| characterization of the cassava geminivirus transcription activation protein putative nuclear localization signal. | the geminivirus transcription activation protein (trap) localizes to the nucleus and contains a putative nuclear localization signal (nls) ((28)prrrr(32)) on the n-terminus. the role of individual residues of this putative nls on nuclear localization and symptom induction was investigated using trap of east african cassava mosaic cameroon virus (eacmcv). subcellular localization was conducted using the green fluorescent protein (gfp). results showed that the proline residue at position 28 (pro-2 ... | 2009 | 19665038 |
| proteolysis of a negative regulator of innate immunity is dependent on resistance genes in tomato and nicotiana benthamiana and induced by multiple bacterial effectors. | rpm1-interacting protein 4 (rin4), a negative regulator of the basal defense response in plants, is targeted by multiple bacterial virulence effectors. we show that rin4 degradation is induced by the effector avrpto from pseudomonas syringae and that this degradation in solanaceous plants is dependent on the resistance protein, pto, a protein kinase, and prf, a nucleotide binding site-leucine-rich repeat protein. our data demonstrate overlap between two of the best-characterized pathways for rec ... | 2009 | 19671880 |
| protein mislocalization in plant cells using a gfp-binding chromobody. | a key challenge in cell biology is to directly link protein localization to function. the green fluorescent protein (gfp)-binding protein, gbp, is a 13-kda soluble protein derived from a llama heavy chain antibody that binds with high affinity to gfp as well as to some gfp variants such as yellow fluorescent protein (yfp). a gbp fusion to the red fluorescent protein (rfp), a molecule termed a chromobody, was previously used to trace in vivo the localization of various animal antigens. in this st ... | 2009 | 19686537 |
| a novel mapkk involved in cell death and defense signaling. | a high-throughput in planta overexpression screen of a nicotiana benthamiana cdna library identified a mitogen activated protein kinase kinase (mapkk), nbmkk1, as a potent inducer of hypersensitive response (hr)-like cell death. nbmkk1-mediated cell death was attenuated in plants whereby expression of nbsipk, an ortholog of tobacco sipk and arabidopsis atmpk6, was knocked down by virus-induced gene silencing (vigs), suggesting that nbmkk1 functions upstream of nbsipk. in accordance with this res ... | 2007 | 19704612 |
| distinct rna motifs mediate systemic rna trafficking. | systemic rna trafficking is emerging as a new paradigm in gene regulation at the whole plant level. the mechanisms that ensure delivery of all sorts of rnas to their respective cellular destinations remain poorly understood. several lines of research suggest that specific sequence/structural motif(s) within an rna directs its trafficking. the first motif was identified in potato spindle tuber viroid (pstvd), which mediates trafficking from bundle sheath to mesophyll in young tobacco leaves. our ... | 2008 | 19704772 |
| the colletotrichum orbiculare ssd1 mutant enhances nicotiana benthamiana basal resistance by activating a mitogen-activated protein kinase pathway. | plant basal resistance is activated by virulent pathogens in susceptible host plants. a colletotrichum orbiculare fungal mutant defective in the ssd1 gene, which regulates cell wall composition, is restricted by host basal resistance responses. here, we identified the nicotiana benthamiana signaling pathway involved in basal resistance by silencing the defense-related genes required for restricting the growth of the c. orbiculare mutant. only silencing of map kinase kinase2 or of both salicylic ... | 2009 | 19706796 |
| regions of the cf-9b disease resistance protein able to cause spontaneous necrosis in nicotiana benthamiana lie within the region controlling pathogen recognition in tomato. | the tomato cf-9 and cf-9b genes both confer resistance to the leaf mold fungus cladosporium fulvum but only cf-9 confers seedling resistance and recognizes the avirulence (avr) protein avr9 produced by c. fulvum. using domain swaps, leucine-rich repeats (lrr) 5 to 15 of cf-9 were shown to be required for cf-9-specific resistance to c. fulvum in tomato, and the entire n-terminus up to lrr15 of cf-9b was shown to be required for cf-9b-specific resistance. finer domain swaps showed that nine amino- ... | 2009 | 19737095 |
| rnai-mediated transgenic tospovirus resistance broken by intraspecies silencing suppressor protein complementation. | extension of an inverted repeat transgene cassette, containing partial nucleoprotein (n) gene sequences from four different tomato-infecting tospovirus spp. with a partial n gene sequence from the tomato strain of tomato yellow ring virus (tyrv-t), renders transgenic nicotiana benthamiana plants additionally resistant to this strain but not to the soybean strain of this tospovirus sp. (tyrv-s), both strains exhibiting 14.4% nucleotide sequence divergence in their n genes. surprisingly, coinocula ... | 2009 | 19737098 |
| complementation analysis of triple gene block of potato virus s (pvs) revealed its capability to support systemic infection and aphid transmissibility of recombinant potato virus x. | triple gene block (tgb) sequences derived from isolates of ordinary potato virus s (pvs-o) and chenopodium-systemic (pvs-cs) were analyzed. although the tgb sequences did not reveal any specific difference within the 7k protein, some specific differences within the 25k and 12k orfs were found. in order to investigate a possible functional divergence of pvs-o and pvs-cs tgb variants, these genes were propagated in chimeric potato virus x (pvx). both pvs tgb variants partly complemented pvx tgb in ... | 2009 | 19748533 |
| molecular characterization and experimental host range of an isolate of wissadula golden mosaic st. thomas virus. | partial genome segments of a begomovirus were previously amplified from wissadula amplissima exhibiting yellow-mosaic and leaf-curl symptoms in the parish of st. thomas, jamaica and this isolate assigned to a tentative begomovirus species, wissadula golden mosaic st. thomas virus. to clone the complete genome of this isolate of wissadula golden mosaic st. thomas virus, abutting primers were designed to pcr amplify its full-length dna-a and dna-b components. sequence analysis of the complete bego ... | 2009 | 19768650 |
| production and characterization of an orally immunogenic plasmodium antigen in plants using a virus-based expression system. | increasing numbers of plant-made vaccines and pharmaceuticals are entering the late stage of product development and commercialization. despite the theoretical benefits of such production, expression of parasite antigens in plants, particularly those from plasmodium, the causative parasites for malaria, have achieved only limited success. we have previously shown that stable transformation of tobacco plants with a plant-codon optimized form of the plasmodium yoelii merozoite surface protein 4/5 ... | 2009 | 19781007 |
| expression of hemagglutinin-neuraminidase glycoprotein of newcastle disease virus in agroinfiltrated nicotiana benthamiana plants. | the worldwide need for producing safer and less expensive vaccines with minor manufacture and processing requirements, together with the advances made through biotechnology, has promoted the development of efficient alternative tools to conventional vaccines. one of these is the use of plants or plant cell culture as production platforms of vaccine antigens with potential use as immunogens. we have already described the use of transgenic potato plants as immunogens against newcastle disease viru ... | 2009 | 19799942 |
