Publications

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transient production of artemisinin in nicotiana benthamiana is boosted by a specific lipid transfer protein from a. annua.our lack of full understanding of transport and sequestration of the heterologous products currently limit metabolic engineering in plants for the production of high value terpenes. for instance, although all genes of the artemisinin/arteannuin b (an/ab) biosynthesis pathway (an-pw) from artemisia annua have been identified, ectopic expression of these genes in nicotiana benthamiana yielded mostly glycosylated pathway intermediates and only very little free (dihydro)artemisinic acid [(dh)aa]. he ...201627421621
the conformation of a plasma membrane-localized somatic embryogenesis receptor kinase complex is altered by a potato aphid-derived effector.somatic embryogenesis receptor kinases (serks) are transmembrane receptors involved in plant immunity. tomato (solanum lycopersicum) carries three serk members. one of these, slserk1, is required for mi-1.2-mediated resistance to potato aphids (macrosiphum euphorbiae). mi-1.2 encodes a coiled-coil nucleotide-binding leucine-rich repeat protein that in addition to potato aphids confers resistance to two additional phloem-feeding insects and to root-knot nematodes (meloidogyne spp.). how slserk1 p ...201627208261
rna1-independent replication and gfp expression from tomato marchitez virus isolate m cloned cdna.tomato marchitez virus (tomarv; synonymous with tomato apex necrosis virus) is a positive-strand rna virus in the genus torradovirus within the family secoviridae. tomarv is an emergent whitefly-transmitted virus that causes important diseases in tomato (solanum lycopersicum) in mexico. here, the genome sequence of the tomarv isolate m (tomarv-m) was determined. we engineered full-length cdna clones of the tomarv-m genomic rna (rna1 and rna2), separately, into a binary vector. coinfiltration of ...201626756828
the exs domain of pho1 participates in the response of shoots to phosphate deficiency via a root-to-shoot signal.the response of shoots to phosphate (pi) deficiency implicates long-distance communication between roots and shoots, but the participating components are poorly understood. we have studied the topology of the arabidopsis (arabidopsis thaliana) phosphate1 (pho1) pi exporter and defined the functions of its different domains in pi homeostasis and signaling. the results indicate that the amino and carboxyl termini of pho1 are both oriented toward the cytosol and that the protein spans the membrane ...201626546667
a duf-246 family glycosyltransferase-like gene affects male fertility and the biosynthesis of pectic arabinogalactans.pectins are a group of structurally complex plant cell wall polysaccharides whose biosynthesis and function remain poorly understood. the pectic polysaccharide rhamnogalacturonan-i (rg-i) has two types of arabinogalactan side chains, type-i and type-ii arabinogalactans. to date few enzymes involved in the biosynthesis of pectin have been described. here we report the identification of a highly conserved putative glycosyltransferase encoding gene, pectic arabinogalactan synthesis-related (pagr), ...201627091363
athd2d gene plays a role in plant growth, development, and response to abiotic stresses in arabidopsis thaliana.the histone deacetylases play important roles in the regulation of gene expression and the subsequent control of a number of important biological processes, including those involved in the response to environmental stress. a specific group of histone deacetylase genes, hd2, is present in plants. in arabidopsis, hd2s include hd2a, hd2b, hd2c, and hd2d. previous research showed that hd2a, hd2b, and hd2c are more related in terms of expression and function, but not hd2d. in this report, we studied ...201627066015
developments in the production of mucosal antibodies in plants.recombinant mucosal antibodies represent attractive target molecules for the development of next generation biopharmaceuticals for passive immunization against various infectious diseases and treatment of patients suffering from mucosal antibody deficiencies. as these polymeric antibodies require complex post-translational modifications and correct subunit assembly, they are considered as difficult-to-produce recombinant proteins. beside the traditional, mammalian-based production platforms, pla ...201626626615
specific control of arabidopsis bak1/serk4-regulated cell death by protein glycosylation.precise control of cell death is essential for the survival of all organisms. arabidopsis thaliana brassinosteroid insensitive 1-associated receptor kinase 1 (bak1) and somatic embryogenesis receptor kinase 4 (serk4) redundantly and negatively regulate cell death through elusive mechanisms. by deploying a genetic screen for suppressors of cell death triggered by virus-induced gene silencing of bak1/serk4 on arabidopsis knockout collections, we identified stt3a, a protein involved in n-glycosylat ...201627250875
