TitleAbstractYear(sorted ascending)
trophic activity of rabies g protein-pseudotyped equine infectious anemia viral vector mediated igf-i motor neuron gene transfer in vitro.the present study examines gene delivery to cultured motor neurons (mns) with the rabies g protein (rabg)-pseudotyped lentiviral equine infectious anemia virus (rabg.eiav) vector. rabg.eiav-mediated beta-galactosidase (rabg.eiav-lacz) gene expression in cultured mns plateaus 120 h after infection. the rate and percent of gene expression observed are titer-dependent (p < 0.001). the rat igf-i cdna sequence was then cloned into a rabg.eiav vector (rabg.eiav-igf-i) and was shown to induce igf-i exp ...200516005636
lymphocyte proliferation responses induced to broadly reactive th peptides did not protect against equine infectious anemia virus challenge.the effect of immunization with five lipopeptides, three containing t-helper (th) epitopes and two with both th and cytotoxic t-lymphocyte (ctl) epitopes, on equine infectious anemia virus (eiav) challenge was evaluated. peripheral blood mononuclear cells from eiav lipopeptide-immunized horses had significant proliferative responses to th peptides compared with those preimmunization, and the responses were attributed to significant responses to peptides gag from positions 221 to 245 (gag 221-245 ...200516085917
comparison of hiv- and eiav-based vectors on their efficiency in transducing murine and human hematopoietic repopulating cells.the use of lentiviral vectors for gene transfer into hematopoietic stem cells has raised considerable interest as these vectors can permanently integrate their genome into quiescent cells. vectors based on alternative lentiviruses would theoretically be safer than hiv-1-based vectors and could also be used in hiv-positive patients, minimizing the risk of generating replication-competent virus. here we report the use of third-generation equine infectious anemia virus (eiav)- and hiv-1-based vecto ...200516099415
antibodies and pmbc from eiav infected carrier horses recognize gp45 and p26 synthetic peptides.equine infectious anemia virus (eiav) is a lentivirus causing a persistent infection in horses characterized by recurrent febrile episodes and high levels of viremia associated with a novel antigenic strain of the virus. the virus contains two envelope glycoproteins, gp90 and gp45, and four internal proteins, p26, p15, p11 and p9. considering that the most infected horses are able to restrict eiav replication to very low levels and that gp45 and p26 contain highly conserved epitopes among lentiv ...200516105689
evaluation of high functional avidity ctl to gag epitope clusters in eiav carrier horses.cytotoxic t lymphocytes (ctl) are critical for lentivirus control including eiav. since ctl from most eiav carrier horses recognize gag epitope clusters (ec), the hypothesis that carrier horses would have high functional avidity ctl to optimal epitopes in gag ec was tested. twenty-two optimal ec epitopes were identified; two in ec1, six in ec2, and seven each in ec3 and 4. however, only five of nine horses had high functional avidity ctl (<or=11 nm) recognizing six epitopes in ec; four in relati ...200516139857
a conservative domain shared by hiv gp120 and eiav gp90: implications for hiv vaccine design.both hiv and eiav belong to the retroviridae family and lentivirus genus. two variable regions (v3 and v4) of equine infectious anemia virus (eiav) gp90 and two variable regions (v1 and v2) of hiv gp120 possibly adopt the same topology. we have studied the n-glycosylation properties and b cell linear epitope distribution profile of these two regions. our results indicated that v3 and v4 of eiav gp90 are very similar to v1 and v2 of hiv gp120. the differences between eiav virulent and vaccine str ...200516379610
optimization of equine infectious anemia derived vectors for hematopoietic cell lineage gene transfer.gene transfer into hematopoietic cells may allow correction of a variety of hematopoietic and metabolic disorders. optimized hiv-1 based lentiviral vectors have been developed for improved gene transfer and transgene expression into hematopoietic cells. however, the use of hiv-1 based vectors for human gene therapy may be limited due to ethical and biosafety issues. we report that vectors based on the non-primate equine infectious anemia virus (eiav) transduce a variety of human hematopoietic ce ...200515550928
molecular analysis of the proviral dna of equine infectious anemia virus in mules in greece.molecular analysis of the regulatory and structurally important genetic segments of equine infectious anemia virus (eiav) in mules is presented. we have previously reported clinicopathological and laboratory findings in mules infected with eiav, both naturally and after experimental inoculation. in this study the fragment coding for integrase, gp90, tat and the fusion domain of gp45 of the proviral genome from these animals was sequenced and compared with one another and with that of eiav strain ...200515567035
specificity of serum neutralizing antibodies induced by transient immune suppression of inapparent carrier ponies infected with a neutralization-resistant equine infectious anemia virus envelope has been previously reported that transient corticosteroid immune suppression of ponies experimentally infected with a highly neutralization resistant envelope variant of equine infectious anemia virus (eiav), designated eiav(deltapnd), resulted in the appearance of type-specific serum antibodies to the infecting eiav(deltapnd) virus. the current study was designed to determine if this induction of serum neutralizing antibodies was associated with changes in the specificity of envelope determ ...200515604441
differential effects of virulent and avirulent equine infectious anemia virus on macrophage cytokine expression.equine infectious anemia virus (eiav) causes rapid development of acute disease followed by recurring episodes of fever, thrombocytopenia, and viremia. most infected equid eventually bring the virus under immunological control. we recently reported the development of an equine-specific ribonuclease protection assay (rpa) to quantitate mrna levels of 10 cytokines. using this newly developed rpa, we now show significant differences in cytokine induction in equine monocyte-derived macrophages (emdm ...200515661161
modulation of human dendritic-cell function following transduction with viral vectors: implications for gene therapy.genetic modification of dendritic-cell (dc) function is an attractive approach to treat disease, either using mature dcs (mdcs) to immunize patients, or immature dcs (idcs) to induce tolerance. viral vectors are efficient at transducing dcs, and we have investigated the effect of transduction with a variety of viral vectors on the phenotype and function of dcs. adenovirus (ad), human immunodeficiency virus (hiv), equine anemia virus (eiav), and moloney murine leukemia virus (mmlv) all up-regulat ...200515671441
