Publications

subcellular localization and immunodetection of movement proteins of olive latent virus 1.	polyclonal sera raised to escherichia coli-expressed movement proteins encoded by orf 3 (p8k) and orf 4 (p6k) of olive latent virus 1, were used for their immunodetection in infected nicotiana benthamiana plants. in subfractionated locally infected tissues 4 days post inoculation (d.p.i.) that were analysed by western blot, p8k was found in the fast-sedimenting fractions p1 and p30 containing membranous material and/or cell organelles and, likely, the fibrous structures mentioned below, but not ...	2005	15747053
molecular genetic evidence for the role of sgt1 in the intramolecular complementation of bs2 protein activity in nicotiana benthamiana.	pepper plants (capsicum annuum) containing the bs2 resistance gene are resistant to strains of xanthomonas campestris pv vesicatoria (xcv) expressing the bacterial effector protein avrbs2. avrbs2 is delivered directly to the plant cell via the type iii protein secretion system (ttss) of xcv. upon recognition of avrbs2 by plants expressing the bs2 gene, a signal transduction cascade is activated leading to a bacterial disease resistance response. here, we describe a novel pathosystem that consist ...	2005	15749757
molecular basis for the rin4 negative regulation of rps2 disease resistance.	recent studies have demonstrated that rps2, a plasma membrane-localized nucleotide binding site/leucine-rich repeat protein from arabidopsis thaliana, associates with rpm1 interacting protein 4 (rin4) and that this association functions to modulate the rps2-mediated defense pathway in response to the bacterial effector protein avrrpt2. in addition to negatively regulating rps2 activity, rin4 is also a target of avrrpt2, a cys protease and cognate bacterial effector protein of rps2. nicotiana ben ...	2005	15749765
human-derived, plant-produced monoclonal antibody for the treatment of anthrax.	the unpredictable nature of bio-terrorism compels us to develop medical countermeasures that will enable authorities to treat individuals exposed to agents such as anthrax. we report the feasibility of producing a protective, human-derived, monoclonal antibody directed against the protective antigen (pa) of bacillus anthracis in plants. this was achieved by transient expression using agroinfiltration of nicotiana benthamiana plants. the resulting antibody was able to neutralize toxin activity in ...	2005	15755575
transgenic plants regenerated from hairy roots of nicotiana benthamiana: a promising host for transient expression of foreign proteins.	hairy root cultures of nicotiana benthamiana have been obtained by co-cultivation of leaf explants with agrobacterium rhizogenes strain a4 harboring a binary vector plasmid, and transgenic nature of the obtained cultures was confirmed by pcr analysis. transgenic plants were regenerated from hairy roots. the biomass yield of transgenic plants grown in vitro was almost two-fold higher than those of wild-type n. benthamiana plants. they differed from untransformed plants by short internodes, reinfo ...	2005	16396315
optimization of agrobacterium-mediated transient assays of gene expression in lettuce, tomato and arabidopsis.	agrobacterium-mediated transient assays for gene function are increasingly being used as alternatives to genetic complementation and stable transformation. however, such assays are variable and not equally successful in different plant species. we analysed a range of genetic and physiological factors affecting transient expression following agroinfiltration, and developed a protocol for efficient and routine transient assays in several plant species. lettuce exhibited high levels of transient ex ...	2005	17173625
high-yield production of authentic human growth hormone using a plant virus-based expression system.	we describe here a high-yield transient expression system for the production of human growth hormone (hgh, or somatotropin) in transfected nicotiana benthamiana leaves. the system is based on a recently described plant virus-based modular expression vector [gleba, y., marillonnet, s. and klimyuk, v. (2004) engineering viral expression vectors for plants: the 'full virus' and the 'deconstructed virus' strategies. curr. opin. plant biol. 7, 182-188; marillonnet, s., giritch, a., gils, m., kandzia, ...	2005	17147632
virus induction of heat shock protein 70 reflects a general response to protein accumulation in the plant cytosol.	different cytoplasmically replicating rna viruses were shown to induce a specific subset of heat-inducible heat shock protein 70 (hsp70) genes in arabidopsis (arabidopsis thaliana). to identify the inducing principle, a promoterreporter system was developed for the facile analysis of differentially responding arabidopsis hsp70 genes, by infiltration into nicotiana benthamiana leaves. through transient expression of individual viral cistrons or through deletion analysis of a viral replicon, we we ...	2005	15805473
gene profiling of a compatible interaction between phytophthora infestans and solanum tuberosum suggests a role for carbonic anhydrase.	late blight of potato, caused by the oomycete pathogen phytophthora infestans, is a devastating disease that can cause the rapid death of plants. to investigate the molecular basis of this compatible interaction, potato cdna microarrays were utilized to identify genes that were differentially expressed in the host during a compatible interaction with p. infestans. of the 7,680 cdna clones represented on the array, 643 (12.9%) were differentially expressed in infected plants as compared with mock ...	2005	16167762
rapid one-step protein purification from plant material using the eight-amino acid strepii epitope.	beyond the rewards of plant genome analysis and gene identification, characterisation of protein activities, post-translational modifications and protein complex composition remains a challenge for plant biologists. ideally, methods should allow rapid isolation of proteins from plant material achieving a high degree of purity. we tested three purification strategies based on the eight-amino acid strepii, six-amino acid his(6) and 181-amino acid tandem affinity purification (tap) affinity tags fo ...	2004	15604670
analysis of thermal stress-mediated pstvd variation and biolistic inoculation of progeny of viroid "thermomutants" to tomato and brassica species.	thermal stress of pstvd-infected nicotiana benthamiana led to appearance of a broad pstvd sequence distribution, where most of mutations accumulated in the left half of the viroid's secondary structure including the "pathogenicity" domain. a similar effect had been reported for hop latent viroid [virology 287 (2001) 349]. the pool of viroid "thermomutants" progenies was transcribed into cdna and used for biolistic inoculation of raphanus sativa, where the pstvd infection was detectable by revers ...	2004	15165815
characterization and genetic diversity of potato yellow mosaic virus from the caribbean.	the begomovirus potato yellow mosaic virus (pymv) is responsible of significant yield losses in tomato in guadeloupe. four field isolates from guadeloupe were analyzed in term of their host range using three inoculation methods (mechanical, grafting and insect vector), sequences analysis of pcr fragments and phylogenetic analysis of an infectious clone, pymv-[gp]. capsicum annuum, datura stramonium, nicotiana benthamiana, n. tabacum 'xanthi nc', petunia hybrida, and solanum tuberosum were found ...	2004	14745605
biological and molecular characterization of lettuce mosaic virus from tehran province in iran.	in this study, lettuce samples having lmv infection symptoms were collected from tehran fields during 2003. samples tested for lmv infection by immuno printing. three positive samples in immuno printing collected and their characteristics were determined. in mechanical inoculation, these isolates produced symptoms on chenopodium quinoa, c. amaranticolor, gomphrena globosa, nicotiana benthamiana, lactuca sativa cv. mantilia and cv. terocadero (which contains the mol1 resistance gene and susceptib ...	2004	15756833