a gene expression microarray for nicotiana benthamiana based on de novo transcriptome sequence assembly.nicotiana benthamiana has been widely used in laboratories around the world for studying plant-pathogen interactions and posttranscriptional gene expression silencing. yet the exploration of its transcriptome has lagged behind due to the lack of both adequate sequence information and genome-wide analysis tools, such as dna microarrays. despite the increasing use of high-throughput sequencing technologies, the dna microarrays still remain a popular gene expression tool, because they are cheaper a ...201627213006
targeted mutagenesis in plant cells through transformation of sequence-specific nuclease mrna.plant genome engineering using sequence-specific nucleases (ssns) promises to advance basic and applied plant research by enabling precise modification of endogenous genes. whereas dna is an effective means for delivering ssns, dna can integrate randomly into the plant genome, leading to unintentional gene inactivation. further, prolonged expression of ssns from dna constructs can lead to the accumulation of off-target mutations. here, we tested a new approach for ssn delivery to plant cells, na ...201627176769
construction and validation of a dual-transgene vector system for stable transformation in plants.in this study, we constructed dual-transgene vectors (pdt1, pdt7, and pdt7g) that simultaneously co-expressed two genes in plants. actin2 and ubq10 promoters were used to control the expression of these two genes. the 4×myc, 3×ha, and 3×flag reporter genes allowed for the convenient identification of a tunable co-expression system in plants, whereas the dexamethasone (dex) inducible reporter gene c-terminus of the glucocorticoid receptor (cgr) provided dex-dependent translocation of the fusion g ...201627157807
the er-membrane transport system is critical for intercellular trafficking of the nsm movement protein and tomato spotted wilt tospovirus.plant viruses move through plasmodesmata to infect new cells. the plant endoplasmic reticulum (er) is interconnected among cells via the er desmotubule in the plasmodesma across the cell wall, forming a continuous er network throughout the entire plant. this er continuity is unique to plants and has been postulated to serve as a platform for the intercellular trafficking of macromolecules. in the present study, the contribution of the plant er membrane transport system to the intercellular traff ...201626863622
identification of ourmiavirus 30k movement protein amino acid residues involved in symptomatology, viral movement, subcellular localization and tubule formation.several plant viruses encode movement proteins (mps) classified in the 30k superfamily. despite a great functional diversity, alignment analysis of mp sequences belonging to the 30k superfamily revealed the presence of a central core region, including amino acids potentially critical for mp structure and functionality. we performed alanine-scanning mutagenesis of the ourmia melon virus (oumv) mp, and studied the effects of amino acid substitutions on mp properties and virus infection. we identif ...201626637973
arabidopsis cml38, a calcium sensor that localizes to ribonucleoprotein complexes under hypoxia stress.during waterlogging and the associated oxygen deprivation stress, plants respond by the induction of adaptive programs, including the redirected expression of gene networks toward the synthesis of core hypoxia-response proteins. among these core response proteins in arabidopsis (arabidopsis thaliana) is the calcium sensor cml38, a protein related to regulator of gene silencing calmodulin-like proteins (rgscams). cml38 transcripts are up-regulated more than 300-fold in roots within 6 h of hypoxia ...201626634999
fertilization independent endosperm genes repress nbgh3.6 and regulate the auxin level during shoot development in nicotiana benthamiana.the fertilization independent endosperm (fie) gene is required to restrict endosperm development without fertilization, and it represses flowering during embryo and seedling development in arabidopsis thaliana however, the regulatory mechanism of the fie gene in postembryonic shoot development is not well understood. silencing of nicotiana benthamiana homologues of the fie gene, nbfie1 and nbfie2, resulted in the enhanced outgrowth of axillary buds and the impairment of secondary xylem different ...201626873977