su proteins from virulent and avirulent eiav demonstrate distinct biological properties.biologic activity of equine infectious anemia virus (eiav) surface (su) glycoprotein was assayed in a mouse model. recombinant su from virulent eiav17 (su17), administered intraperitoneally to mouse pups, induced dose-dependent diarrheal responses similar to those reported for siv su (virology 277 (2000) 250). su17 caused fluid accumulation without histological lesions in mouse intestinal loops, induced chloride secretory currents in ussing chambers and increased inositol 1,4,5 triphosphate (ip3 ...200515708599
discerning an effective balance between equine infectious anemia virus attenuation and vaccine efficacy.among the diverse experimental vaccines evaluated in various animal lentivirus models, live attenuated vaccines have proven to be the most effective, thus providing an important model for examining critical immune correlates of protective vaccine immunity. we previously reported that an experimental live attenuated vaccine for equine infectious anemia virus (eiav), based on mutation of the viral s2 accessory gene, elicited protection from detectable infection by virulent virus challenge (f. li e ...200515708986
amino acid preferences for a critical substrate binding subsite of retroviral proteases in type 1 cleavage sites.the specificities of the proteases of 11 retroviruses representing each of the seven genera of the family retroviridae were studied using a series of oligopeptides with amino acid substitutions in the p2 position of a naturally occurring type 1 cleavage site (val-ser-gln-asn-tyr pro-ile-val-gln; the arrow indicates the site of cleavage) in human immunodeficiency virus type 1 (hiv-1). this position was previously found to be one of the most critical in determining the substrate specificity differ ...200515767422
in vivo evaluation of an eiav vector for the systemic genetic delivery of therapeutic antibodies.lentiviral-based vectors hold great promise as gene delivery vehicles for the treatment of a wide variety of diseases. we have previously reported the development of a nonprimate lentiviral vector system based on the equine infectious anaemia virus (eiav), which is able to efficiently transduce dividing and nondividing cells both in vitro and in vivo. here, we report on the application of eiav vectors for the systemic delivery of an antibody fusion protein designed for the treatment of cancer. t ...200515772687
attenuation of dna replication by hiv-1 reverse transcriptase near the central termination sequence.previous pre-steady-state kinetic studies of equine infectious anemia virus-1 (eiav) reverse transcriptase (rt) showed two effects of dna substrates containing the central termination sequence (cts) on the polymerization reaction: reduction of burst amplitude in single nucleotide addition experiments and accumulation of termination products during processive dna synthesis [berdis, a. j., stetor, s. r., le grice, s. f. j., and barkley, m. d. (2001) biochemistry 40, 12140-12149]. the present study ...200515807528
evolution of the equine infectious anemia virus long terminal repeat during the alteration of cell tropism.equine infectious anemia virus (eiav) is a lentivirus with in vivo cell tropism primarily for tissue macrophages; however, in vitro the virus can be adapted to fibroblasts and other cell types. tropism adaptation is associated with both envelope and long terminal repeat (ltr) changes, and findings strongly suggest that these regions of the genome influence cell tropism and virulence. furthermore, high levels of genetic variation have been well documented in both of these genomic regions. however ...200515827180
equine infectious anemia virus-infected dendritic cells retain antigen presentation determine if equine monocyte-derived dendritic cells (dc) were susceptible to equine infectious anemia virus (eiav) infection, ex vivo-generated dc were infected with virus in vitro. eiav antigen was detected by immunofluorescence 3 days post-infection with maximum antigen being detected on day 4, whereas there was no antigen detected in dc incubated with the same amount of heat-inactivated eiav. no cytolytic activity was observed after eiav(wsu5) infection of dc. these monocyte-derived dc we ...200515840514
genetic immunization with codon-optimized equine infectious anemia virus (eiav) surface unit (su) envelope protein gene sequences stimulates immune responses in the context of dna vaccines the native equine infectious anemia virus (eiav)-envelope gene has proven to be an extremely weak immunogen in horses probably because the rna transcripts are poorly expressed owing to an unusual codon-usage bias, the possession of multiple rna splice sites and potential adenosine-rich rna instability elements. to overcome these problems a synthetic version of sequences encoding the eiav surface unit (su) envelope glycoprotein was produced (synsu) in which the codo ...200515885929
[comparison of several viral vectors for gene therapy of corneal endothelial cells].in this paper we compare the transduction efficiency, toxicity, and safety of retroviral vectors [equine infectious anemia virus (eiav), human immunodeficiency virus-1 (hiv-1), human foamy virus (pfv] and adenovirus (ad) for potential use in gene therapy of corneal endothelial cells.200515886987
comparison of hiv-1 and eiav-based lentiviral vectors in corneal this study we compare the ability of self-inactivating human immunodeficiency virus 1 (hiv-1) and equine infectious anaemia virus (eiav)-based vectors to mediate gene transfer to rabbit and human corneas and to a murine corneal endothelial cell line. both vectors were pseudotyped with vesicular stomatitis virus-g (vsv-g) envelope and contained marker transgenes under the control of an internal cmv promoter. for specificity of action, the heterologous promoter in the eiav-vector was exchanged ...200515939034
early detection of dominant env-specific and subdominant gag-specific cd8+ lymphocytes in equine infectious anemia virus-infected horses using major histocompatibility complex class i/peptide tetrameric complexes.cytotoxic t lymphocytes (ctl) are critical for control of lentiviruses, including equine infectious anemia virus (eiav). measurement of equine ctl responses has relied on chromium-release assays, which do not allow accurate quantitation. recently, the equine mhc class i molecule 7-6, associated with the ela-a1 haplotype, was shown to present both the gag-gw12 and env-rw12 eiav ctl epitopes. in this study, 7-6/gag-gw12 and 7-6/env-rw12 mhc class i/peptide tetrameric complexes were constructed and ...200515979679
[induction of eiav-specific cellular immune response by attenuated eiav vaccine].to elucidate cellular immune protective mechanism of eiav.200616388761
influenza m2 envelope protein augments avian influenza hemagglutinin pseudotyping of lentiviral vectors.lentivirus-based gene transfer has the potential to efficiently deliver dna-based therapies into non-dividing epithelial cells of the airway for the treatment of lung diseases such as cystic fibrosis. however, significant barriers both to lung-specific gene transfer and to production of lentivirus vectors must be overcome before these vectors can be routinely used for applications to the lung. in this study, we investigated whether the ability to produce lentiviral vectors pseudotyped with fowl ...200616397505