properties of a virus isolated from vernonia amygdalina del. in lagos, nigeria.	a previously uncharacterized virus tentatively named vernonia green vein-banding virus (vgvbv) was isolated from vernonia amygdalina del. ("bitterleaf") from lagos, nigeria. the virus was mechanically transmissible but had a narrow host range restricted to nicotiana benthamiana, chenopodium quinoa and c. amaranticolor. it was also transmissible in a non-persistent manner by myzus persicae. the virus was purified from n. benthamiana and about 750 nm long flexuous rod-shaped particles were observe ...	2004	15745049
construction and properties of a gene-silencing vector based on poplar mosaic virus (genus carlavirus).	a gene-silencing vector based on a full-length genomic clone of poplar mosaic virus (popmv) was constructed, with coat protein and movement protein genes removed, and containing instead, the coding sequence for green fluorescent protein (gfp). this paper demonstrates that the popmv-derived gene-silencing vector was able to silence gfp expression in gfp transgenic nicotiana benthamiana plants. the full-length genome of an oxford isolate of popmv (pv275) was cloned and sequenced. a full-length pop ...	2004	15664047
a geminivirus-induced gene silencing system for gene function validation in cassava.	we have constructed an african cassava mosaic virus (acmv) based gene-silencing vector as a reverse genetics tool for gene function analysis in cassava. the vector carrying a fragment from the nicotiana tabacum sulfur gene (su), encoding one unit of the chloroplast enzyme magnesium chelatase, was used to induce the silencing of the cassava orthologous gene resulting in yellow-white spots characteristic of the inhibition of su expression. this result suggests that well developed sequence database ...	2004	15630624
plant sterol biosynthesis: identification of two distinct families of sterol 4alpha-methyl oxidases.	in plants, the conversion of cycloartenol into functional phytosterols requires the removal of the two methyl groups at c-4 by an enzymic complex including a sterol 4alpha-methyl oxidase (smo). we report the cloning of candidate genes for smos in arabidopsis thaliana, belonging to two distinct families termed smo1 and smo2 and containing three and two isoforms respectively. smo1 and smo2 shared low sequence identity with each other and were orthologous to the erg25 gene from saccharomyces cerevi ...	2004	14653780
the tomato resistance protein bs4 is a predicted non-nuclear tir-nb-lrr protein that mediates defense responses to severely truncated derivatives of avrbs4 and overexpressed avrbs3.	the lycopersicon esculentum bs4 resistance (r) gene specifies recognition of xanthomonas campestris pv. vesicatoria (xcv) strains that express the cognate avrbs4 avirulence protein. bs4 was isolated by positional cloning and is predicted to encode a nucleotide-binding leucine-rich repeat (nb-lrr) protein that is homologous to tobacco n and potato y-1 resistance proteins. xcv infection tests demonstrate that bs4 confers perception of avrbs4 but not the 97% identical avrbs3 protein. however, when ...	2004	14675431
molecular chaperone hsp90 associates with resistance protein n and its signaling proteins sgt1 and rar1 to modulate an innate immune response in plants.	sgt1 and rar1 are important signaling components of resistance (r) gene-mediated plant innate immune responses. here we report that sgt1 and rar1 associate with the molecular chaperone hsp90. in addition, we show that hsp90 associates with the resistance protein n that confers resistance to tobacco mosaic virus. this suggests that hsp90-sgt1-rar1 and r proteins might exist in one complex. suppression of hsp90 in nicotiana benthamiana plants shows that it plays an important role in plant growth a ...	2004	14583611
pseudomonas syringae type iii secretion system targeting signals and novel effectors studied with a cya translocation reporter.	pseudomonas syringae pv. tomato strain dc3000 is a pathogen of tomato and arabidopsis: the hrp-hrc-encoded type iii secretion system (ttss), which injects bacterial effector proteins (primarily called hop or avr proteins) into plant cells, is required for pathogenicity. in addition to being regulated by the hrpl alternative sigma factor, most avr or hop genes encode proteins with n termini that have several characteristic features, including (i) a high percentage of ser residues, (ii) an aliphat ...	2004	14702323
arg-16 and arg-21 in the n-terminal region of the triple-gene-block protein 1 of bamboo mosaic virus are essential for virus movement.	the protein encoded by the first gene of the triple gene block (tgbp1) of potexviruses is required for movement of the viruses. it has been reported that single arg-->ala substitutions at position 11, 16 or 21 of tgbp1 of bamboo mosaic virus (bamv) eliminate its rna-binding activity, while substitutions at position 16 or 21 only affect its ntpase activity (liou et al., virology 277, 336-344, 2000). however, it remains unclear whether these arg-->ala substitutions also affect the movement of bamv ...	2004	14718640
proteomic analysis of the oxygen-evolving complex of photosystem ii under biotec stress: studies on nicotiana benthamiana infected with tobamoviruses.	we have previously shown that tobamovirus infection induces an inhibition of photosystem ii electron transport, disturbing the oxygen-evolving complex (oec). in the infected plants, the oec polypeptide pattern was modified when compared to healthy plants, the levels of the psbp and psbq extrinsic proteins being lowered to different extents. in this work we have further investigated by two-dimensional polyacrylamide gel electrophoresis (2-de) the changes on the oec protein pattern of thylakoid me ...	2004	14760711
plant virus expression systems for transient production of recombinant allergens in nicotiana benthamiana.	in recent years, several studies have demonstrated the use of autonomously replicating plant viruses as vehicles to express a variety of therapeutic molecules of pharmaceutical interest. plant virus vectors for expression of heterologous proteins in plants represent an attractive biotechnological tool to complement the conventional production of recombinant proteins in bacterial, fungal, or mammalian cells. virus vectors are advantageous when high levels of gene expression are desired within a s ...	2004	14962756
a cysteine-rich plant protein potentiates potyvirus movement through an interaction with the virus genome-linked protein vpg.	we have identified a cellular factor that interacts with the virus genome-linked proteins (vpgs) of a diverse range of potyviruses. the factor, called potyvirus vpg-interacting protein (pvip), is a plant-specific protein with homologues in all the species examined, i.e., pea, arabidopsis thaliana, and nicotiana benthamiana. the sequence of pvip does not identify a specific function, although the existence of a "phd finger" domain may implicate the protein in transcriptional control through chrom ...	2004	14963126
host-specific generation and maintenance of tomato bushy stunt virus defective interfering rnas.	the accumulation of tomato bushy stunt virus (tbsv) defective interfering rnas (dis) has been observed in several species of plants, but the involvement of host-specific processes and the functional role of dis are still poorly understood. in this study, the accumulation of dis was compared after several passages of tbsv through nicotiana benthamiana and pepper (capsicum annuum). as anticipated, passages of wild-type tbsv through n. benthamiana resulted in the accumulation of significant levels ...	2004	14964533