the calmodulin-like proteins atcml4 and atcml5 are single-pass membrane proteins targeted to the endomembrane system by an n-terminal signal anchor sequence.calmodulins (cams) are important mediators of ca(2+) signals that are found ubiquitously in all eukaryotic organisms. plants contain a unique family of calmodulin-like proteins (cmls) that exhibit greater sequence variance compared to canonical cams. the arabidopsis thaliana proteins atcml4 and atcml5 are members of cml subfamily vii and possess a cam domain comprising the characteristic double pair of ef-hands, but they are distinguished from other members of this subfamily and from canonical c ...201627029353
phosphate deficiency induces the jasmonate pathway and enhances resistance to insect herbivory.during their life cycle, plants are typically confronted by simultaneous biotic and abiotic stresses. low inorganic phosphate (pi) is one of the most common nutrient deficiencies limiting plant growth in natural and agricultural ecosystems, while insect herbivory accounts for major losses in plant productivity and impacts ecological and evolutionary changes in plant populations. here, we report that plants experiencing pi deficiency induce the jasmonic acid (ja) pathway and enhance their defense ...201627016448
the histone deacetylase complex 1 protein of arabidopsis has the capacity to interact with multiple proteins including histone 3-binding proteins and histone 1 variants.intrinsically disordered proteins can adopt multiple conformations, thereby enabling interaction with a wide variety of partners. they often serve as hubs in protein interaction networks. we have previously shown that the histone deacetylase complex 1 (hdc1) protein from arabidopsis (arabidopsis thaliana) interacts with histone deacetylases and quantitatively determines histone acetylation levels, transcriptional activity, and several phenotypes, including abscisic acid sensitivity during germin ...201626951436
a glycosyltransferase from nicotiana alata pollen mediates synthesis of a linear (1,5)-α-l-arabinan when expressed in arabidopsis.the walls of nicotiana alata pollen tubes contain a linear arabinan composed of (1,5)-α-linked arabinofuranose residues. although generally found as a side chain on the backbone of the pectic polysaccharide rhamnogalacturonan i, the arabinan in n. alata pollen tubes is considered free, as there is no detectable rhamnogalacturonan i in these walls. carbohydrate-specific antibodies detected arabinan epitopes at the tip and along the shank of n. alata pollen tubes that are predominantly part of the ...201626850276
asparagus irx9, irx10, and irx14a are components of an active xylan backbone synthase complex that forms in the golgi apparatus.heteroxylans are abundant components of plant cell walls and provide important raw materials for the food, pharmaceutical, and biofuel industries. a number of studies in arabidopsis (arabidopsis thaliana) have suggested that the irregular xylem9 (irx9), irx10, and irx14 proteins, as well as their homologs, are involved in xylan synthesis via a golgi-localized complex termed the xylan synthase complex (xsc). however, both the biochemical and cell biological research lags the genetic and molecular ...201626951434
optimization of engineered production of the glucoraphanin precursor dihomomethionine in nicotiana benthamiana.glucosinolates are natural products characteristic of the brassicales order, which include vegetables such as cabbages and the model plant arabidopsis thaliana. glucoraphanin is the major glucosinolate in broccoli and associated with the health-promoting effects of broccoli consumption. toward our goal of creating a rich source of glucoraphanin for dietary supplements, we have previously reported the feasibility of engineering glucoraphanin in nicotiana benthamiana through transient expression o ...201626909347
eril1, the plant homologue of eri-1, is involved in the processing of chloroplastic rrnas.proteins belonging to the enhancer of rna interference-1 subfamily of 3'-5' exoribonucleases participate in divergent rna pathways. they degrade small interfering rnas (sirnas), thus suppressing rna interference, and are involved in the maturation of ribosomal rnas and the degradation of histone messenger rnas (mrnas). here, we report evidence for the role of the plant homologue of these proteins, which we termed enhanced rna interference-1-like-1 (eril1), in chloroplast function. in vitro assay ...201627531275
mouse fat storage-inducing transmembrane protein 2 (fit2) promotes lipid droplet accumulation in plants.fat storage-inducing transmembrane protein 2 (fit2) is an endoplasmic reticulum (er)-localized protein that plays an important role in lipid droplet (ld) formation in animal cells. however, no obvious homologue of fit2 is found in plants. here, we tested the function of fit2 in plant cells by ectopically expressing mouse (mus musculus) fit2 in nicotiana tabacum suspension-cultured cells, nicotiana benthamiana leaves and arabidopsis thaliana plants. confocal microscopy indicated that the expressi ...201627987528