recruitment of the adaptor protein 2 complex by the human immunodeficiency virus type 2 envelope protein is necessary for high levels of virus release.the envelope (env) protein of human immunodeficiency virus type 2 (hiv-2) and the hiv-1 vpu protein stimulate the release of retroviral particles from human cells that restrict virus production, an activity that we call the enhancement of virus release (evr). we have previously shown that two separate domains in the hiv-2 envelope protein are required for this activity: a glycine-tyrosine-x-x-hydrophobic (gyxxtheta) motif in the cytoplasmic tail and an unmapped region in the ectodomain of the pr ...200616501101
combined amino acid mutations occurring in the envelope closely correlate with pathogenicity of eiav.the chinese equine infectious anemia virus (eiav) donkey-leukocyte attenuated vaccine (dlv) provides a unique natural model system to study the attenuation mechanism and immunological control of lentivirus replication. critical consensus mutations were identified between virulent chinese eiav strains and vaccine strains. based on a full-length infectious clone of eiav vaccine strain plgfd3, two molecular clones, mfd5-4-7 and mfd7-2-11, were successfully constructed, in which 4 and 6 critical con ...200616502285
the s2 accessory gene of equine infectious anemia virus is essential for expression of disease in ponies.equine infectious anemia virus (eiav) is a macrophage-tropic lentivirus that persistently infects horses and causes a disease that is characterized by periodic episodes of fever, thrombocytopenia, and viremia. eiav encodes only four regulatory/accessory genes, (tat, rev, ttm, and s2) and is the least genetically complex of all known lentiviruses. we sought to determine the role of the eiav s2 accessory gene of eiav by introducing mutations that would prevent s2 expression on the p19/wenv17 infec ...200616503341
reversal of neurochemical changes and pain-related behavior in a model of neuropathic pain using modified lentiviral vectors expressing this study, we evaluated the possible use of lentiviral vectors in the treatment of neuropathic pain. we chose to administer gdnf-expressing vectors because of the known beneficial effect of this trophic factor in alleviation of neuropathic pain in adult rodents. lentiviral vectors expressing either gdnf or control, green fluorescent protein or beta-galactosidase, were injected unilaterally into the spinal dorsal horn 5 weeks before a spinal nerve ligation was induced (or sham surgery for the ...200616504588
the integration profile of eiav-based vectors.lentiviral vectors based on equine infectious anemia virus (eiav) stably integrate into dividing and nondividing cells such as neurons, conferring long-term expression of their transgene. the integration profile of an eiav vector was analyzed in dividing hek293t cells, alongside an hiv-1 vector as a control, and compared to a random dataset generated in silico. a multivariate regression model was generated and the influence of the following parameters on integration site selection determined: (a ...200616950499
association between the presence of serum antibodies against neospora spp. and fetal loss in equines.a study of the association between the presence of serum antibodies against neospora spp. and fetal loss was performed using serum samples of horses submitted to the laboratory for the detection of antibodies to equine herpesvirus-1 and equine infectious anemia virus. the sera submitted for equine infectious anemia testing were from horses declared healthy and those submitted for the detection of antibodies to equine herpesvirus-1 were from mares with late clinical signs of reproductive disorder ...200616962708
lentiviral vector expressing retinoic acid receptor beta2 promotes recovery of function after corticospinal tract injury in the adult rat spinal cord.spinal cord injury often results in permanent and devastating neurological deficits and disability. this is due to the limited regenerative capacity of neurones in the central nervous system (cns). we recently demonstrated that a transcription factor retinoic acid receptor beta2 (rarbeta2) promoted axonal regeneration in adult sensory neurones located peripherally. however, it is not known if rarbeta2 can promote axonal regeneration in cortical neurones of the cns. here, we demonstrate that deli ...200616984961
mutation of ymyl in the nipah virus matrix protein abrogates budding and alters subcellular localization.matrix (m) proteins reportedly direct the budding of paramyxoviruses from infected cells. in order to begin to characterize the assembly process for the highly lethal, emerging paramyxovirus nipah virus (niv), we have examined the budding of niv m. we demonstrated that expression of the niv m protein is sufficient to produce budding virus-like particles (vlps) that are physically and morphologically similar to niv. we identified in niv m a sequence, ymyl, with similarity to the ypdl late domain ...200617005661
characterization of functional domains of equine infectious anemia virus rev suggests a bipartite rna-binding domain.equine infectious anemia virus (eiav) rev is an essential regulatory protein that facilitates expression of viral mrnas encoding structural proteins and genomic rna and regulates alternative splicing of the bicistronic tat/rev mrna. eiav rev is characterized by a high rate of genetic variation in vivo, and changes in rev genotype and phenotype have been shown to coincide with changes in clinical disease. to better understand how genetic variation alters rev phenotype, we undertook deletion and m ...200616571801
eiav vector-mediated delivery of endostatin or angiostatin inhibits angiogenesis and vascular hyperpermeability in experimental cnv.we evaluated the efficacy of equine infectious anaemia virus (eiav)-based lentiviral vectors encoding endostatin (eiav.endostatin) or angiostatin (eiav.angiostatin) in inhibiting angiogenesis and vascular hyperpermeability in the laser-induced model of choroidal neovascularisation (cnv). equine infectious anaemia virus.endostatin, eiav.angiostatin or control (eiav.null) vectors were administered into the subretinal space of c57bl/6j mice. two weeks after laser injury cnv areas and the degree of ...200616572190
structural features in eiav ncp11: a lentivirus nucleocapsid protein with a short linker.lentiviral nucleocapsid proteins are a class of multifunctional proteins that play an essential role in rna packaging and viral infectivity. they contain two cx(2)cx(4)hx(4)c zinc binding motifs connected by a basic linker of variable length. the 3d structure of a 37-aa peptide corresponding to sequence 22-58 from lentiviral eiav nucleocapsid protein ncp11, complexed with zinc, has been determined by 2d (1)h nmr spectroscopy, simulated annealing, and molecular dynamics. the solution structure co ...200616634633