short deletions in nuclear targeting sequences of african cassava mosaic virus coat protein prevent geminivirus twinned particle formation.	coat proteins (cps) of geminiviruses are multifunctional proteins. using transient expression experiments, we have recently identified putative sequence motifs of african cassava mosaic virus (acmv) cp involved in nuclear import (nls) and export (nes) (virology 286 (2001) 373). here, we report on the effect of corresponding deletion mutants in the context of infecting viruses. since nls and nes may overlap with dna binding and multimerisation domains, we have investigated their effect on viral i ...	2004	14972538
interaction between a geminivirus replication protein and the plant sumoylation system.	geminiviruses are small dna viruses that replicate in nuclei of infected plant cells after accumulation of host replication machinery. tomato golden mosaic virus (tgmv) and tomato yellow leaf curl sardinia virus (tylcsv) encode a protein, repac1 (or rep), that is essential for viral replication. rep/repac1 is an oligomeric protein that binds to double-stranded dna, catalyzes cleavage and ligation of single-stranded dna, and is sufficient for host induction. it also interacts with several host pr ...	2004	14990696
cell-to-cell, but not long-distance, spread of rna silencing that is induced in individual epidermal cells.	a turnip crinkle virus (tcv)-based system was devised to discriminate cell-to-cell and systemic long-distance spread of rna silencing in plants. modified tcv-gfpdeltacp, constructed by replacing the coat protein (cp) gene with the green fluorescent protein (gfp) gene, replicated in single epidermal cells but failed to move from cell to cell in nicotiana benthamiana. mechanical inoculation of tcv-gfpdeltacp induced effective rna silencing in single epidermal cells which spread from cell to cell t ...	2004	14990735
in vitro recombinants of two nearly identical potyviral isolates express novel virulence and symptom phenotypes in plants.	six novel chimeric viruses were constructed by sequentially exchanging segments of the viral genomes between the infectious cdna clone (ppva-b11) of potato virus a (isolate pva-b11) and pufl, an almost identical infectious cdna of pva (isolate u) made in this study. the infectious in vitro transcripts of pufl and ppva-b11 caused similar severe mosaic and leaf malformation phenotypes in systemically infected leaves of nicotiana benthamiana. in contrast, one chimera induced a unique phenotype of y ...	2004	14993660
satellite panicum mosaic virus capsid protein elicits symptoms on a nonhost plant and interferes with a suppressor of virus-induced gene silencing.	the capsid protein (cp) of satellite panicum mosaic virus (spmv) has been implicated as a pathogenicity factor, inducing severe chlorosis on millet plants co-infected with spmv and its helper virus, panicum mosaic virus (pmv). in this study, we tested the effects of spmv cp on nicotiana benthamiana, a plant that does not support pmv+spmv infections. spmv cp expressed from a potato virus x (pvx) gene vector elicited necrotic lesions on n. benthamiana. pathogenicity factors often have the addition ...	2004	15000393
red clover necrotic mosaic virus replication proteins accumulate at the endoplasmic reticulum.	red clover necrotic mosaic virus (rcnmv) encodes n-terminally overlapping proteins of 27 and 88 kda (p27 and p88) known to be required for replication. green fluorescent protein (gfp) fusions were used to visualize the location of p27 and p88 within nicotiana benthamiana cells. gfp:p27 fusions localized to the endoplasmic reticulum (er), co-localized with er-targeted yellow fluorescent protein and caused membrane restructuring and proliferation. cellular fractionation of virus-inoculated n. bent ...	2004	15016550
virus survival of rna silencing without deploying protein-mediated suppression in nicotiana benthamiana.	rna silencing represents an evolutionarily conserved defence mechanism that plays a key antiviral role in protecting plants and animals against virus infection. to counterattack, plant, animal and fungal viruses produce proteins capable of suppressing rna silencing. here, we report an unprecedented phenomenon that potato virus x, a single-stranded positive rna virus, is able to survive rna silencing without deploying protein-mediated anti-silencing by revealing an unexpected symptom re-emergence ...	2004	15044003
sida micrantha mosaic is associated with a complex infection of begomoviruses different from abutilon mosaic virus.	we report on the nucleotide sequences of geminiviruses of the genus bemogovirus infecting sida micrantha schr., a common weed in brazil. for decades, the mosaic frequently associated with sida plants was considered to be caused by a brazilian strain of abutilon mosaic virus (abmv). by infection studies and sequence comparisons, we demonstrate that it is associated with a complex of at least two begomoviruses as different from abmv as most south american geminiviruses. two molecules of dna a (a1, ...	2004	15045569
functional analysis of the final steps of the 1-deoxy-d-xylulose 5-phosphate (dxp) pathway to isoprenoids in plants using virus-induced gene silencing.	isoprenoid biosynthesis in plant plastids occurs via the 1-deoxy-d-xylulose 5-phosphate (dxp) pathway. we used tobacco rattle virus (trv) to posttranscriptionally silence the expression of the last two enzymes of this pathway, the ispg-encoded (e)-4-hydroxy-3-methylbut-2-enyl diphosphate synthase (hds) and the isph-encoded isopentenyl/dimethylallyl diphosphate synthase (idds), as well as isopentenyl/dimethylallyl diphosphate isomerase (idi), the enzyme that interconverts ipp and dmapp. trv-ispg ...	2004	15064370
a geminivirus ayvv-derived shuttle vector for tobacco by2 cells.	we have developed a plant- escherichia coli pasv shuttle vector from the essential elements of the ageratum yellow vein virus (ayvv). the geminivirus vector contains the ayvv genome with the coat-protein deletion, the e. coli vector backbone of puc19, a unique cloning site and gene expression cassettes for plant selection and reporter gene activity. the replication of pasv vectors was compared in nicotiana benthamiana and n. tabacum by2 cells, and the latter were found to be suitable for long-te ...	2004	15069580
salicylic acid is part of the mi-1-mediated defense response to root-knot nematode in tomato.	the mi-1 gene of tomato confers resistance against three species of root-knot nematode in tomato (lycopersicon esculentum). transformation of tomato carrying mi-1 with a construct expressing nahg, which encodes salicylate hydroxylase, a bacterial enzyme that degrades salicylic acid (sa) to catechol, results in partial loss of resistance to root-knot nematodes. exogenous sa was toxic to roots expressing nahg but not to control roots. this toxicity is most likely due to the production of catechol ...	2004	15077667
a natural variant of a host rna-dependent rna polymerase is associated with increased susceptibility to viruses by nicotiana benthamiana.	nicotiana benthamiana often displays more intense symptoms after infection by rna viruses than do other nicotiana species. here, we examined the role of rna-dependent rna polymerases (rdrps) in n. benthamiana antiviral defense. cdnas representing only two genes encoding rdrps were identified in n. benthamiana. one rdrp was similar in sequence to sde1/sgs2 required for maintenance of transgene silencing, whereas the second, named nbrdrp1m, was >90% identical in sequence to the salicylic acid (sa) ...	2004	15079073
potato virus x-induced gene silencing in leaves and tubers of potato.	virus induced gene silencing (vigs) is increasingly used to generate transient loss-of-function assays and has potential as a powerful reverse-genetics tool in functional genomic programs as a more rapid alternative to stable transformation. a previously described potato virus x (pvx) vigs vector has been shown to trigger silencing in the permissive host nicotiana benthamiana. this paper demonstrates that a pvx-based vigs vector is also effective in triggering a vigs response in both diploid and ...	2004	15084725