| processing of mirna precursors. | plant microrna (mirna) processing requires at least two cleavage steps of respective precursors. the first cleavage step is from pri-mirna to pre-mirna, and the second cleavage step is from pre-mirna to mature mirna. using northern blot analysis, we previously showed that the rnase iii enzyme dicer-like protein 1 (dcl1) and the double-stranded rna-binding (drb) protein hyl1 are involved in the processing reactions of mirna precursors. the processing of plant mirnas differs from that of animal mi ... | 2010 | 19802599 |
| transient assays for the analysis of mirna processing and function. | transient assays provide a convenient alternative to stable transformation. for small rna analysis in plants, the most widely used method, commonly named agroinfiltration, makes use of agrobacterium tumefaciens to deliver transgenes into leaf cells of nicotiana benthamiana. compared to the generation of stably transformed plants, agroinfiltration is more rapid, and samples can be analyzed a few days after inoculation. agroinfiltration has been used successfully in many different applications, in ... | 2010 | 19802601 |
| differing requirements for actin and myosin by plant viruses for sustained intercellular movement. | the actin cytoskeleton has been implicated in the intra- and intercellular movement of a growing number of plant and animal viruses. however, the range of viruses influenced by actin for movement and the mechanism of this transport are poorly understood. here we determine the importance of microfilaments and myosins for the sustained intercellular movement of a group of rna-based plant viruses. we demonstrate that the intercellular movement of viruses from different genera [tobacco mosaic virus ... | 2009 | 19805075 |
| loss of the plant dead-box protein ise1 leads to defective mitochondria and increased cell-to-cell transport via plasmodesmata. | plants have intercellular channels, plasmodesmata (pd), that span the cell wall to enable cell-to-cell transport of micro- and macromolecules. we identified an arabidopsis thaliana embryo lethal mutant increased size exclusion limit 1 (ise1) that results in increased pd-mediated transport of fluorescent tracers. the ise1 mutants have a higher frequency of branched and twinned pd than wild-type embryos. silencing of ise1 in mature nicotiana benthamiana leaves also leads to increased pd transport, ... | 2009 | 19805190 |
| transcriptional changes and oxidative stress associated with the synergistic interaction between potato virus x and potato virus y and their relationship with symptom expression. | many virus diseases of economic importance to agriculture result from mixtures of different pathogens invading the host at a given time. this contrasts with the relatively scarce studies available on the molecular events associated with virus-host interactions in mixed infections. compared with single infections, co-infection of nicotiana benthamiana with potato virus x (pvx) and potato virus y (pvy) resulted in increased systemic symptoms (synergism) that led to necrosis of the newly emerging l ... | 2009 | 19810812 |
| identification of two rna silencing suppressors from banana bunchy top virus. | in order to suppress rna silencing, many plant and some animal viruses encode rna silencing suppressors to achieve infection. in this study, we report that b3 and b4, encoded by dna3 and dna4 of banana bunchy top virus (bbtv), exhibit rna silencing suppression activity. b3 and b4 were able to increase the transient expression of green fluorescent protein (gfp) and dramatically enhanced the pathogenicity of potato virus x (pvx) in nicotiana benthamiana. b4 was able to reverse established gene sil ... | 2009 | 19816653 |
| differential resistance to citrus psorosis virus in transgenic nicotiana benthamiana plants expressing hairpin rna derived from the coat protein and 54k protein genes. | citrus psorosis virus (cpsv), genus ophiovirus, family ophioviridae, is the causal agent of a serious disease affecting citrus trees in many countries. the viral genome consists of three ssrnas of negative polarity. post-transcriptional gene silencing (ptgs), a mechanism of plant defence against viruses, can be induced by transgenic expression of virus-derived sequences encoding hairpin rnas. since the production of transgenic citrus lines and their evaluation would take years, a herbaceous mode ... | 2009 | 19820946 |
| changes induced by the pepper mild mottle tobamovirus on the chloroplast proteome of nicotiana benthamiana. | we have analyzed the chloroplast proteome of nicotiana benthamiana using two-dimensional gel electrophoresis and mass spectrometry followed by a database search. in order to improve the resolution of the two-dimensional electrophoresis gels, we have made separate maps for the low and the high ph range. at least 200 spots were detected. we identified 72 polypeptides, some being isoforms of different multiprotein families. in addition, changes in this chloroplast proteome induced by the infection ... | 2010 | 19823941 |
| subcellular localization and in vivo interactions of the arabidopsis thaliana ethylene receptor family members. | the gaseous phytohormone ethylene regulates many developmental processes and responses to environmental conditions in higher plants. in arabidopsis thaliana, ethylene perception and initiation of signaling are mediated by a family of five receptors which are related to prokaryotic two-component sensor histidine kinases. the transient expression of fluorescence-tagged receptors in tobacco (nicotiana benthamiana) epidermal leaf cells demonstrated that all ethylene receptors are targeted to the er ... | 2008 | 19825542 |
| molecular mechanisms of generation for nitric oxide and reactive oxygen species, and role of the radical burst in plant immunity. | rapid production of nitric oxide (no) and reactive oxygen species (ros) has been implicated in the regulation of innate immunity in plants. a potato calcium-dependent protein kinase (stcdpk5) activates an nadph oxidase strboha to d by direct phosphorylation of n-terminal regions, and heterologous expression of stcdpk5 and strbohs in nicotiana benthamiana results in oxidative burst. the transgenic potato plants that carry a constitutively active stcdpk5 driven by a pathogen-inducible promoter of ... | 2009 | 19830396 |
| a leaf-based assay using interchangeable design principles to rapidly assemble multistep recombinant pathways. | the assembly of multistep recombinant pathways in stably transformed plants is a cornerstone of crops producing new products yet can be a laborious and time-consuming process. any heterologous expression platform capable of providing a rapid estimation of the functional assembly of an entire pathway would guide the design of such transgenic traits. in this study, we use a nicotiana benthamiana transient leaf expression system to simultaneously express five genes, from five independent t(dna) bin ... | 2009 | 19843252 |
| nucleolin/nsr1p binds to the 3' noncoding region of the tombusvirus rna and inhibits replication. | previous genome-wide screens identified >100 host genes affecting tombusvirus replication using yeast model host. one of those factors was nsr1p (nucleolin), which is an abundant rna-binding shuttle protein involved in rrna maturation and ribosome assembly. we find that overexpression of nsr1p in yeast or in nicotiana benthamiana inhibited the accumulation of tombusvirus rna by approximately 10-fold. regulated overexpression of nsr1p revealed that nsr1p should be present at the beginning of vira ... | 2010 | 19861225 |
| high-level expression of camelid nanobodies in nicotiana benthamiana. | nanobodies (or vhhs) are single-domain antigen-binding fragments derived from camelid heavy chain-only antibodies. their small size, monomeric behaviour, high stability and solubility, and ability to bind epitopes not accessible to conventional antibodies make them especially suitable for many therapeutic and biotechnological applications. here we describe high-level expression, in nicotiana benthamiana, of three versions of an anti-hen egg white lysozyme (hewl) nanobody which include the origin ... | 2010 | 19862637 |