the crystal structure of the c-terminal domain of vps28 reveals a conserved surface required for vps20 recruitment.the endosomal sorting complex i required for transport (escrt-i) is composed of the three subunits vps23/tsg101, vps28 and vps37. escrt-i is recruited to cellular membranes during multivesicular endosome biogenesis and by enveloped viruses such as hiv-1 to mediate budding from the cell. here, we describe the crystal structure of a conserved c-terminal domain from sacharomyces cerevisiae vps28 (vps28-ctd) at 3.05 a resolution which folds independently into a four-helical bundle structure. co-expr ...200616749904
[construction of recombinant lentivirus vaccine with single round replication].to develop a safe and effective lentivirus vaccine model and provide insights into the development of other lentivirus vaccines.200616792898
[elevation of ifn-gamma transcription level in peripheral blood mononuclear cells of eiav vaccinated horses].to evaluate the relationship between the transcriptional level of ifn-gamma mrna in peripheral blood mononuclear cells (pbmc) and immune protective response driven by inoculated horses with donkey leukocyte attenuated vaccine of eiav(dlv), and to elucidate the immune mechanism of dlv.200616806002
from mice to macaques--animal models of hiv nervous system disease.lentiviral diseases of animals have been recognized for over a century, long before hiv was recognized as the cause of aids. all lentiviruses cause neurological disease and productive virus replication in the cns occurs exclusively in cells of macrophage lineage. the ability to molecularly engineer the inoculum virus, to sample the brain at many different time points from acute through terminal infection and to correlate in vivo with in vitro findings are significant advantages of animal models ...200616842082
steps to prevent outbreaks of swamp fever. 200616921663
amino acid mutations of the infectious clone from chinese eiav attenuated vaccine resulted in reversion of virulence.the chinese equine infectious anemia virus (eiav) donkey-leukocyte attenuated vaccine (dlv) provides a unique natural model system by which attenuated mechanism and immunological control of lentivirus replication may be studied. we analyzed the critical consensus mutations that occurred during the viral passages in vitro and in vivo for vaccine's preparation. based on the full-length infectious clone plgfd3 (eiav vaccine background) and according to mutations displayed during viral attenuation, ...200616202485
a replication competent lentivirus (rcl) assay for equine infectious anaemia virus (eiav)-based lentiviral vectors.lentiviral vectors are being developed to satisfy a wide range of currently unmet medical needs. vectors destined for clinical evaluation have been rendered multiply defective by deletion of all viral coding sequences and nonessential cis-acting sequences from the transfer genome. the viral envelope and accessory proteins are excluded from the production system. the vectors are produced from separate expression plasmids that are designed to minimize the potential for homologous recombination. th ...200616208418
do alix and alg-2 really control endosomes for better or for worse?alix/aip1 (alg-2-interacting protein x/apoptosis-linked-gene-2-interacting protein 1) is an adaptor protein that was first described for its capacity to bind to the calcium-binding protein alg-2 (apoptosis-linked gene 2), the expression of which seemed necessary for cell death. over-expression of truncated forms of alix blocks caspase-dependent and -independent mechanisms of cell death. numerous observations in yeast and in mammalian cells suggest that alix controls the making of and trafficking ...200616354163
stable and efficient intraocular gene transfer using pseudotyped eiav lentiviral vectors.we have developed minimal non-primate lentiviral vectors based on the equine infectious anaemia virus (eiav). we evaluated the in vivo expression profiles of these vectors delivered regionally to ocular tissues to define their potential utility in ocular gene therapy.200616299834
apparent elimination of eiav ancestral species in a long-term inapparent carrier.equine infectious anemia virus (eiav) envelope variation produces newly dominant quasispecies with each sequential disease cycle; new populations arise, and previous plasma quasispecies, including the original inoculum, become undetectable. the question remains whether these ancestral variants exist in tissue reservoirs or if the immune system eliminates quasispecies from persistent infections. to examine this, an eiav long-term inapparent carrier was immune suppressed with dexamethasone. immune ...200616226288
distribution of equine infectious anemia in horses in the north of minas gerais state, brazil.the paper examines the prevalence of equine infectious anemia (eia) in horse populations in the northern part (comprising 89 cities) of minas gerais state, brazil, from january 2002 to december 2004. data on 8,981 agar gel immunodiffusion test results from the region were used as input for a statistical and autoregressive analysis model to construct a city-level map of the distribution of eia prevalence. the following eia prevalence (p) levels were found: 49 cities with 0 < p < or = 0.5%, 26 wit ...200617037619
equine infectious anaemia in ireland: characterisation of the virus. 200617056658
a single amino acid difference within the alpha-2 domain of two naturally occurring equine mhc class i molecules alters the recognition of gag and rev epitopes by equine infectious anemia virus-specific ctl.although ctl are critical for control of lentiviruses, including equine infectious anemia virus, relatively little is known regarding the mhc class i molecules that present important epitopes to equine infectious anemia virus-specific ctl. the equine class i molecule 7-6 is associated with the equine leukocyte ag (ela)-a1 haplotype and presents the env-rw12 and gag-gw12 ctl epitopes. some ela-a1 target cells present both epitopes, whereas others are not recognized by gag-gw12-specific ctl, sugge ...200617082657
identifying interaction sites in "recalcitrant" proteins: predicted protein and rna binding sites in rev proteins of hiv-1 and eiav agree with experimental data.protein-protein and protein nucleic acid interactions are vitally important for a wide range of biological processes, including regulation of gene expression, protein synthesis, and replication and assembly of many viruses. we have developed machine learning approaches for predicting which amino acids of a protein participate in its interactions with other proteins and/or nucleic acids, using only the protein sequence as input. in this paper, we describe an application of classifiers trained on ...200617094257
serodiagnosis of equine infectious anemia by agar gel immunodiffusion and elisa using a recombinant p26 viral protein expressed in escherichia coli as antigen.we used a p26 recombinant protein (p26r) from equine infectious-anemia virus (eiav) expressed in escherichia coli as antigen to standardize an agar-gel immunodiffusion (agidp26r) test and an indirect elisa (elisap26r) for the detection of antibodies against eiav in 720 equine sera from brazil. we evaluated the tests's relative diagnostic sensitivities (relse) and relative diagnostic specificities (relsp) against a commercial agid kit (idexx, usa). we used three sera panels: panel a--196 agid-neg ...200717109980