phloem specific expression driven by wheat dwarf geminivirus v-sense promoter in transgenic dicotyledonous species.	wheat dwarf geminivirus (wdv) is a single-stranded dna mastrevirus. the large intergenic region (lir) of wdv contains cis-acting elements essential for the replication of the genome as well as for the bidirectional transcription of virus genes. the lir was fused to the gus (uida) reporter gene and the wdv viral sense (v-sense) promoter activity derived from the stable integration of that promoter was analysed in transgenic dicot plants. various dicot species were tested, including nicotiana taba ...	2004	15086824
expression of the b subunit of the heat-labile enterotoxin of escherichia coli in tobacco mosaic virus-infected nicotiana benthamiana plants and its characterization as mucosal immunogen and adjuvant.	we have produced biologically active recombinant (r) ltb, the nontoxic b subunit of heat-labile toxin (lt) of escherichia coli in tobacco mosaic virus (tmv)-infected nicotiana benthamiana plants. we amplified the ltb encoding sequence with its leader and introduced a hexahistidyl tag and an endoplasmic reticulum retention signal. the resulting product was ligated into a tmv-based plant viral expression vector that was used for the generation of recombinant viral rna. eighty-nine percent of n. be ...	2004	15099768
rice dwarf phytoreovirus segment s6-encoded nonstructural protein has a cell-to-cell movement function.	rice dwarf virus (rdv) is a member of the genus phytoreovirus, which is composed of viruses with segmented double-stranded rna genomes. proteins that support the intercellular movement of these viruses in the host have not been identified. microprojectile bombardment was used to determine which open reading frames (orfs) support intercellular movement of a heterologous virus. a plasmid containing an infectious clone of potato virus x (pvx) defective in cell-to-cell movement and expressing either ...	2004	15113917
silencing of subfamily i of protein phosphatase 2a catalytic subunits results in activation of plant defense responses and localized cell death.	the central importance of protein phosphorylation in plant defense responses has been demonstrated by the isolation of several disease-resistance genes that encode protein kinases. in addition, there are many reports of changes in protein phosphorylation accompanying plant responses to pathogens. in contrast, little is known about the role of protein dephosphorylation in regulating plant defenses. we report that expression of the lepp2ac1 gene, which encodes a catalytic subunit of the heterotrim ...	2004	15125764
citrx thioredoxin interacts with the tomato cf-9 resistance protein and negatively regulates defence.	to identify proteins involved in tomato cf-9 resistance protein function, a yeast two-hybrid screen was undertaken using the cytoplasmic c-terminus of cf-9 as bait. a thioredoxin-homologous clone, interacting specifically with cf-9, was identified and called citrx (cf-9-interacting thioredoxin). virus-induced gene silencing (vigs) of citrx resulted in an accelerated cf-9/avr9-triggered hypersensitive response in both tomato and nicotiana benthamiana, accompanied by enhanced accumulation of react ...	2004	15131698
a modified viral satellite dna that suppresses gene expression in plants.	dnabeta is a type of single-stranded (ss) circular satellite dna found in association with monopartite-genome begomoviruses, such as tomato yellow leaf curl china virus isolate y10 (tylccnv-y10). y10 dnabeta is required for symptom expression in plants but depends on tylccnv-y10 genomic dna (dna-a) for replication and encapsidation. when we converted dnabeta into a gene-silencing vector (modified dnabeta (dnambeta)) by replacing its c1 open-reading frame (orf) with a multiple cloning site (mcs), ...	2004	15144385
molecular characterization of nbchmp1 encoding a homolog of human chmp1 in nicotiana benthamiana.	in humans, chmp1 encodes a protein of dual function that plays a role in both modification of chromatin structure and endosomal vesicle trafficking. recently, it was found that sal1, a chmp1 homolog in maize, is important for the development of the aleurone cell layers in maize endosperm. in this study, we investigated the structure and function of a nicotiana benthamiana chmp1 homolog designated nbchmp1. nbchmp1 encodes a small protein with a bipartite nuclear localization signal at its n-termi ...	2004	15179039
a method of high frequency virus-induced gene silencing in chili pepper (capsicum annuum l. cv. bukang).	using a tobacco rattle virus (trv)-based virus-induced gene silencing (vigs) system, expression of phytogene desaturase (pds) and ribulose-1,5-bisphosphate carboxylase small-subuit (rbcs) genes was suppressed in nicotiana benthamiana and pepper plants (capsicum annuum l. cv. bukang). the silenced phenotypes of pale yellow (rbcs), and photobleached leaves (pds), were invariably obvious 2 weeks after inoculation with the trv-based vector. in a parallel experiment, the same set of genes was silence ...	2004	15179058
the dna beta satellite component associated with ageratum yellow vein disease encodes an essential pathogenicity protein (betac1).	ageratum yellow vein disease (ayvd) is caused by the geminivirus ageratum yellow vein virus (ayvv) and an associated dna beta satellite. we have mapped a dna beta transcript to a highly conserved open reading frame (betac1 orf). the most abundant transcript 5'-terminus is located 8 bases upstream of the betac1 orf putative initiation codon while the transcript terminates at multiple sites downstream from the putative termination codon. disruption of betac1 protein expression by the introduction ...	2004	15183051
geminate structures of african cassava mosaic virus.	two types of geminate structures were purified from african cassava mosaic geminivirus (acmv)-infected nicotiana benthamiana plants and analyzed by electron cryomicroscopy and image reconstruction. after cesium sulfate density gradient centrifugation, they were separated into lighter top (t) and heavier bottom (b) components. t particles comigrated with host proteins, whereas b particles were concentrated in a cesium density typical for complete virions. both particles were composed of two incom ...	2004	15194750
single-chain antibodies against a plant viral rna-dependent rna polymerase confer virus resistance.	crop loss due to viral diseases is still a major problem for agriculture today. we present a strategy to achieve virus resistance based on the expression of single-chain fv fragments (scfvs) against a conserved domain in a plant viral rna-dependent rna polymerase (rdrp), a key enzyme in virus replication. the selected scfvs inhibited complementary rna synthesis of different plant virus rdrps in vitro and virus replication in planta. moreover, the scfvs also bound to the rdrp of the distantly rel ...	2004	15195103
molecular characterization of a melon necrotic spot virus strain that overcomes the resistance in melon and nonhost plants.	resistance of melon (cucumis melo l.) to melon necrotic spot virus (mnsv) is inherited as a single recessive gene, denoted nsv. no mnsv isolates described to date (e.g., mnsv-malpha5), except for the mnsv-264 strain described here, are able to overcome the resistance conferred by nsv. analysis of protoplasts of susceptible (nsv/-) and resistant (nsv/nsv) melon cultivars inoculated with mnsv-264 or mnsv-malpha5 indicated that the resistance trait conferred by this gene is expressed at the single- ...	2004	15195949