| evidence for similarity-assisted recombination and predicted stem-loop structure determinant in potato virus x rna recombination. | virus rna recombination, one of the main factors for genetic variability and evolution, is thought to be based on different mechanisms. here, the recently described in vivo potato virus x (pvx) recombination assay [draghici, h.-k. & varrelmann, m. (2009). j virol 83, 7761-7769] was applied to characterize structural parameters of recombination. the assay uses an agrobacterium-mediated expression system incorporating a pvx green fluorescent protein (gfp)-labelled full-length clone. the clone cont ... | 2010 | 19864501 |
| isolation and characterization of aawrky1, an artemisia annua transcription factor that regulates the amorpha-4,11-diene synthase gene, a key gene of artemisinin biosynthesis. | amorpha-4,11-diene synthase (ads) of artemisia annua catalyzes the conversion of farnesyl diphosphate into amorpha-4,11-diene, the first committed step in the biosynthesis of the antimalarial drug artemisinin. the promoters of ads contain two reverse-oriented ttgacc w-box cis-acting elements, which are the proposed binding sites of wrky transcription factors. a full-length cdna (aawrky1) was isolated from a cdna library of the glandular secretory trichomes (gsts) in which artemisinin is synthesi ... | 2009 | 19880398 |
| plastocyanin transit peptide interacts with potato virus x coat protein, while silencing of plastocyanin reduces coat protein accumulation in chloroplasts and symptom severity in host plants. | potato virus x coat protein (pvxcp) is, through communication with host proteins, involved in processes such as virus movement and symptom development. here, we report that pvxcp also interacts with the precursor of plastocyanin, a protein involved in photosynthesis, both in vitro and in vivo. yeast two-hybrid analysis indicated that pvxcp interacted with only the plastocyanin transit peptide. in subsequent bimolecular fluorescence complementation assays, both proteins were collocated within chl ... | 2009 | 19888818 |
| the nuclear interactor pyl8/rcar3 of fagus sylvatica fspp2c1 is a positive regulator of abscisic acid signaling in seeds and stress. | the functional protein phosphatase type 2c from beechnut (fagus sylvatica; fspp2c1) was a negative regulator of abscisic acid (aba) signaling in seeds. in this report, to get deeper insight on fspp2c1 function, we aim to identify pp2c-interacting partners. two closely related members (pyl8/rcar3 and pyl7/rcar2) of the arabidopsis (arabidopsis thaliana) betv i family were shown to bind fspp2c1 in a yeast two-hybrid screening and in an aba-independent manner. by transient expression of fspp2c1 and ... | 2010 | 19889877 |
| the rice yellow mottle virus p1 protein exhibits dual functions to suppress and activate gene silencing. | in plants rna silencing is a host defense mechanism against viral infection, in which double-strand rna is processed into 21-24-nt short interfering rna (sirna). silencing spreads from cell to cell and systemically through a sequence-specific signal to limit the propagation of the virus. to counteract this defense mechanism, viruses encode suppressors of silencing. the p1 protein encoded by the rice yellow mottle virus (rymv) displays suppression activity with variable efficiency, according to t ... | 2010 | 19891703 |
| tomato leaf curl java virus v2 protein is a determinant of virulence, hypersensitive response and suppression of posttranscriptional gene silencing. | we previously identified the tomato leaf curl java virus-a (tolcjv-a[id]) from southeast asia as a new member of the emerging group of monopartite begomoviruses that require a betasatellite component for symptom induction. in this study, the role of v2 in viral pathogenesis and posttranscriptional gene silencing (ptgs) was studied. our results showed v2 of tolcjv-a[id] elicits a reaction resembling the hypersensitive response (hr) associated with the induction of necrosis and a systemic burst of ... | 2010 | 19896687 |
| potatovirus x and tobacco mosaic virus-based vectors compatible with the gateway cloning system. | virus-based expression vectors are important tools for high-level production of foreign proteins and for gene function analysis through virus induced gene silencing. to exploit further their advantages as fast, high yield replicons, a set of vectors was produced by converting and adapting potato virus x (pvx) and tobacco mosaic virus (tmv)-based vectors to allow easy cloning of foreign sequences by the gateway cloning system. target genes were cloned efficiently by recombination and successfully ... | 2010 | 19903495 |
| satellite panicum mosaic virus coat protein enhances the performance of plant virus gene vectors. | the coat protein of satellite panicum mosaic virus (spcp) is known to effectively protect its cognate rna from deleterious events, and here, we tested its stabilizing potential for heterologous virus-based gene vectors in planta. in support of this, a potato virus x (pvx) vector carrying the spmv capsid protein (pvx-spcp) gene was stable for at least three serial systemic passages through nicotiana benthamiana. to test the effect of spcp in trans, pvx-spcp was co-inoculated onto n. benthamiana t ... | 2010 | 19903565 |
| evolutionary trajectory of turnip mosaic virus populations adapting to a new host. | little is known about how some plant viruses establish successful cross-species transmission whilst others do not; the genetic basis for adaptation is largely unknown. this study investigated the genetic changes that occurred using the progeny of an infectious clone, p35tunos, derived from the turnip mosaic virus (tumv) uk 1 isolate, which has a brassica host type, but rarely infects raphanus systemically and then only asymptomatically. the genetic trajectory leading to viral adaptation was stud ... | 2010 | 19906944 |
| a virus-induced gene silencing screen identifies a role for thylakoid formation1 in pseudomonas syringae pv tomato symptom development in tomato and arabidopsis. | pseudomonas syringae pv tomato dc3000 (pst dc3000), which causes disease in tomato (solanum lycopersicum) and arabidopsis (arabidopsis thaliana), produces coronatine (cor), a non-host-specific phytotoxin. cor, which functions as a jasmonate mimic, is required for full virulence of pst dc3000 and for the induction of chlorosis in host plants. previous genetic screens based on insensitivity to cor and/or methyl jasmonate identified several potential targets for cor and methyl jasmonate. in this st ... | 2010 | 19915014 |
| production of biologically active human myelocytokines in plants. | an effective system for expression of human granulocyte and granulocyte macrophage colony-stimulating factors (hg-csf and hgm-csf) in nicotiana benthamiana plants was developed using viral vector based on tobacco mosaic virus infecting cruciferous plants. the genes of target proteins were cloned into the viral vector driven by actin promoter of arabidopsis thaliana. the expression vectors were delivered into plant cells by agroinjection. maximal synthesis rate was detected 5 days after injection ... | 2009 | 19916932 |
| high level protein expression in plants through the use of a novel autonomously replicating geminivirus shuttle vector. | we constructed a novel autonomously replicating gene expression shuttle vector, with the aim of developing a system for transiently expressing proteins at levels useful for commercial production of vaccines and other proteins in plants. the vector, pric, is based on the mild strain of the geminivirus bean yellow dwarf virus (beydv-m) and is replicationally released into plant cells from a recombinant agrobacterium tumefaciens ti plasmid. pric differs from most other geminivirus-based vectors in ... | 2010 | 19929900 |