ledgf/p75 interacts with divergent lentiviral integrases and modulates their enzymatic activity in vitro.transcriptional co-activator ledgf/p75 is the major cellular interactor of hiv-1 integrase (in), critical to efficient viral replication. in this work, a series of ins from the betaretrovirus, gammaretrovirus, deltaretrovirus, spumavirus and lentivirus retroviral genera were tested for interaction with the host factor. none of the non-lentiviral ins possessed detectable affinity for ledgf in either pull-down or yeast two-hybrid assays. in contrast, all lentiviral ins examined, including those fr ...200717158150
an alix fragment potently inhibits hiv-1 budding: characterization of binding to retroviral ypxl late domains.the retroviral structural protein, gag, contains small peptide motifs known as late domains that promote efficient virus release from the infected cell. in addition to the well characterized ptap late domain, the p6 region of hiv-1 gag contains a binding site for the host cell protein alix. to better understand the functional role of the gag/alix interaction, we overexpressed an alix fragment composed of residues 364-716 (alix 364-716) and examined the effect on release of wild type (wt) and ali ...200717158451
differential sensitivity of viruses to the antiviral activity of shiga toxin 1 a subunit.the non-toxic enzymic a subunit of shiga toxin 1 (stxa1) reduces expression and replication of the bovine retroviruses, bovine leukemia virus and bovine immunodeficiency virus (biv). here, the impact of stxa1 on representative positive and negative stranded rna viruses was compared. biv and equine infectious anemia virus were sensitive to picomolar concentrations of stxa1 while poliovirus, rhinovirus, and vesicular stomatitis virus were only marginally sensitive to nanomolar concentrations of to ...200717197049
oviduct-specific expression of two therapeutic proteins in transgenic hens.recent advances in avian transgenesis have led to the possibility of utilizing the laying hen as a production platform for the large-scale synthesis of pharmaceutical proteins. ovalbumin constitutes more than half of the protein in the white of a laid egg, and expression of the ovalbumin gene is restricted to the tubular gland cells of the oviduct. here we describe the use of lentiviral vectors to deliver transgene constructs comprising regulatory sequences from the ovalbumin gene designed to di ...200717259305
standardization and validation of an agar gel immunodiffusion test for the diagnosis of equine infectious anemia using a recombinant p26 antigen.we developed and validated an agar gel immunodiffusion test (agid) test for the diagnosis of equine infectious anemia (eia) using as antigen the p26 protein of equine infectious anemia virus (eiav) produced in the escherichia coli expression system. the developed rp26-agid test showed an excellent diagnostic relative sensitivity (100%) and specificity (100%) compared to a commercial agid assay when 1855 field serum samples were analyzed. in addition, the rp26-agid demonstrated to be a precise as ...200717292568
correlation between the induction of th1 cytokines by an attenuated equine infectious anemia virus vaccine and protection against disease progression.the equine infectious anemia virus (eiav) donkey-leukocyte attenuated vaccine (dlv) has been used to protect against equine infectious anaemia (eia) disease for several decades in china. the attenuated mechanism and immunological protective mechanisms remain to be elucidated. to identify responses that correlate with the protection against disease, we immunized horses with dlv, followed by challenge with an eiav wild-type strain ln. all vaccinated horses were asymptomatic and had a low level of ...200717325374
immune selection of equine infectious anemia virus env variants during the long-term inapparent stage of disease.the principal neutralizing domain (pnd) of equine infectious anemia virus (eiav) is located in the v3 region of su. genetic variation in the pnd is considered to play an important role in immune escape and eiav persistence; however, few studies have characterized genetic variation in su during the inapparent stage of disease. to better understand the mechanisms of virus persistence, we undertook a longitudinal study of su variation in a pony experimentally inoculated with the virulent eiav(wyo). ...200717328936
reduction of anti-hiv-1 gag immune responses during co-immunization: immune interference by the hiv-1 envelope.immunization with more than one immunogen (co-immunization) is an efficient regimen to induce immunity to multiple antigens. however, immune interference has been reported using multi-plasmid dna immunizations. hiv-1 envelope (env) and gag gene products are the predominant immunogens used in current aids vaccines, although, few studies have evaluated possible immune interference when these two antigens are co-administered. therefore, in this study, immune interference during co-inoculation was e ...200717346134
structural and biochemical studies of alix/aip1 and its role in retrovirus budding.alix/aip1 functions in enveloped virus budding, endosomal protein sorting, and many other cellular processes. retroviruses, including hiv-1, siv, and eiav, bind and recruit alix through ypx(n)l late-domain motifs (x = any residue; n = 1-3). crystal structures reveal that human alix is composed of an n-terminal bro1 domain and a central domain that is composed of two extended three-helix bundles that form elongated arms that fold back into a "v." the structures also reveal conformational flexibil ...200717350572
envelope-specific t-helper and cytotoxic t-lymphocyte responses associated with protective immunity to equine infectious anemia virus.equine infectious anemia virus (eiav) infection of horses provides a valuable model for examining the natural immunological control of lentivirus infection and disease and the mechanisms of protective and enhancing vaccine immunity. we have previously hypothesized that the eiav envelope (env) proteins gp90 and gp45 are major determinants of vaccine efficacy, and that the development of protective immunity by attenuated viral vaccines may be associated with the progressive redirection of immune r ...200717374779
real-time quantitative rt-pcr and pcr assays for a novel european field isolate of equine infectious anaemia virus based on sequence determination of the gag 2006, an outbreak of equine infectious anaemia (eia) occurred in ireland. the initial source of the outbreak is believed to have been contaminated plasma imported from italy. this paper presents the nucleotide sequence of the gag gene of the virus identified in ireland (eiav(ire)), the first for a european strain of eiav. comparison of the gag gene with north american and asian strains of the virus showed that the gag gene is less well conserved than previously believed, and that eiav strains ...200717483378