short interfering rna accumulation correlates with host recovery in dna virus-infected hosts, and gene silencing targets specific viral sequences.	viruses are both inducers and targets of posttranscriptional gene silencing (ptgs), a natural defense mechanism in plants. here we report molecular evidence of the ability of single-stranded dna (ssdna) viruses to induce ptgs in infected plants irrespective of the severity of or recovery from the symptoms. our results reveal that five distinct species of cassava-infecting geminiviruses were capable of triggering ptgs by producing two classes of virus-specific short interfering rnas (sirnas) of 2 ...	2004	15220420
virus-induced gene silencing in solanum species.	virus-induced gene silencing (vigs) has been used routinely in nicotiana benthamiana to assess functions of candidate genes and as a way to discover new genes required for diverse pathways, especially disease resistance signalling. vigs has recently been shown to work in arabidopsis thaliana and in tomato. here, we report that vigs using the tobacco rattle virus (trv) viral vector can be used in several solanum species, although the choice of vector and experimental conditions vary depending on ...	2004	15225290
mutation of phe50 to ser50 in the 126/183-kda proteins of odontoglossum ringspot virus abolishes virus replication but can be complemented and restored by exact reversion.	sequence comparison of a non-biologically active full-length cdna clone of odontoglossum ringspot virus (orsv) pot1 with a biologically active orsv cdna clone pot2 revealed a single nucleotide change of t-->c at position 211. this resulted in the change of phe50 in ot2 to ser50 in ot1. it was not the nucleotide but the amino acid change of phe50 that was responsible for the inability of ot1 to replicate. time-course experiments showed that no minus-strand rna synthesis was detected in mutants wi ...	2004	15269387
vdnep, an elicitor from verticillium dahliae, induces cotton plant wilting.	verticillium wilt is a vascular disease of cotton. the causal fungus, verticillium dahliae, secretes elicitors in culture. we have generated approximately 1,000 5'-terminal expressed sequence tags (ests) from a cultured mycelium of v. dahliae. a number of ests were found to encode proteins harboring putative signal peptides for secretion, and their cdnas were isolated. heterologous expression led to the identification of a protein with elicitor activities. this protein, named v. dahliae necrosis ...	2004	15294839
relocalization of nuclear aly proteins to the cytoplasm by the tomato bushy stunt virus p19 pathogenicity protein.	the p19 protein of tomato bushy stunt virus (tbsv) is a multifunctional pathogenicity determinant involved in suppression of posttranscriptional gene silencing, virus movement, and symptom induction. here, we report that p19 interacts with the conserved rna-binding domain of an as yet uncharacterized family of plant aly proteins that, in animals, are involved in export of rnas from the nucleus and transcriptional coactivation. we show that the four aly proteins encoded by the arabidopsis genome ...	2004	15299117
transient coexpression of individual genes encoded by the triple gene block of potato mop-top virus reveals requirements for tgbp1 trafficking.	tgbp1, tgbp2, and tgbp3, three plant virus movement proteins encoded by the "triple gene block" (tgb), may act in concert to facilitate cell-to-cell transport of viral rna genomes. transient expression of potato mop-top virus (genus pomovirus) movement proteins was used as a model to reconstruct interactions between tgb proteins. in bombarded epidermal cells of nicotiana benthamiana, green fluorescent protein (gfp)-tgbp1 was distributed uniformly. however, in the presence of tgbp2 and tgbp3, gfp ...	2004	15305613
preparation of polyclonal antibodies of rubisco large and small subunits and their application in the functional analysis of the genes.	spinach rubisco (ribulose-1,5-bisphosphate carboxylase/oxygenase) large (rbcl) and small (rbcs) subunits were separated by sds-page, and protein amount and purity were determined by bradford assay. polyclonal antibodies against rbcl and rbcs subunit were generated in female balb/c mice and had no cross-reaction with each other. a total of 81 microg antigens were used and 0.3 ml anti-sera with titer of 1:5000 were yielded. the antibodies were also applicable to study rbcl and rbcs in tobacco plan ...	2004	15346203
virus induced gene silencing of a deficiens ortholog in nicotiana benthamiana.	traditionally, developmental studies in plant biology have suffered from the lack of a convenient means to study gene function in non-model plant species. here we show that virus-induced gene silencing (vigs) is an effective new tool to study the function of orthologs of floral homeotic genes such as deficiens (def) in non-model systems. we used a tobacco rattle virus (trv)-based vigs approach to study the function of the nicotiana benthamiana def ortholog (nbdef). silencing of nbdef in n. benth ...	2004	15356389
tête à tête of tomato yellow leaf curl virus and tomato yellow leaf curl sardinia virus in single nuclei.	since 1997 two distinct geminivirus species, tomato yellow leaf curl sardinia virus (tylcsv) and tomato yellow leaf curl virus (tylcv), have caused a similar yellow leaf curl disease in tomato, coexisted in the fields of southern spain, and very frequently doubly infected single plants. tomatoes as well as experimental test plants (e.g., nicotiana benthamiana) showed enhanced symptoms upon mixed infections under greenhouse conditions. viral dna accumulated to a similar extent in singly and doubl ...	2004	15367638
dna gyrase is involved in chloroplast nucleoid partitioning.	dna gyrase, which catalyzes topological transformation of dna, plays an essential role in replication and transcription in prokaryotes. virus-induced gene silencing of nbgyra or nbgyrb, which putatively encode dna gyrase subunits a and b, respectively, resulted in leaf yellowing phenotypes in nicotiana benthamiana. nbgyra and nbgyrb complemented the gyra and gyrb temperature-sensitive mutations of escherichia coli, respectively, which indicates that the plant and bacterial subunits are functiona ...	2004	15367714
preserved antigenicity of hiv-1 p24 produced and purified in high yields from plants inoculated with a tobacco mosaic virus (tmv)-derived vector.	production of structural proteins from foot-and-mouth disease virus (fmdv) and bovine herpes virus (bhv-1) in nicotiana benthamiana through the use of a tobacco mosaic virus-based vector (tmv-30b) has been reported previously. the development of the tmv-30b-hisc vector, a new version that adds a c-terminal histidine (his) sequence to the foreign protein expressed is described. coding sequences from the fmdv vpl protein and the core protein, p24, from a clade c hiv-1 isolate from zambia were clon ...	2004	15381357
agrodrench: a novel and effective agroinoculation method for virus-induced gene silencing in roots and diverse solanaceous species.	summary virus-induced gene silencing (vigs) is an extremely powerful tool for plant functional genomics. we used tobacco rattle virus (trv)-derived vigs vectors expressed from binary vectors within agrobacterium to induce rna silencing in plants. leaf infiltration is the most common method of agroinoculation used for vigs but this method has limitations as it is laborious for large-scale screening and some plants are difficult to infiltrate. here we have developed a novel and simple method of ag ...	2004	15447657
expression of tombusvirus open reading frames 1 and 2 is sufficient for the replication of defective interfering, but not satellite, rna.	yeast cells co-expressing the replication proteins p36 and p95 of carnation italian ringspot virus (cirv) support the rna-dependent replication of several defective interfering (di) rnas derived from either the genome of cirv or the related cymbidium ringspot virus (cymrsv), but not the replication of a satellite rna (sat rna) originally associated with cymrsv. di, but not sat rna, was replicated in yeast cells co-expressing both di and sat rna. using transgenic nicotiana benthamiana plants cons ...	2004	15448375