| high-level hiv-1 nef transient expression in nicotiana benthamiana using the p19 gene silencing suppressor protein of artichoke mottled crinckle virus. | in recent years, different hiv antigens have been successfully expressed in plants by either stable transformation or transient expression systems. among hiv proteins, nef is considered a promising target for the formulation of a multi-component vaccine due to its implication in the first steps of viral infection. attempts to express nef as a single protein product (not fused to a stabilizing protein) in transgenic plants resulted in disappointingly low yields (about 0.5% of total soluble protei ... | 2009 | 19930574 |
| characterization of tomato yellow vein streak virus, a begomovirus from brazil. | tomato yellow vein streak virus (toyvsv) is a tentative begomovirus (family geminiviridae) species that seriously affects tomato and potato production in brazil. here, we have determined the genomic and biological characteristics of a toyvsv isolate (ba3) from a potato plant sampled in rio grande do sul state, brazil. the dna-a nucleotide sequence of ba3 and another previously reported toyvsv isolate share 89.7% sequence identity. these toyvsv isolates should be classified as a new species in th ... | 2010 | 19937269 |
| the tobacco etch virus p3 protein forms mobile inclusions via the early secretory pathway and traffics along actin microfilaments. | plant potyviruses encode two membrane proteins, 6k and p3. the 6k protein has been shown to induce virus replication vesicles. however, the function of p3 remains unclear. in this study, subcellular localization of the tobacco etch virus (tev) p3 protein was investigated in nicotiana benthamiana leaf cells. the tev p3 protein localized on the endoplasmic reticulum (er) membrane and formed punctate inclusions in association with the golgi apparatus. the trafficking of p3 to the golgi was mediated ... | 2010 | 19945728 |
| p0 proteins of european beet-infecting poleroviruses display variable rna silencing suppression activity. | post-transcriptional gene silencing (ptgs), or rna silencing, is one of the key mechanisms of antiviral defence used by plants. to counter this defence response, viruses produce suppressor proteins that are able to inhibit the ptgs pathway or to interfere with some of its function. the aim of this study was to evaluate the rna silencing suppressor (rss) activity of p0 proteins from selected european isolates of the beet-infecting poleroviruses beet chlorosis virus (bchv) and beet mild yellowing ... | 2010 | 19955562 |
| myosin xi is required for actin-associated movement of plastid stromules. | stromules are highly dynamic stroma-filled tubules extending from the surface of plastids and occasionally interconnecting individual plastids, allowing the movement of complex biological molecules between the interconnected plastids. experiments with inhibitors of cytoskeleton assembly have indicated the involvement of an actin-based system in stromule movement. however, the motor protein associated with the system had not been identified. here, we present direct evidence that myosin xi is invo ... | 2009 | 19995729 |
| a myosin xi tail domain homologous to the yeast myosin vacuole-binding domain interacts with plastids and stromules in nicotiana benthamiana. | the actin cytoskeleton plays a role in mobility of many different organelles in plant cells, including chloroplasts, mitochondria, golgi, and peroxisomes. while progress has been made in identifying the myosin motors involved in trafficking of various plant organelles, not all of the cargoes mobilized by different members of the myosin xi family have yet been identified. the involvement of myosins in chloroplast positioning and mitochondrial movement was demonstrated by expression of a virus-ind ... | 2009 | 19995734 |
| significance of eukaryotic translation elongation factor 1a in tobacco mosaic virus infection. | eukaryotic translation elongation factor 1a (eef1a) has been shown to interact with both the viral rna-dependent rna polymerase and the 3'-terminal genomic rna of tobacco mosaic virus (tmv). in this study, we demonstrated that the down-regulation of eef1a mrna levels by virus-induced gene silencing using potato virus x vector dramatically reduced the accumulation of tmv rna and the spread of tmv infection. the translation activity of the eef1a-silenced nicotiana benthamiana leaves was not severe ... | 2010 | 20012112 |
| an efficient system to detect protein ubiquitination by agroinfiltration in nicotiana benthamiana. | the ubiquitination proteasome pathway has been demonstrated to regulate all plant developmental and signaling processes. e3 ligase/substrate-specific interactions and ubiquitination play important roles in this pathway. however, due to technical limitations only a few instances of e3 ligase-substrate binding and protein ubiquitination in plants have been directly evidenced. an efficient in vivo and in vitro ubiquitination assay was developed for analysis of protein ubiquitination reactions by ag ... | 2010 | 20015064 |
| rna-dependent rna polymerase 6 delays accumulation and precludes meristem invasion of a viroid that replicates in the nucleus. | the detection of viroid-derived small rnas (vd-srnas) similar to the small interfering rnas (sirnas, 21 to 24 nucleotides [nt]) in plants infected by nuclear-replicating members of the family pospiviroidae (type species, potato spindle tuber viroid [pstvd]) indicates that they are inducers and targets of the rna-silencing machinery of their hosts. rna-dependent rna polymerase 6 (rdr6) catalyzes an amplification circuit producing the double-stranded precursors of secondary sirnas. recently, the r ... | 2010 | 20015979 |
| inhibition of sterol biosynthesis reduces tombusvirus replication in yeast and plants. | the replication of plus-strand rna viruses depends on subcellular membranes. recent genome-wide screens have revealed that the sterol biosynthesis genes erg25 and erg4 affected the replication of tomato bushy stunt virus (tbsv) in a yeast model host. to further our understanding of the role of sterols in tbsv replication, we demonstrate that the downregulation of erg25 or the inhibition of the activity of erg25p with an inhibitor (6-amino-2-n-pentylthiobenzothiazole; apb) leads to a 3- to 5-fold ... | 2010 | 20015981 |
| hydrophobin fusions for high-level transient protein expression and purification in nicotiana benthamiana. | insufficient accumulation levels of recombinant proteins in plants and the lack of efficient purification methods for recovering these valuable proteins have hindered the development of plant biotechnology applications. hydrophobins are small and surface-active proteins derived from filamentous fungi that can be easily purified by a surfactant-based aqueous two-phase system. in this study, the hydrophobin hfbi sequence from trichoderma reesei was fused to green fluorescent protein (gfp) and tran ... | 2010 | 20018596 |
| nicotiana tabacum eil2 directly regulates expression of at least one tobacco gene induced by sulphur starvation. | sulphur deficiency severely affects plant growth and their agricultural productivity leading to diverse changes in development and metabolisms. molecular mechanisms regulating gene expression under low sulphur conditions remain largely unknown. atslim1, a member of the ein3-like (eil) family was reported to be a central transcriptional regulator of the plant sulphur response, however, no direct interaction of this protein with any sulphur-responsive promoters was demonstrated. the focus of this ... | 2010 | 20018902 |
| pathogenicity of alternanthera mosaic virus is affected by determinants in rna-dependent rna polymerase and by reduced efficacy of silencing suppression in a movement-competent tgb1. | four biologically active cdna clones were derived from the alternanthera mosaic virus (altmv; genus potexvirus) isolate, altmv-sp, which differ in symptoms in infected nicotiana benthamiana plants. two clones induced necrosis and plant death; a mixture of all four clones induced milder symptoms than altmv-sp. replication of all clones was enhanced by a minimum of fourfold at 15 degrees c. a mixture of clones 4-7 (severe) and 3-1 (mild) was indistinguishable from altmv-sp, but the ratio of 4-7 to ... | 2010 | 20019006 |