cloning and large-scale expansion of epitope-specific equine cytotoxic t lymphocytes using an anti-equine cd3 monoclonal antibody and human recombinant il-2.cytotoxic t lymphocytes are involved in controlling intracellular pathogens in many species, including horses. particularly, ctl are critical for the control of equine infectious anemia virus (eiav), a lentivirus that infects horses world-wide. in humans and animal models, ctl clones are valuable for evaluating the fine specificity of epitope recognition, and for adoptive immunotherapy against infectious and neoplastic diseases. cloned ctl would be equally useful for similar studies in the horse ...200717498813
long terminal repeat sequences from virulent and attenuated equine infectious anemia virus demonstrate distinct promoter the early 1970s, the chinese equine infectious anemia virus (eiav) vaccine, eiav(dla), was developed through successive passages of a wild-type virulent virus (eiav(l)) in donkeys in vivo and then in donkey macrophages in vitro. eiav attenuation and cell tropism adaptation are associated with changes in both envelope and long terminal repeat (ltr). however, specific ltr changes during chinese eiav attenuation have not been demonstrated. in this study, we compared ltr sequences from both virul ...200717499380
association of gag multimers with filamentous actin during equine infectious anemia virus assembly.a role for the actin cytoskeleton in retrovirus assembly has long been speculated. however, specific mechanisms by which actin facilitates the assembly process remain elusive. we previously demonstrated differential effects of experimentally modified actin dynamics on virion production of equine infectious anemia virus (eiav), a lentivirus related to hiv-1, suggesting an involvement of actin dynamics in retrovirus production. in the current study, we used bimolecular fluorescence complementation ...200717504173
cyclin box structure of the p-tefb subunit cyclin t1 derived from a fusion complex with eiav tat.the positive transcription elongation factor b (p-tefb) is an essential regulator of viral gene expression during the life cycle of human immunodeficiency virus type 1 (hiv-1). its cyclin t1 subunit forms a ternary complex with the viral transcriptional transactivator (tat) protein and the transactivation response (tar) rna element thereby activating cyclin dependent kinase 9 (cdk9), which stimulates transcription at the level of chain elongation. we report the structure of the cyclin box domain ...200717540406
both hiv- and eiav-based lentiviral vectors mediate gene delivery to pancreatic cancer cells and human pancreatic primary patient xenografts.few effective treatments for pancreatic cancer exist, especially for patients with advanced disease. gene therapy alone, or combined with current treatments, offers an alternative approach. here we examined the potential of primate and nonprimate lentivectors to mediate gene delivery to this cancer type. vsv-g pseudotyped lentivectors based on human immunodeficiency type-1 virus (hiv-1) and equine infectious anemia virus (eiav), containing the enhanced green fluorescent protein (egfp) reporter g ...200717571071
cryopreservation and lentiviral-mediated genetic modification of human primary cultured corneal endothelial determine the viability and potential usefulness of cryopreserved human primary cultured corneal endothelial cells by characterizing their morphology, gene expression, and ability for genetic modification by the lentiviral vector equine infectious anemia virus (eiav).200717591873
an investigation of equine infectious anaemia infection in the central anatolia region of this study, 162 horses, 80 donkeys and 51 mule serum samples were collected in konya city. additionally, 64 horse serum samples from ankara and 49 samples from kayseri city were included in the study. a total of 406 serum samples were examined by agar gel immunodiffusion (agid) and enzyme-linked immunosorbent assay (elisa) for antibody to equine infectious anaemia virus (eiav) and no positive result was detected.200717665759
distinct intracellular trafficking of equine infectious anemia virus and human immunodeficiency virus type 1 gag during viral assembly and budding revealed by bimolecular fluorescence complementation assays.retroviral gag polyproteins are necessary and sufficient for virus budding. numerous studies of human immunodeficiency virus type 1 (hiv-1) gag assembly and budding mechanisms have been reported, but relatively little is known about these fundamental pathways among animal lentiviruses. while there may be a general assumption that lentiviruses share common assembly mechanisms, studies of equine infectious anemia virus (eiav) have indicated alternative cellular pathways and cofactors employed amon ...200717686839
systematic epitope analysis of the p26 eiav core protein.the major core protein of equine infectious anemia virus (eiav), p26, is one of the primary immunogenic structural proteins during a persistent infection of horses and is highly conserved among antigenically variants of viral isolates. in order to investigate its immune profile in more detail for a better diagnostic, an epitope mapping was carried out by means of two libraries of overlapping peptide fragments prepared by simultaneous and parallel spps on derivatized cellulose membranes (spot syn ...200717705340
gag genetic heterogeneity of equine infectious anemia virus (eiav) in naturally infected horses in canada.gag genetic heterogeneity of equine infectious anemia virus (eiav) variants in naturally infected horses in canada was studied since very limited information is available on the variability of eiav gag sequences in public database. a phylogenetic analysis based on 414nts of gag gene sequences amplified by a nested polymerase chain reaction (pcr) revealed the distinct divergence of these variants compared to other published strains in a corresponding region. significant predicted amino acid seque ...200717767972
envelope variation as a primary determinant of lentiviral vaccine efficacy.lentiviral envelope antigenic variation and associated immune evasion are believed to present major obstacles to effective vaccine development. although this perception is widely assumed by the scientific community, there is, to date, no rigorous experimental data assessing the effect of increasing levels of lentiviral env variation on vaccine efficacy. it is our working hypothesis that env is, in fact, a primary determinant of vaccine effectiveness. we previously reported that a successful expe ...200717846425
[study on the role of cytotoxic t lymphocytes during equine infectious anemia virus infection and its epitope specificity]. 200717886727
experimental rhodococcus equi and equine infectious anemia virus dna vaccination in adult and neonatal horses: effect of il-12, dose, and route.improving the ability of dna-based vaccines to induce potent type1/th1 responses against intracellular pathogens in large outbred species is essential. rhodoccocus equi and equine infectious anemia virus (eiav) are two naturally occurring equine pathogens that also serve as important large animal models of neonatal immunity and lentiviral immune control. neonates present a unique challenge for immunization due to their diminished immunologic capabilities and apparent th2 bias. in an effort to au ...200717889970