orf6 of tobacco mosaic virus is a determinant of viral pathogenicity in nicotiana benthamiana.	tobacco mosaic virus (tmv) contains a sixth open reading frame (orf6) that potentially encodes a 4.8 kda protein. elimination of orf6 from tmv attenuated host responses in nicotiana benthamiana without alteration in virus accumulation. furthermore, heterologous expression of tmv orf6 from either potato virus x (pvx) or tobacco rattle virus (trv) vectors enhanced the virulence of both viruses in n. benthamiana, also without effects on their accumulation. by contrast, the presence or absence of tm ...	2004	15448376
potato virus x tgbp1 induces plasmodesmata gating and moves between cells in several host species whereas cp moves only in n. benthamiana leaves.	experiments were conducted to compare the plasmodesmal transport activities of potato virus x (pvx) tgbp1 and coat protein (cp) in several plant species. microinjection experiments indicated that tgbp1 gates plasmodesmata in nicotiana tabacum leaves. these results support previous microinjection studies indicating that tgbp1 gates plasmodesmata in nicotiana benthamiana and nicotiana clevelandii leaves. to study protein movement, plasmids expressing the green fluorescent protein (gfp) gene fused ...	2004	15464839
hoppton is a pseudomonas syringae hrp (type iii secretion system) cysteine protease effector that suppresses pathogen-induced necrosis associated with both compatible and incompatible plant interactions.	pseudomonas syringae pv. tomato dc3000 causes bacterial speck disease in tomato, and it elicits the hypersensitive response (hr) in non-host plants such as nicotiana tabacum and nicotiana benthamiana. the compatible and incompatible interactions of dc3000 with tomato and nicotiana spp., respectively, result in plant cell death, but the hr cell death occurs more rapidly and is associated with effective plant defense. both interactions require the hrp (hr and pathogenicity) type iii secretion syst ...	2004	15469508
unaltered complex n-glycan profiles in nicotiana benthamiana despite drastic reduction of beta1,2- n -acetylglucosaminyltransferase i activity.	udp-glcnac:alpha3-d-mannoside beta1,2- n -acetylglucosaminyltransferase i (gnti; ec 2.4.1.101) is a golgi-resident glycosyltransferase that is essential for the processing of oligomannose to hybrid and complex n-glycans in higher eukaryotes. the cdna of nicotiana tabacum gnti has been cloned and characterised previously. to assess the influence of gnti expression levels on the formation of complex n-glycans we used posttranscriptional gene silencing to knock down the expression of gnti in the to ...	2004	15486460
a conserved secondary structure in the hypervariable region at the 5' end of bamboo mosaic virus satellite rna is functionally interchangeable.	satellite rna (satrna) associated with bamboo mosaic virus (bamv) is dependent on bamv for replication and encapsidation. molecular analyses of total rna extracted from bamboo species collected worldwide revealed that 26 out of 61 bamv isolates harbored satbamv. among them, two phylogenetically distinguishable groups, a and b, with a genetic diversity of 6.9 +/- 0.7% were identified. greatest sequence diversity occurred in the 5' untranslated region (utr) that contained one hypervariable region ...	2004	15527838
high-level expression of alternative oxidase protein sequences enhances the spread of viral vectors in resistant and susceptible plants.	the alternative oxidase (aox) is the terminal oxidase of the cyanide-resistant alternative respiratory pathway in plants and has been implicated in resistance to viruses. when tobacco mosaic virus (tmv) vectors were used to drive very high levels of expression of either aox or aox mutated in its active site (aox-e), virus spread was enhanced. this was visualized as the induction of larger hypersensitive-response lesions after inoculation onto nn-genotype tobacco than those produced by vectors be ...	2004	15557251
a dnabeta associated with tomato yellow leaf curl china virus is required for symptom induction.	we report here that all 25 isolates of tomato yellow leaf curl china virus (tylccnv) collected from tobacco, tomato, or siegesbeckia orientalis plants in different regions of yunnan province, china, were associated with dnabeta molecules. to investigate the biological role of dnabeta, full-length infectious clones of viral dna and dnabeta of tylccnv isolate y10 (tylccnv-y10) were agroinoculated into nicotiana benthamiana, nicotiana glutinosa, nicotiana. tabacum samsun (nn or nn), tomato, and pet ...	2004	15564504
gene shuffling-generated and natural variants of the tomato resistance gene cf-9 exhibit different auto-necrosis-inducing activities in nicotiana species.	tomato cf genes encode membrane-bound proteins with extracellular leucine-rich repeats, and confer resistance to the fungal tomato pathogen cladosporium fulvum, and a hypersensitive response (hr) to c. fulvum-derived race-specific elicitors. several cf genes, including cf-4 and cf-9, are members of the highly homologous hcr9 (homologues of c. fulvumresistance gene cf-9) gene family. hcr9s evolve mainly by sequence exchange between paralogues, by which novel cf genes may be generated. to mimic th ...	2004	15584959
efficient virus-induced gene silencing in roots using a modified tobacco rattle virus vector.	due to their capability of eliciting a form of posttranscriptional gene silencing (termed virus-induced gene silencing or vigs), plant viruses are increasingly used as reverse-genetics tools for functional characterization of plant genes. rna viruses have been shown to trigger silencing in a variety of host plants, including members of solanacae and arabidopsis (arabidopsis thaliana). several factors affect the silencing response, including host range and viral tropism within the plant. the work ...	2004	15591447
pseudomonas type iii effector avrpto suppresses the programmed cell death induced by two nonhost pathogens in nicotiana benthamiana and tomato.	many gram-negative bacterial pathogens rely on a type iii secretion system to deliver a number of effector proteins into the host cell. though a number of these effectors have been shown to contribute to bacterial pathogenicity, their functions remain elusive. here we report that avrpto, an effector known for its ability to interact with pto and induce pto-mediated disease resistance, inhibited the hypersensitive response (hr) induced by nonhost pathogen interactions. pseudomonas syringae pv. to ...	2004	15597738
dissecting rna silencing in protoplasts uncovers novel effects of viral suppressors on the silencing pathway at the cellular level.	short interfering rna (sirna)-mediated rna silencing plays an important role in cellular defence against viral infection and abnormal gene expression in multiple organisms. many viruses have evolved silencing suppressors for counter-defence. we have developed an rna silencing system in the protoplasts of nicotiana benthamiana to investigate the functions of viral suppressors at the cellular level. we showed that rna silencing against a green fluorescent protein (gfp) reporter gene in the protopl ...	2004	15601991
pvx-cre-mediated marker gene elimination from transgenic plants.	cre recombinase gene from bacteriophage p1 was transiently expressed by a potato virus x (pvx)-based vector in transgenic lox -target nicotiana benthamiana plants to remove the selectable marker gene. the target construct consisted of two directly oriented lox sites flanking a bar gene located between a gfp coding region and an upstream camv 35s promoter. the cre-mediated excision of intervening sequence placed the gfp coding region under the transcriptional control of the camv 35s promoter. gfp ...	2004	15604695