| cloning and molecular analysis of hlbzip1 and hlbzip2 transcription factors putatively involved in the regulation of the lupulin metabolome in hop (humulus lupulus l.). | hop (humulus lupulus l.), the essential source of beer flavor is of interest from a medicinal perspective in view of its high content in health-beneficial terpenophenolics including prenylflavonoids. the dissection of biosynthetic pathway(s) of these compounds in lupulin glands, as well as its regulation by transcription factors (tfs), is important for efficient biotechnological manipulation of the hop metabolome. tfs of the bzip class were preselected from the hop transcriptome using a cdna-afl ... | 2010 | 20028133 |
| analysis of an autoproteolytic activity of rice yellow mottle virus silencing suppressor p1. | ectopically expressed rice yellow mottle virus p1 fusion proteins were found to be cleaved in planta and in escherichia coli. cleavage takes place in the absence of bacterial protease activity, indicating that the p1 fusion is autocatalytically processed independently of host factors. n-terminal sequencing of the c-terminal cleavage product of transiently expressed p1/gfp (green fluorescence protein) in nicotiana benthamiana showed that the cleavage site is located between the first two amino ac ... | 2010 | 20030588 |
| high-level rapid production of full-size monoclonal antibodies in plants by a single-vector dna replicon system. | plant viral vectors have great potential in rapid production of important pharmaceutical proteins. however, high-yield production of hetero-oligomeric proteins that require the expression and assembly of two or more protein subunits often suffers problems due to the "competing" nature of viral vectors derived from the same virus. previously we reported that a bean yellow dwarf virus (beydv)-derived, three-component dna replicon system allows rapid production of single recombinant proteins in pla ... | 2010 | 20047189 |
| covalent dimerization of camelidae anti-human tnf-alpha single domain antibodies by the constant kappa light chain domain improves neutralizing activity. | the tumor necrosis factor-alpha (tnfalpha) plays an important role in a number of chronic inflammatory disorders. monoclonal camelidae variable heavy chain domain-only antibodies (v(h)h) have been developed to antagonize the action of human tnfalpha (anti-tnf-v(h)h). here we describe a strategy to obtain functional dimeric anti-tnf-v(h)h molecules, based on the c-terminal fusion of a kappa light chain domain to the anti-tnf-v(h)h. the resulting fusion protein was transiently expressed by use of ... | 2010 | 20047190 |
| tomato 14-3-3 protein 7 positively regulates immunity-associated programmed cell death by enhancing protein abundance and signaling ability of mapkkk {alpha}. | programmed cell death (pcd) is triggered when pto, a ser-thr protein kinase, recognizes either the avrpto or avrptob effector from pseudomonas syringae pv tomato. this pcd requires mitogen-activated protein kinase kinase kinase (mapkkk alpha ) as a positive regulator in tomato (solanum lycopersicum) and nicotiana benthamiana. to examine how pcd-eliciting activity of the tomato mapkkk alpha protein is regulated, we screened for mapkkk alpha -interacting proteins in tomato and identified a 14-3-3 ... | 2010 | 20061552 |
| immunomodulatory activity of a plant extract containing human papillomavirus 16-e7 protein in human monocyte-derived dendritic cells. | this study reports the immunomodulatory activity on human monocyte derived dendritic cells (mddcs) of a vaccine preparation shown to be effective against an hpv16-related tumour in an animal model. the vaccine is composed of extract from nicotiana benthamiana leaves containing hpv16 e7 protein expressed by a potato virus x-derived vector (nbpvx-e7). the effect of the extract was evaluated on mddc differentiation and maturation by monitoring the phenotypic expression of specific markers. the resu ... | 2009 | 20074460 |
| the pathogenicity determinant of citrus tristeza virus causing the seedling yellows syndrome maps at the 3'-terminal region of the viral genome. | citrus tristeza virus (ctv) (genus closterovirus, family closteroviridae) causes some of the more important viral diseases of citrus worldwide. the ability to map disease-inducing determinants of ctv is needed to develop better diagnostic and disease control procedures. a distinctive phenotype of some isolates of ctv is the ability to induce seedling yellows (sy) in sour orange, lemon and grapefruit seedlings. in florida, the decline isolate of ctv, t36, induces sy, whereas a widely distributed ... | 2010 | 20078776 |
| identification of the virulence factors and suppressors of posttranscriptional gene silencing encoded by ageratum yellow vein virus, a monopartite begomovirus. | ageratum yellow vein disease (ayvd) is caused by the association of a tomato leaf curl java betasatellite [indonesia:indonesia 1:2003] (tolcjb-[id:id1:03]) with a begomovirus component. our previous results demonstrated that tolcjb-[id:id:03] is essential for induction of leaf curl symptoms in plants and transgene expression of its betac1 gene in nicotiana benthamiana plants induces virus-like symptoms. here we show that ageratum yellow vein virus-indonesia [indonesia: tomato] (ayvv-id[id:tom]) ... | 2010 | 20079777 |
| identification of tomato phosphatidylinositol-specific phospholipase-c (pi-plc) family members and the role of plc4 and plc6 in hr and disease resistance. | the perception of pathogen-derived elicitors by plants has been suggested to involve phosphatidylinositol-specific phospholipase-c (pi-plc) signalling. here we show that plc isoforms are required for the hypersensitive response (hr) and disease resistance. we characterised the tomato [solanum lycopersicum (sl)] plc gene family. six sl plc-encoding cdnas were isolated and their expression in response to infection with the pathogenic fungus cladosporium fulvum was studied. we found significant reg ... | 2010 | 20088897 |
| molecular characterization and infectivity of papaya leaf curl china virus infecting tomato in china. | papaya leaf curl china virus (palcucnv) was previously reported as a distinct begomovirus infecting papaya in southern china. based on molecular diagnostic survey, 13 palcucnv isolates were obtained from tomato plants showing leaf curl symptoms in henan and guangxi provinces of china. complete nucleotide sequences of 5 representative isolates (aj558116, aj558117, aj704604, fn256260, and fn297834) were determined to be 2738-2751 nucleotides, which share 91.7%-97.9% sequence identities with palcuc ... | 2010 | 20104645 |
| s-glycoprotein-like protein regulates defense responses in nicotiana plants against ralstonia solanacearum. | rsrga4 (for ralstonia solanacearum-responsive gene a4) encodes a polypeptide similar to s-locus glycoprotein (sgp) from brassica rapa and sgp-like proteins from ipomoea trifida and medicago truncatula. therefore, we designated rsrga4 as ntsglp (for nicotiana tabacum sgp-like protein) and nbsglp (its nicotiana benthamiana ortholog). nbsglp is expressed in root, leaf, petal, gynoecium, and stamen. expression of nbsglp was strongly induced by inoculation with an avirulent strain of r. solanacearum ... | 2010 | 20118275 |
| viral-induced systemic necrosis in plants involves both programmed cell death and the inhibition of viral multiplication, which are regulated by independent pathways. | resistant plants respond rapidly to invading avirulent plant viruses by triggering a hypersensitive response (hr). an hr is accompanied by a restraint of virus multiplication and programmed cell death (pcd), both of which have been observed in systemic necrosis triggered by a successful viral infection. here, we analyzed signaling pathways underlying the hr in resistance genotype plants and those leading to systemic necrosis. we show that systemic necrosis in nicotiana benthamiana, induced by pl ... | 2010 | 20121450 |