comparative requirements for the restriction of retrovirus infection by trim5alpha and trimcyp.the restriction factors, trim5alpha in most primates and trimcyp in owl monkeys, block infection of various retroviruses soon after virus entry into the host cell. rhesus monkey trim5alpha (trim5alpha rh) inhibits human immunodeficiency virus (hiv-1) and feline immunodeficiency virus (fiv) more potently than human trim5alpha (trim5alpha hu). trimcyp restricts infection of hiv-1, simian immunodeficiency virus of african green monkeys (siv agm) and fiv. early after infection, trimcyp, like trim5al ...200717920096
western blot assay using recombinant p26 antigen for detection of equine infectious anemia virus-specific antibodies.we analyzed the performance of a single-band western blot (wb) test using recombinant p26 (rp26) capsid protein of equine infectious anemia virus. according to the results obtained, the rp26 wb test is a reliable confirmatory diagnostic tool to be used as a complementary test after an enzyme-linked immunosorbent assay or agar gel immunodiffusion test yielding doubtful results.200717959820
development of photoreceptor-specific promoters and their utility to investigate eiav lentiviral vector mediated gene transfer to photoreceptors.we wanted to investigate the ability of recombinant equine infectious anemia virus (eiav) vectors to transduce photoreceptor cells by developing a series of photoreceptor-specific promoters that drive strong gene expression in photoreceptor cells.200717963276
diagnosis of equine infectious anaemia during the 2006 outbreak in 2006 there was an outbreak of equine infectious anaemia (eia) in ireland. this paper describes the use of the diagnosis of clinical and subclinical cases of the disease. in acute cases the elisas and the immunoblot were more sensitive than the agid. in one mare, fluctuating antibody levels were observed in all the serological assays before it seroconverted by agid. viral rna and dna were detected by rt-pcr and pcr in all the tissues from the infected animals examined postmortem. the pcr detec ...200717993655
[establishment of a 293-cell line containing luciferase reporter for eiav receptor and ltr functions].to accurately and conveniently detect neutralizing antibodies and receptor binding affinities of different equine infectious anemia virus (eiav) strains, the cdna of eiav receptor, elr1, was cloned and inserted in an eukaryotic expression vector pcdna3.1(+). this recombinant plasmid was designated as pelr1. the 293 cell line was transiently transfected with pelr1 and the expression of elr1 on transfected cells was verified by western blot and indirect immunofluorescence assay (ifa). furthermore, ...200718271266
long terminal repeats are not the sole determinants of virulence for equine infectious anemia virus.the long terminal repeats (ltrs) of equine infectious anemia virus donkey leukocyte-attenuated virus (eiav-dla) were substituted with those of the wild-type eiav-l (wt eiav-l, the parent virus of eiav-dla). the resulting chimeric plasmid was designated pok-ltr dla/l. purified pok-ltr dla/l was transfected into monocyte-derived macrophage (mdm) cultures prepared from eiav-negative, heparinized whole blood from a donkey. eighth-passage cell cultures developed the typical cytopathogenic effects (cp ...200716932982
immune suppression of challenged vaccinates as a rigorous assessment of sterile protection by lentiviral vaccines.we previously reported that an experimental live-attenuated equine infectious anemia virus (eiav) vaccine, containing a mutated s2 accessory gene, provided protection from disease and detectable infection after virulent virus (eiav(pv)) challenge [li f, craigo jk, howe l, steckbeck jd, cook s, issel c, et al. a live-attenuated equine infectious anemia virus proviral vaccine with a modified s2 gene provides protection from detectable infection by intravenous virulent virus challenge of experiment ...200717023099
role of psip1/ledgf/p75 in lentiviral infectivity and integration replicate, lentiviruses such as hiv must integrate dna copies of their rna genomes into host cell chromosomes. lentiviral integration is favored in active transcription units, which allows efficient viral gene expression after integration, but the mechanisms directing integration targeting are incompletely understood. a cellular protein, psip1/ledgf/p75, binds tightly to the lentiviral-encoded integrase protein (in), and has been reported to be important for hiv infectivity and integration ta ...200718092005
molecular characterization of feline immunodeficiency virus budding.infection of domestic cats with feline immunodeficiency virus (fiv) is an important model system for studying human immunodeficiency virus type 1 (hiv-1) infection due to numerous similarities in pathogenesis induced by these two lentiviruses. however, many molecular aspects of fiv replication remain poorly understood. it is well established that retroviruses use short peptide motifs in gag, known as late domains, to usurp cellular endosomal sorting machinery and promote virus release from infec ...200818094166
solution nmr characterizations of oligomerization and dynamics of equine infectious anemia virus matrix protein and its interaction with pip2.budding of retroviruses requires the structural precursor polyprotein, gag, to target the plasma membrane through its n-terminal matrix (ma) domain. for hiv-1, the interaction between membrane signaling molecule phosphatidylinositol 4,5-diphosphate (pip2) and ma induces the exposure of myristate and promotes membrane binding. here we studied oligomerization of the naturally unmyristylated equine infectious anemia virus (eiav) ma and its interaction with pip2-c4 primarily using solution nmr spect ...200818220420
envelope determinants of equine infectious anemia virus vaccine protection and the effects of sequence variation on immune recognition.a highly effective attenuated equine infectious anemia virus (eiav) vaccine (eiav(d9)) capable of protecting 100% of horses from disease induced by a homologous env challenge strain (eiav(pv)) was recently tested in ponies to determine the level of protection against divergent env challenge strains (j. k. craigo, b. s. zhang, s. barnes, t. l. tagmyer, s. j. cook, c. j. issel, and r. c. montelaro, proc. natl. acad. sci. usa 104:15105-15110, 2007). an inverse correlation between challenge strain e ...200818234792
development and applications of non-hiv-based lentiviral vectors in neurological disorders.vectors based on non-hiv lentiviruses are opening up new approaches for the treatment of human disorders. these vectors efficiently deliver genes into many different types of cells from a broad range of species including man and the resulting gene expression is long-term. these features make them very attractive to be transformed into tools for gene therapy. hiv-1 based lentiviral vectors were initially developed, a process which provided valuable insights into the biology of these vectors allow ...200819075624