cotton leaf curl gezira virus-satellite dnas represent a divergent, geographically isolated nile basin lineage: predictive identification of a satdna rep-binding motif.	cotton leaf curl gezira virus (clcugv), a species of the genus begomovirus (family geminiviridae), was recently cloned from cotton, okra, and sida alba plants exhibiting leaf-curling and vein-thickening symptoms in sudan. here, we describe a previously unknown lineage of single-stranded dna satellite (satdna) molecules, which are associated with clcugv, and are required for development of characteristic disease symptoms. co-inoculation of cotton and nicotiana benthamiana plants with satdnas clon ...	2004	15826909
host-dependent differences during synergistic infection by potyviruses with potato virus x.	summary a comparative analysis of the synergistic interaction between pvx and either pvy or tev potyviruses was performed in nicotiana benthamiana and n. tabacum plants. in each pvx/potyvirus combination, doubly infected plants developed much more severe symptoms than singly infected ones. however, while pvx accumulation increased in doubly infected n. tabacum plants compared with singly infected plants, the accumulation of pvx did not vary drastically in doubly infected n. benthamiana plants wi ...	2004	20565579
beet yellows virus: the importance of being different.	summary taxonomic relationship: type member of the genus closterovirus, family closteroviridae. a member of the alphavirus-like supergroup of positive-strand rna viruses. physical properties: virions are flexuous filaments of approximately 1300 nm in length and approximately 12 nm in diameter that are made up of a approximately 15.5 kb rna and five proteins. the major capsid protein forms virion body of helical symmetry that constitutes approximately 95% of the virion length. the short virion ta ...	2003	20569367
cytosolic hsp90 and hsp70 are essential components of inf1-mediated hypersensitive response and non-host resistance to pseudomonas cichorii in nicotiana benthamiana.	summary mitogen-activated protein kinases (mapks) play pivotal roles in the signal transduction pathway of plant defence responses against pathogens. a search for mapk-interacting proteins revealed an interaction between a nicotiana benthamiana mapk, sipk (nbsipk) and cytosolic hsp90 (nbhsp90c-1) in yeast two-hybrid assay. to study the function of hsp90 in disease resistance, we silenced nbhsp90c-1 in n. benthamiana by virus-induced gene silencing (vigs) with potato virus x (pvx). nbhsp90c-1 sil ...	2003	20569398
tomato leaf curl gujarat virus, a new begomovirus species causing a severe leaf curl disease of tomato in varanasi, india.	abstract the biological and molecular properties of tomato leaf curl gujarat virus from varanasi, india (tolcgv-[var]) were characterized. tolcgv-[var] could be transmitted by grafting and through whitefly transmission in a persistent manner. the full-length genome of dna-a and dna-b of tolcgv-[var] was cloned in puc18. sequence analysis revealed that dna-a (ay190290) is 2,757 bp and dna-b (ay190291) is 2,688 bp in length. tolcgv-[var] could infect and cause symptoms in tomato, pepper, nicotiana ...	2003	18943612
interactions between geminiviruses in a naturally occurring mixture: pepper huasteco virus and pepper golden mosaic virus.	abstract pepper huasteco virus (phv) and pepper golden mosaic virus (pepgmv) are found in mixtures in many horticultural crops in mexico. this combination constitutes an interesting, naturally occurring model system to study several aspects of virus-virus interactions. possible interactions between phv and pepgmv were studied at four levels: symptom expression, gene expression, replication, and movement. in terms of symptom expression, the interaction was shown to be host-dependent because antag ...	2003	18944336
cucumovirus- and bromovirus-encoded movement functions potentiate cell-to-cell movement of tobamo- and potexviruses.	cucumber mosaic virus (cmv, a cucumovirus) and brome mosaic virus (bmv, a bromovirus) require the coat protein (cp) in addition to the 3a movement protein (mp) for cell-to-cell movement, while cowpea chlorotic mottle virus (ccmv, a bromovirus) does not. using bombardment-mediated transcomplementation assays, we investigated whether the movement functions encoded by these viruses potentiate cell-to-cell movement of movement-defective tomato mosaic virus (tomv, a tobamovirus) and potato virus x (p ...	2003	14592759
high throughput virus-induced gene silencing implicates heat shock protein 90 in plant disease resistance.	virus-induced gene silencing was used to assess the function of random nicotiana benthamiana cdnas in disease resistance. out of 4992 cdnas tested from a normalized library, there were 79 that suppressed a hypersensitive response (hr) associated with pto-mediated resistance against pseudomonas syringae. however, only six of these clones blocked the pto-mediated suppression of p.syringae growth. the three clones giving the strongest loss of pto resistance had inserts corresponding to hsp90 and al ...	2003	14592968
synthesis of l-ascorbic acid in the phloem.	although plants are the main source of vitamin c in the human diet, we still have a limited understanding of how plants synthesise l-ascorbic acid (asa) and what regulates its concentration in different plant tissues. in particular, the enormous variability in the vitamin c content of storage organs from different plants remains unexplained. possible sources of asa in plant storage organs include in situ synthesis and long-distance transport of asa synthesised in other tissues via the phloem. in ...	2003	14633288
novel system for the simultaneous analysis of geminivirus dna replication and plant interactions in nicotiana benthamiana.	the origin of replication of african cassava mosaic virus (acmv) and a gene expression vector based on potato virus x were exploited to devise an in planta system for functional analysis of the geminivirus replication-associated protein (rep) in transgenic nicotiana benthamiana line pori-2. this line contains an integrated copy of a tandem repeat of the acmv origin of replication flanking nonviral sequences that can be mobilized and replicated by rep as an episomal replicon. a rep-gfp fusion pro ...	2003	14645587
gene expression analysis of plant host-pathogen interactions by supersage.	the type iii restriction endonuclease ecop15i was used in isolating fragments of 26 bp from defined positions of cdnas. we call this substantially improved variant to the conventional serial analysis of gene expression (sage) procedure "supersage." by applying supersage to magnaporthe grisea (blast)-infected rice leaves, gene expression profiles of both the rice host and blast fungus were simultaneously monitored by making use of the fully sequenced genomes of both organisms, revealing that the ...	2003	14676315
negative-strand tospoviruses and tenuiviruses carry a gene for a suppressor of gene silencing at analogous genomic positions.	posttranscriptional silencing of a green fluorescent protein (gfp) transgene in nicotiana benthamiana plants was suppressed when these plants were infected with tomato spotted wilt virus (tswv), a plant-infecting member of the bunyaviridae: infection with tswv resulted in complete reactivation of gfp expression, similar to the case for potato virus y, but distinct from that for cucumber mosaic virus, two viruses known to carry genes encoding silencing suppressor proteins. agrobacterium-based lea ...	2003	12502849
the importance of alfalfa mosaic virus coat protein dimers in the initiation of replication.	deletion and substitution mutations affecting the oligomerization of alfalfa mosaic virus (amv) coat protein (cp) were studied in protoplasts to determine their effect on genome activation, an early step in amv replication. the cp mutants that formed dimers, cpdeltac9 and cpc-a(r)f, were highly active in initiating replication with 63-84% of wild-type (wt) cp activity. however, all mutants that did not form dimers, cpdeltac18, cpdeltac19, cpc-wfp, and cpc-w, were much less active with 19-33% of ...	2003	12504539