| a progeny virus from a cucumovirus pseudorecombinant evolved to gain the ability to accumulate its rna-silencing suppressor leading to systemic infection in tobacco. | two isolates of tomato aspermy virus (tav), v-tav and c-tav, can systemically infect nicotiana benthamiana but only c-tav can move systemically in n. tabacum. any pseudorecombinants between the two strains could not move systemically in tobacco as efficiently as c-tav. however, a pseudorecombinant consisting of rnas 1 and 3 of v-tav and rna 2 of c-tav (v1c2v3), which cannot infect tobacco systemically, generated progeny with a mutation in v1 and a recombination in c2 (v1(m)c2(r)v3), enabling the ... | 2010 | 20121454 |
| monoclonal antibody produced in plants efficiently treats west nile virus infection in mice. | over the past decade, west nile virus (wnv) has spread to all 48 of the lower united states as well as to parts of canada, mexico, the caribbean, and south america, with outbreaks of neuroinvasive disease occurring annually. at present, no therapeutic or vaccine is available for human use. epidemics of wnv and other emerging infectious disease threats demand cost-efficient and scalable production technologies that can rapidly transfer effective therapeutics into the clinical setting. we have pre ... | 2010 | 20133644 |
| immunogenic properties of chimeric potato virus x particles displaying the hepatitis c virus hypervariable region i peptide r9. | the immunogenic properties of chimeric potato virus x (pvx) particles engineered to display the synthetic r9 peptide have been evaluated. the r9 peptide is a consensus sequence derived from diverse variants of the hypervariable region 1 from the hepatitis c virus (hcv) envelope protein e2. two different constructs were designed, with the r9 peptide expressed either as an indirect fusion via the ribosomal skip 2a (pvx(r9-2a)cp) sequence or as a direct pvx coat protein fusion (pvx(r9)cp). systemic ... | 2010 | 20138085 |
| in vitro and in vivo interaction of atrma2 e3 ubiquitin ligase and auxin binding protein 1. | e3 ubiquitin (ub) ligases play diverse roles in cellular regulation in eukaryotes. three homologous atrmas (atrma1, atrma2, and atrma3) were recently identified as er-localized arabidopsis homologs of human ring membrane-anchor e3 ub ligase. here, auxin binding protein 1 (abp1), one of the auxin receptors in arabidopsis, was identified as a potential substrate of atrma2 through a yeast two-hybrid assay. an in vitro pull-down assay confirmed the interaction of full-length atrma2 with abp1. atrma2 ... | 2010 | 20152813 |
| hsp70 and its cochaperone cpip promote potyvirus infection in nicotiana benthamiana by regulating viral coat protein functions. | this study demonstrates that heat shock protein 70 (hsp70) together with its cochaperone cpip regulates the function of a potyviral coat protein (cp), which in turn can interfere with viral gene expression. hsp70 was copurified as a component of a membrane-associated viral ribonucleoprotein complex from potato virus a-infected plants. downregulation of hsp70 caused a cp-mediated defect associated with replication. when pva cp was expressed in trans, it interfered with viral gene expression and r ... | 2010 | 20154150 |
| hc-pro, a potyvirus rna silencing suppressor, cancels cycling of cucumber mosaic virus in nicotiana benthamiana plants. | the mixed infection of cucumber mosaic virus (cmv) and a potyvirus has been known to increase cmv titer in nicotiana benthamiana plants, resulting in synergistic viral symptoms. we found that among three potyviruses--potato virus y (pvy), turnip mosaic virus (tumv), and clover yellow vein virus (c1yvv)--synergistic effects on cmv (or a recombinant cmv vector) titers were most efficiently induced by a co-infection with pvy in n. benthamiana plants. in addition, the helper component-proteinase (hc ... | 2010 | 20162445 |
| binding of monoclonal antibodies to the movement protein (mp) of tobacco mosaic virus: influence of subcellular mp localization and phosphorylation. | monoclonal antibodies (mabs) to recombinant movement protein (mp(rec)) of tobacco mosaic virus (tmv) were used to reveal the dependence of mp epitope accessibility to mabs on subcellular mp localization and post-translational mp phosphorylation. leaves of nicotiana benthamiana or n. tabacum were inoculated mechanically with tmv or agroinjected with an mp expression vector. at different time post-inoculation, er membrane- and cell wall-enriched fractions (er-mp and cw-mp, respectively) were isola ... | 2010 | 20164264 |
| handygun: an improved custom-designed, non-vacuum gene gun suitable for virus inoculation. | particle bombardment with a non-vacuum gene gun is an efficient method for transfection of plant cells with cloned viruses and initiation of virus infection. the handygun developed in this study is an improved version of a non-vacuum gene gun. bombardment parameters were studied by inoculating an infectious, 35s promoter-driven cdna of potato virus a (pva; potyvirus) to the potato clone 'a6', nicotiana benthamiana and n. tabacum as plasmid dna coated on microprojectiles (gold particles). the lar ... | 2010 | 20188761 |
| multiple coat protein mutations abolish recognition of pepino mosaic potexvirus (pepmv) by the potato rx resistance gene in transgenic tomatoes. | despite the fact that pepino mosaic virus (pepmv) and potato virus x (pvx) share less than 40% identity in their coat proteins (cp), the known pvx elicitor of rx, transgenic tomato (cv. microtom) plants expressing a functional potato rx resistance gene showed resistance toward pepmv. however, in a low percentage of plants, pepmv accumulation was observed and back inoculation experiments demonstrated that these plants contained resistance-breaking pepmv variants. sequencing of the cp gene of thes ... | 2010 | 20192825 |
| a plastid-targeted heat shock cognate 70kda protein interacts with the abutilon mosaic virus movement protein. | the movement protein (mp) of bipartite geminiviruses facilitates cell-to-cell as well as long-distance transport within plants and influences viral pathogenicity. yeast two-hybrid assays identified a chaperone, the nuclear-encoded and plastid-targeted heat shock cognate 70kda protein (cphsc70-1) of arabidopsis thaliana, as a potential binding partner for the abutilon mosaic virus (abmv) mp. in planta, bimolecular fluorescence complementation (bifc) analysis showed cphsc70-1/mp complexes and mp h ... | 2010 | 20193958 |
| semicontinuous bioreactor production of a recombinant human therapeutic protein using a chemically inducible viral amplicon expression system in transgenic plant cell suspension cultures. | plant cell culture is an alternative for the production of recombinant human therapeutic proteins because of improved product safety, lower production cost, and capability for eukaryotic post-translational modification. in this study, bioreactor production of recombinant human alpha-1-antitrypsin (raat) glycoprotein using a chemically inducible cucumber mosaic virus (cmv) viral amplicon expression system in transgenic nicotiana benthamiana cell culture is presented. optimization of a chemically ... | 2010 | 20198659 |
| malvastrum yellow vein yunnan virus is amonopartite begomovirus. | nine samples of diseased malvastrum coromandelianum plants collected from the fields in yunnan province of china were found to be infected with malvastrum yellow vein yunnan virus (myvynv), when tested by pcr using specific primers. the results of pcr and southern blot analysis showed that only 4 samples out of 9 were associated with the satellite dnab molecules. their sequence analysis indicated that dnab molecules share 97.8%-99.4% of nucleotide sequence identities with dnab associated with my ... | 2010 | 20201610 |
| northern blotting techniques for small rnas. | in eukaryotes, rna silencing encompasses a range of biochemical processes mediated by approximately 20-25 nt small rnas (smrnas). this chapter describes northern blot hybridization techniques optimized for detection of such smrnas, whether extracted from plant or animal tissues. the basic protocol is described, and control blots illustrate the detection specificity and sensitivity of this method using dna oligonucleotide probes. known endogenous smrnas are analyzed in samples prepared from sever ... | 2010 | 20204871 |