biochemical characterization of a recombinant trim5alpha protein that restricts human immunodeficiency virus type 1 replication.the rhesus monkey intrinsic immunity factor trim5alpha(rh) recognizes incoming capsids from a variety of retroviruses, including human immunodeficiency virus type 1 (hiv-1) and equine infectious anemia virus (eiav), and inhibits the accumulation of viral reverse transcripts. however, direct interactions between restricting trim5alpha proteins and retroviral capsids have not previously been demonstrated using pure recombinant proteins. to facilitate structural and mechanistic studies of retrovira ...200818799573
functional replacement of a retroviral late domain by ubiquitin fusion.retroviral gag polyprotein precursors are both necessary and sufficient for the assembly and release of virus-like particles (vlps) from infected cells. it is well established that small gag-encoded motifs, known as late domains, promote particle release by interacting with components of the cellular endosomal sorting and ubiquitination machinery. the gag proteins of a number of different retroviruses are ubiquitinated; however, the role of gag ubiquitination in particle egress remains undefined ...200818817521
equine infectious anemia virus resists the antiretroviral activity of equine apobec3 proteins through a packaging-independent mechanism.equine infectious anemia virus (eiav), uniquely among lentiviruses, does not encode a vif gene product. other lentiviruses, including human immunodeficiency virus type 1 (hiv-1), use vif to neutralize members of the apobec3 (a3) family of intrinsic immunity factors that would otherwise inhibit viral infectivity. this suggests either that equine cells infected by eiav in vivo do not express active a3 proteins or that eiav has developed a novel mechanism to avoid inhibition by equine a3 (ea3). her ...200818818324
effect of two synthetic peptides mimicking conserved regions of equine infectious anemia virus proteins gp90 and gp45 upon cytokine mrna expression.gp90 and gp45 synthetic peptides, which mimic conserved sequences of native viral proteins, are recognized by antibodies to equine infectious anemia virus (eiav) in asymptomatic carrier horses and generate humoral and cellular responses in immunized mice. cytokine mrna levels were evaluated in equine peripheral blood mononuclear cells (pbmcs) after in vitro stimulation with gp90 and gp45 with the aim of determining the cytokine profile associated with the proliferative response. stimulation inde ...200818825485
eiav vector-mediated co-delivery of endostatin and angiostatin driven by the rpe-specific vmd2 promoter inhibits choroidal neovascularization.equine infectious anemia virus (eiav) is a non-primate lentivirus that does not cause human disease. subretinal injection in mice of a recombinant eiav lentiviral vector in which lacz is driven by a cmv promoter (eiav cmv lacz) resulted in rapid and strong expression of lacz in retinal pigmented epithelial (rpe) cells and some other cells including ganglion cells resulting in x-gal within the optic nerve. substitution of the rpe-specific vmd2 promoter for the cmv promoter resulted in prolonged ( ...200818840069
a comparative analysis of viral matrix proteins using disorder predictors.a previous study (goh g.k.-m., dunker a.k., uversky v.n. (2008) protein intrinsic disorder toolbox for comparative analysis of viral proteins. bmc genomics. 9 (suppl. 2), s4) revealed that hiv matrix protein p17 possesses especially high levels of predicted intrinsic disorder (pid). in this study, we analyzed the pid patterns in matrix proteins of viruses related and unrelated to hiv-1.200818947403
[development of a cfse-based flow cytometry for evaluating eiav-stimulated proliferation of t lymphocytes].to develop a flow cytometry using (5-carboxyfluorescein diacetate succinmidyl ester, cfse) to detect the proliferation of specific t lymphocytes from equine infectious anemia virus (eiav).200818992187
structural insights into the cyclin t1-tat-tar rna transcription activation complex from eiav.the replication of many retroviruses is mediated by a transcriptional activator protein, tat, which activates rna polymerase ii at the level of transcription elongation. tat interacts with cyclin t1 of the positive transcription-elongation factor p-tefb to recruit the transactivation-response tar rna, which acts as a promoter element in the transcribed 5' end of the viral long terminal repeat. here we present the structure of the cyclin box domain of cyclin t1 in complex with the tat protein fro ...200819029897
[comparison of proviral genomes between the chinese eiav donkey leukocyte-attenuated vaccine and its parental virulent strain].the donkey leukocyte-attenuated vaccine of equine infectious anemia virus (eiav) was the first lentiviral vaccine that induced solid protection from the infection of virulent strains. to elucidate the mechanism of increased immunogenicity and attenuated virulence of the vaccine, the proviral genomic dna of an eiav vaccine strain, eiav(dlv121) was analyzed and compared with the genome of a parental virulent strain eiav(dv117). full length viral genomic dnas were amplified as two segments by la-pc ...200819226953
an outbreak of equine infectious anaemia in ireland during 2006: investigation methodology, initial source of infection, diagnosis and clinical presentation, modes of transmission and spread in the meath cluster. 200819165942
an outbreak of equine infectious anaemia in ireland during 2006: the modes of transmission and spread in the kildare cluster. 200819165943
truncation of cytoplasmic tail of eiav env increases the pathogenic necrosis.equine infectious anemia virus (eiav), like other lentiviruses, has a transmembrane glycoprotein with an unusually long cytoplasmic tail (ct). viral envelope (env) proteins having ct truncations just downstream the putative membrane-spanning domain (pmsd) are assumed to exist among all wild-type budded virions, and also in some cell-adapted strains. to determine whether ct-truncated env proteins can cause particularly deleterious effects on the env expressing cells and/or their neighboring cells ...200818294721
naturally arising point mutations in non-essential domains of equine infectious anemia virus rev alter rev-dependent nuclear-export activity.equine infectious anemia virus (eiav) exhibits a high rate of genetic variation in vivo, and results in a clinically variable disease in infected horses. in vivo populations of eiav have been characterized by the presence of distinct, genetic subpopulations of rev that differ in phenotype and fluctuate in dominance in a manner coincident with each clinical stage of disease. this study examined the specific mutations that arose in vivo and altered the phenotype. the rev protein was found to be hi ...200818343848
the interferon response inhibits hiv particle production by induction of trim22.treatment of human cells with type 1 interferons restricts hiv replication. here we report that the tripartite motif protein trim22 is a key mediator. we used transcriptional profiling to identify cellular genes that were induced by interferon treatment and identified trim22 as one of the most strongly up-regulated genes. we confirmed, as in previous studies, that trim22 over-expression inhibited hiv replication. to assess the role of trim22 expressed under natural inducing conditions, we compar ...200818389079
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