generation and characterisation of functional recombinant antibody fragments against rna replicase nib from plum pox virus.	a monoclonal antibody (mab 2a) able to react against the rna replicase nib from plum pox virus (ppv) was obtained and used for generating a specific scfv fragment. the vh and vl coding sequences were cloned and expressed as a fusion scfv protein to alkaline phosphatase. this fusion protein was able to recognise viral nib in both western and tissue-print elisa blots. the affinity and specificity of scfv2a for nib was similar to that of the parental mab and the region yleafy from ppv-nib was ident ...	2003	12535657
molecular analysis of the cucumber leaf spot virus genome.	full-length clones of the genome of the aureusvirus, cucumber leaf spot virus (clsv), have been constructed and infectious t7 polymerase derived synthetic transcripts have been produced. mutational analysis of the genome indicates a role for p84 in viral rna replication, the cp in systemic movement, p27 in viral cell-to-cell movement and p17 in symptom induction. a clsv mutant lacking orfs for the cp, p27 and p17 (clsv yx) was capable of replication and systemic movement in transgenic nicotiana ...	2003	12573495
dynamics of the vigs-mediated chimeric silencing of the nicotiana benthamiana chlh gene and of the tobacco mosaic virus vector.	the chlh gene, encoding for the h subunit of the magnesium chelatase enzyme, was silenced in nicotiana bentahamiana plants by virus-induced gene silencing (vigs), using tobacco mosaic virus (tmv) expression vector. strong silencing of the chlh target gene was initiated only in the apical tissues, in which the endogenous transcription level of the target gene and the level of tmv vector rna were both very high. the virus vector was also targeted by vigs, and its suppression was correlated with th ...	2003	12575743
agrosuppression: a bioassay for the hypersensitive response suited to high-throughput screening.	we describe a novel method, agrosuppression, that addresses the need for an assay of the hypersensitive response (hr) in intact plants that is rapid and adapted to high-throughput functional screening of plant and pathogen genes. the agrosuppression assay is based on inoculation of intact plants with a mixture of agrobacterium tumefaciens strains carrying (i) a binary plasmid with one or more candidate hr-inducing genes and (ii) a tumor-inducing (oncogenic) t-dna. in the absence of hr induction, ...	2003	12580277
a naturally occurring recombinant dna-a of a typical bipartite begomovirus does not require the cognate dna-b to infect nicotiana benthamiana systemically.	species of the genus begomovirus (family geminiviridae) found in the western hemisphere typically have a bipartite genome that consists of two 2.6 kb dna genomic components, dna-a and dna-b. we have identified and cloned genomic components of a new tomato-infecting begomovirus from brazil, for which the name tomato crinkle leaf yellows virus (tcrlyv) is proposed, and a dna-a variant of tomato chlorotic mottle virus (tocmv-[mg-bt1]). sequence analysis revealed that tcrlyv was most closely related ...	2003	12604824
use of green fluorescent protein to quantify the growth of colletotrichum during infection of tobacco.	to develop a quantitative assay of fungal growth inside plant tissues, strains of colletotrichum destructivum and colletotrichum orbiculare were transformed with a modified green fluorescent protein (gfp) gene fused with a glyceraldehyde-3-phosphate dehydrogenase promoter from aspergillus nidulans. transformants expressed gfp in culture and had the same growth rate and general appearance as the wild type. gfp was observed in all fungal structures during infection of leaves of nicotiana benthamia ...	2003	12609730
nicotiana benthamiana gp91phox homologs nbrboha and nbrbohb participate in h2o2 accumulation and resistance to phytophthora infestans.	active oxygen species (aos) are responsible for triggering defense responses in plants. respiratory burst oxidase homologs (rboh genes) have been implicated in aos generation. we have isolated two rboh cdnas, nbrboha and nbrbohb, from nicotiana benthamiana leaves. nbrboha was expressed constitutively at a low level and the transcripts were increased after mechanical stress of control leaf infiltration, whereas nbrbohb was induced specifically by the protein elicitor inf1 from the potato pathogen ...	2003	12615943
turnip crinkle virus coat protein mediates suppression of rna silencing in nicotiana benthamiana.	all of the protein products of turnip crinkle virus (tcv; tombusviridae, carmovirus) were tested for their ability to suppress rna silencing of a reporter gene after transient expression in agrobacterium-infiltrated nicotiana benthamiana leaves. only the capsid protein, p38, showed suppression activity, although this was not obvious when p38 was expressed as part of a tcv infection of the same tissues. when p38 was expressed from a pvx vector, symptoms with enhanced severity that correlated with ...	2003	12620795
immunodetection and fluorescent microscopy of transgenically expressed hordeivirus tgbp3 movement protein reveals its association with endoplasmic reticulum elements in close proximity to plasmodesmata.	the subcellular localization of the hydrophobic tgbp3 protein of poa semilatent virus (pslv, genus hordeivirus) was studied in transgenic plants using fluorescent microscopy to detect green fluorescent protein (gfp)-tagged protein and immunodetection with monoclonal antibodies (mabs) raised against the gfp-based fusion expressed in e. coli. in western blot analysis, mabs efficiently recognized the wild-type and gfp-fused pslv tgbp3 proteins expressed in transgenic nicotiana benthamiana, but fail ...	2003	12655101
potato mop-top virus: the coat protein-encoding rna and the gene for cysteine-rich protein are dispensable for systemic virus movement in nicotiana benthamiana.	full-length genomic cdna clones of the swedish isolate of potato mop-top virus (pmtv) were transcribed in vitro using t7 rna polymerase. the combination of rna 1, 2 and 3 synthesized in the presence of m(7)gpppg cap analogue was infectious when inoculated onto nicotiana benthamiana plants. also, the combination of rna 1 (encodes the viral replicase) with rna 3 [encodes the triple gene block proteins and a small cysteine-rich protein (crp)] was infectious and both rnas moved systemically in n. be ...	2003	12655103
molecular characterization of nbpaf encoding the alpha6 subunit of the 20s proteasome in nicotiana benthamiana.	the 26s proteasome involved in degradation of proteins covalently modified with polyubiquitin consists of the 20s proteasome and 19s regulatory complex. the nbpaf gene encoding the alpha6 subunit of the 20s proteasome was identified from nicotiana benthamiana. nbpaf exhibits high sequence homology with the corresponding genes from arabidopsis, human and yeast. the deduced amino acid sequence of nbpaf reveals that this protein contains the proteasome alpha-type subunits signature and nuclear loca ...	2003	12661772
c-terminal domain of a hevein-like protein from wasabia japonica has potent antimicrobial activity.	an antimicrobial protein, designated wjamp-1, was purified from leaves of wasabia japonica l. wjamp-1 showed antimicrobial activity against both fungi and bacteria. the deduced amino acid sequence of cdna of wjamp-1 showed 60% and 70% identity with a hevein from hevea brasiliansis and a hevein-like protein from arabidopsis thaliana, respectively. however, matured wjamp-1 lacked the hevein domain and may correspond to the c-terminal domain of hevein. southern blot analysis showed that one or two ...	2003	12668776