| exploring different strategies to express dengue virus envelope protein in a plant system. | dengue virus envelope glycoprotein (e-protein) is the main protein associated with immunity induction. to produce a candidate for subunit vaccines and to provide an antigen for diagnostic kits, it was expressed in a novel plant system using deconstructed viral modules. a truncated version of the e-protein was designed to be expressed alone and co-expressed with dengue virus structural proteins. as well, the critical domain iii of e-protein was fused to hepatitis b core antigen (hbcore). the reco ... | 2010 | 20213522 |
| optimisation of the purification process of a tumour-targeting antibody produced in n. benthamiana using vacuum-agroinfiltration. | it was previously demonstrated that the tumour-targeting antibody mab h10 can be transiently expressed and purified at high levels in nicotiana benthamiana by using a vacuum-agroinfiltration system boosted by the use of a virus silencing suppressor protein. scope of this work was to analyse different steps of protein extraction from agroinfiltrated leaves to optimise the purification process of the secretory mab h10 providing new insights in the field of large-scale plant production. two differe ... | 2010 | 20229286 |
| a key enzyme for flavin synthesis is required for nitric oxide and reactive oxygen species production in disease resistance. | nitric oxide (no) and reactive oxygen species (ros) play key roles in plant immunity. however, the regulatory mechanisms of the production of these radicals are not fully understood. hypersensitive response (hr) cell death requires the simultaneous and balanced production of no and ros. in this study we indentified nbribaencoding a bifunctional enzyme, guanosine triphosphate cyclohydrolase ii/3,4-dihydroxy-2-butanone-4-phosphate synthase, which participates in the biosynthesis of flavin, by scre ... | 2010 | 20230506 |
| disruption of plant carotenoid biosynthesis through virus-induced gene silencing affects oviposition behaviour of the butterfly pieris rapae. | optical plant characteristics are important cues to plant-feeding insects. in this article, we demonstrate for the first time that silencing the phytoene desaturase (pds) gene, encoding a key enzyme in plant carotenoid biosynthesis, affects insect oviposition site selection behaviour. virus-induced gene silencing employing tobacco rattle virus was used to knock down endogenous pds expression in three plant species (arabidopsis thaliana, brassica nigra and nicotiana benthamiana) by its heterologo ... | 2010 | 20298487 |
| [new role for tobacco--production of biopharmaceuticals]. | over one hundred of biopharmaceuticals present on the pharmaceutical market--among them mainly therapeutically peptides and proteins such as hormones, enzymes, interferons, blood factors, monoclonal antibodies, vaccines, growth factors and fusion proteins, are manufactured through recombinant dna technology in cultures of transgenic bacterial (escherichia coli), yeast (saccharomyces cerevisiae) and mammalian cells (cho, bhk) and rarely insect cells or insects (bombyx mori). high costs of their m ... | 2009 | 20301965 |
| biosynthesis of glucosinolates--gene discovery and beyond. | glucosinolates are sulfur-rich secondary metabolites characteristic of the brassicales order with important biological and economic roles in plant defense and human nutrition. application of systems biology tools continues to identify genes involved in the biosynthesis of glucosinolates. recent progress includes genes in all three phases of the pathway, i.e. side-chain elongation of precursor amino acids, formation of the core glucosinolate structure and side-chain decoration. major breakthrough ... | 2010 | 20303821 |
| in planta protein sialylation through overexpression of the respective mammalian pathway. | many therapeutic proteins are glycosylated and require terminal sialylation to attain full biological activity. current manufacturing methods based on mammalian cell culture allow only limited control of this important posttranslational modification, which may lead to the generation of products with low efficacy. here we report in vivo protein sialylation in plants, which have been shown to be well suited for the efficient generation of complex mammalian glycoproteins. this was achieved by the i ... | 2010 | 20305285 |
| development of tobacco necrosis virus a as a vector for efficient and stable expression of fmdv vp1 peptides. | plant virus-based expression systems provide attractive alternatives for production of animal virus-originated antigenic peptides. in the present study, an infectious cdna clone of tobacco necrosis virus a chinese isolate (tnv-a(c)) was used for expression of different peptides derived from foot and mouth disease virus (fmdv) serotype o vp1 fused downstream of the coat protein (cp) open reading frame (orf). chenopodium amaranticolor inoculated with in vitro transcripts of the chimaeras developed ... | 2010 | 20331532 |
| endosome-associated crt1 functions early in resistance gene-mediated defense signaling in arabidopsis and tobacco. | resistance gene-mediated immunity confers protection against pathogen infection in a wide range of plants. a genetic screen for arabidopsis thaliana mutants compromised for recognition of turnip crinkle virus previously identified crt1, a member of the ghkl atpase/kinase superfamily. here, we demonstrate that crt1 interacts with various resistance proteins from different structural classes, and this interaction is disrupted when these resistance proteins are activated. the arabidopsis mutant crt ... | 2010 | 20332379 |
| a role for chloroplast-localized thylakoid formation 1 (thf1) in bacterial speck disease development. | coronatine (cor), a jasmonate mimic produced by pseudomonas syringae pv. tomato dc3000 (pst dc3000) is required for full virulence of pst dc3000 in tomato and arabidopsis. cor is shown to induce a range of physiological processes including chlorosis, root growth inhibition and anthocyanin accumulation in tomato. to elucidate the host/signaling genes involved in cor-responses, we utilized a forward genetics approach using nicotiana benthamiana and virus-induced gene silencing (vigs) and identifie ... | 2010 | 20339321 |
| the complete nucleotide sequence of a new bipartite begomovirus from brazil infecting abutilon. | the complete nucleotide sequence of abutilon mosaic brazil virus (abmbv), a new bipartite begomovirus from bahia, brazil, is described and analyzed phylogenetically. its dna a is most closely related to those of sida-infecting begomoviruses from brazil and forms a phylogenetic cluster with pepper- and euphorbia-infecting begomoviruses from central america. the dna b component forms a cluster with different sida- and okra-infecting begomoviruses from brazil. both components are distinct from thos ... | 2010 | 20349251 |
| tomato chocolate spot virus, a member of a new torradovirus species that causes a necrosis-associated disease of tomato in guatemala. | tomatoes in guatemala have been affected by a new disease, locally known as "mancha de chocolate" (chocolate spot). the disease is characterized by distinct necrotic spots on leaves, stems and petioles that eventually expand and cause a dieback of apical tissues. samples from symptomatic plants tested negative for infection by tomato spotted wilt virus, tobacco streak virus, tobacco etch virus and other known tomato-infecting viruses. a virus-like agent was sap-transmitted from diseased tissue t ... | 2010 | 20376682 |
