mutant ubr1(-/-) ubr2(-/-) mice die as early embryos, the rescued ubr1(-/-) ubr2(-/-) fibroblasts still retain the n-end rule pathway, albeit of lower activity than that of w ...200516055722
viruses activate a genetically conserved cell death pathway in a unicellular organism.given the importance of apoptosis in the pathogenesis of virus infections in mammals, we investigated the possibility that unicellular organisms also respond to viral pathogens by activating programmed cell death. the m1 and m2 killer viruses of saccharomyces cerevisiae encode pore-forming toxins that were assumed to kill uninfected yeast cells by a nonprogrammed assault. however, we found that yeast persistently infected with these killer viruses induce a programmed suicide pathway in uninfecte ...200516061692
a cell-free system for the synthesis of sindbis virus subgenomic rna: importance of the concentration of the initiating ntp.we describe here an in vitro system for template-dependent initiation and synthesis of a sindbis virus (sv) subgenomic (sg) rna transcript. the critical components of this system were (1) a minus-strand promoter-template corresponding to the region of the sv genome from nt 7441 to nt 7772 (-157 to +175 relative to the sg rna transcription initiation site at nt 7598), and (2) a p15 fraction from cells infected with recombinant vaccinia viruses expressing the sv nonstructural proteins, p123 and ns ...200516085228
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