| a tobamovirus expression vector for agroinfection of legumes and nicotiana. | the highest recombinant protein expression levels in plants have been achieved using tobacco mosaic virus (tmv) vectors via agroinoculation of the tobacco, nicotiana benthamiana. these vectors have been utilized for pharmaceutical protein production and also can serve as rapid gene expression screens for proteonomics. we have constructed a similar vector based on the legume-infecting tobamovirus, sunn hemp mosaic virus (shmv), by deleting the coat protein gene (shmv eliminate coat protein gene o ... | 2010 | 20380855 |
| efficiency of vigs and gene expression in a novel bipartite potexvirus vector delivery system as a function of strength of tgb1 silencing suppression. | we have developed plant virus-based vectors for virus-induced gene silencing (vigs) and protein expression, based on alternanthera mosaic virus (altmv), for infection of a wide range of host plants including nicotiana benthamiana and arabidopsis thaliana by either mechanical inoculation of in vitro transcripts or via agroinfiltration. in vivo transcripts produced by co-agroinfiltration of bacteriophage t7 rna polymerase resulted in t7-driven altmv infection from a binary vector in the absence of ... | 2010 | 20381827 |
| virus-based microrna expression for gene functional analysis in plants. | traditional virus-induced gene silencing (vigs) is a powerful virus-based short interfering rna-mediated rna silencing technique for plant functional genomics. besides short interfering rnas, micrornas (mirnas) have also been shown to regulate gene expression by rna silencing in various organisms. however, plant virus-based mirna silencing has not been reported. in addition, a number of plant mirnas have been identified or predicted, while their functions are largely unknown. thus, there is an u ... | 2010 | 20388670 |
| the pro/hel region is indispensable for packaging non-replicating turnip yellow mosaic virus rna, but not replicating viral rna. | turnip yellow mosaic virus (tymv) is a spherical plant virus that has a single 6.3 kb positive strand rna. the genomic rna has a trna-like structure (tls) at the 3'-end. the 3'-tls and hairpins in the 5'-untranslated region supposedly serve as packaging signals; however, recent studies have shown that they do not play a role in tymv rna packaging. in this study, we focused on packaging signals by examining a series of deletion mutants of tymv. analysis of encapsidated viral rna after agroinfiltr ... | 2010 | 20396967 |
| rna-dependent rna polymerase 1 from nicotiana tabacum suppresses rna silencing and enhances viral infection in nicotiana benthamiana. | endogenous eukaryotic rna-dependent rna polymerases (rdrs) produce double-stranded rna intermediates in diverse processes of small rna synthesis in rna silencing pathways. rdr6 is required in plants for posttranscriptional gene silencing induced by sense transgenes (s-ptgs) and has an important role in amplification of antiviral silencing. whereas rdr1 is also involved in antiviral defense in plants, this does not necessarily proceed through triggering silencing. in this study, we show that nico ... | 2010 | 20400679 |
| one-step, zero-background ligation-independent cloning intron-containing hairpin rna constructs for rnai in plants. | *the hairpin-based rna interference (rnai) technique plays an important role in exploring gene function in plants. although there are several methods for making hairpin rna (hprna) constructs, these methods usually need multiple relatively laborious, time-consuming or high-cost cloning steps. here we describe a one-step, zero-background ligation-independent cloning (oz-lic) method for making intron-containing hprna (ihprna) constructs by our vector prnai-lic. *to generate the ihprna constructs w ... | 2010 | 20406406 |
| 2-de proteome maps for the leaf apoplast of nicotiana benthamiana. | we provide 2-d gel reference maps for the apoplastic proteome of nicotiana benthamiana leaves infiltrated or not with the bacterial gene vector agrobacterium tumefaciens. about 90 proteins were analyzed by lc-ms/ms for identification and function assignment. we show, overall, an effective response of the plant to agroinfiltration involving a specific, cell wall maintenance-independent up-regulation of defense protein secretion. the proteome maps described should be a useful tool for systemic stu ... | 2010 | 20422621 |
| green biofactories: recombinant protein production in plants. | until recently, low accumulation levels have been the major bottleneck for plant-made recombinant protein production. however, several breakthroughs have been described in the past few years allowing for very high maccumulation levels, mainly through chloroplast transformation and transient expression, coupled with subcellular targeting and protein fusions. another important factor influencing our ability to use plants for the production of recombinant proteins is the availability of quick and s ... | 2010 | 20423324 |
| the 3'-terminal sequence of bamboo mosaic virus minus-strand rna interacts with rna-dependent rna polymerase and initiates plus-strand rna synthesis. | a 3'-terminal, 77-nucleotide sequence of bamboo mosaic virus (bamv) minus-strand rna (ba-77), comprising a 5' stem-loop, a spacer and a 3'-cuuuu sequence, can be used to initiate plus-strand rna synthesis in vitro. to understand the mechanism of plus-strand rna synthesis, mutations were introduced in the 5' untranslated region of bamv rna, resulting in changes at the 3' end of minus-strand rna. the results showed that at least three uridylate residues in 3'-cuuuu are required and the changes at ... | 2010 | 20447270 |
| the hypersensitive response induced by the v2 protein of a monopartite begomovirus is countered by the c2 protein. | a functional analysis of the v2 protein of two monopartite begomoviruses, papaya leaf curl virus (palcuv) and cotton leaf curl kokhran virus (clcukov), has been performed. expression of the v2 gene from a potato virus x (pvx) vector resulted in severe leaf curling followed by a hypersensitive response (hr) in nicotiana benthamiana and n. tabacum, demonstrating that the v2 protein is a pathogenicity determinant and a target of host defence responses. agroinfiltration of a pvx vector expressing th ... | 2010 | 20447273 |
| role of nitric oxide and reactive oxide species in disease resistance to necrotrophic pathogens. | nitric oxide (no) and reactive oxygen species (ros) are important signaling molecules in plant immunity. however, roles of no and ros in disease resistance to necrotrophic pathogens are not fully understood. we have recently demonstrated that no plays a pivotal role in basal defense against botrytis cinerea and the expression of the salicylic acid (sa) -responsive gene pr-1in nicotiana benthamiana. by contrast, ros function negatively in resistance or positively in expansion of disease lesions d ... | 2010 | 20448455 |
| identification of nicotiana benthamiana genes involved in pathogen-associated molecular pattern-triggered immunity. | in order to identify components of pathogen-associated molecular pattern-triggered immunity (pti) pathways in nicotiana benthamiana, we conducted a large-scale forward-genetics screen using virus-induced gene silencing and a cell-death-based assay for assessing pti. the assay relied on four combinations of pti-inducing nonpathogens and cell-death-causing challenger pathogens and was first validated in plants silenced for fls2 or bak1. over 3,200 genes were screened and 14 genes were identified t ... | 2010 | 20459311 |
| components of the pseudomonas syringae type iii secretion system can suppress and may elicit plant innate immunity. | the type iii secretion system (t3ss) of pseudomonas syringae translocates into plant cells multiple effectors that suppress pathogen-associated molecular pattern (pamp)-triggered immunity (pti). p. syringae pv. tomato dc3000 no longer delivers the t3ss translocation reporter avrpto-cya in nicotiana benthamiana leaf tissue in which pti was induced by prior inoculation with p. fluorescens(pln18). cosmid pln18 expresses the t3ss system of p. syringae pv. syringae 61 but lacks the hopa1(psy61) effec ... | 2010 | 20459312 |