mdmyb93 is a regulator of suberin deposition in russeted apple fruit skins.a comparison of the transcriptomes of russeted vs nonrusseted apple skins previously highlighted a tight relationship between a gene encoding an myb-type transcription factor, mdmyb93, and some key suberin biosynthetic genes. the present work assesses the role of this transcription factor in the suberization process. a phylogenetic analysis of mdmyb93 and arabidopsis thaliana mybs was performed and the function of mdmyb93 was further investigated using agrobacterium-mediated transient overexpres ...201627716944
dimerization of the vacuolar receptors atrmr1 and -2 from arabidopsis thaliana contributes to their localization in the trans-golgi network.in arabidopsis thaliana, different types of vacuolar receptors were discovered. the atvsr (vacuolar sorting receptor) receptors are well known to be involved in the traffic to lytic vacuole (lv), while few evidences demonstrate the involvement of the receptors from atrmr family (receptor membrane ring-h2) in the traffic to the protein storage vacuole (psv). in this study we focused on the localization of two members of atrmr family, atrmr1 and -2, and on the possible interaction between these tw ...201627706038
an easy method for plant polysome profiling.translation of mrna to protein is a fundamental and highly regulated biological process. polysome profiling is considered as a gold standard for the analysis of translational regulation. the method described here is an easy and economical way for fractionating polysomes from various plant tissues. a sucrose gradient is made without the need for a gradient maker by sequentially freezing each layer. cytosolic extracts are then prepared in a buffer containing cycloheximide and chloramphenicol to im ...201627684295
molecular evolution and functional characterization of a bifunctional decarboxylase involved in lycopodium alkaloid biosynthesis.lycopodium alkaloids (las) are derived from lysine (lys) and are found mainly in huperziaceae and lycopodiaceae. las are potentially useful against alzheimer's disease, schizophrenia, and myasthenia gravis. here, we cloned the bifunctional lysine/ornithine decarboxylase (l/odc), the first gene involved in la biosynthesis, from the la-producing plants lycopodium clavatum and huperzia serrata we describe the in vitro and in vivo functional characterization of the l. clavatum l/odc (lcl/odc). the r ...201627303024
knocking down expression of the auxin-amidohydrolase iar3 alters defense responses in solanaceae family plants.in plants, indole-3-acetic acid (iaa) amido hydrolases (ahs) participate in auxin homeostasis by releasing free iaa from iaa-amino acid conjugates. we investigated the role of iar3, a member of the iaa amido hydrolase family, in the response of solanaceous plants challenged by biotrophic and hemi-biotrophic pathogens. by means of genome inspection and phylogenic analysis we firstly identified iaa-ah sequences and putative iar3 orthologs in nicotiana benthamiana, tomato and potato. we evaluated t ...201627968994
identification of phakopsora pachyrhizi candidate effectors with virulence activity in a distantly related pathosystem.phakopsora pachyrhizi is the causal agent of asian soybean rust, a disease that causes enormous economic losses, most markedly in south america. p. pachyrhizi is a biotrophic pathogen that utilizes specialized feeding structures called haustoria to colonize its hosts. in rusts and other filamentous plant pathogens, haustoria have been shown to secrete effector proteins into their hosts to permit successful completion of their life cycle. we have constructed a cdna library from p. pachyrhizi haus ...201627014295
cytosolic activation of cell death and stem rust resistance by cereal mla-family cc-nlr proteins.plants possess intracellular immune receptors designated "nucleotide-binding domain and leucine-rich repeat" (nlr) proteins that translate pathogen-specific recognition into disease-resistance signaling. the wheat immune receptors sr33 and sr50 belong to the class of coiled-coil (cc) nlrs. they confer resistance against a broad spectrum of field isolates of puccinia graminis f. sp. tritici, including the ug99 lineage, and are homologs of the barley powdery mildew-resistance protein mla10. here, ...201627555587
focal effector accumulation in a biotrophic interface at the primary invasion sites of colletotrichum orbiculare in multiple susceptible plants.we identified virulence-related effectors of a hemibiotrophic fungal pathogen colletotrichum orbiculare, and found that a novel interface was generated by a biotrophic interaction between c. orbiculare and the host cucumber, in which the effectors secreted from the pathogen accumulated preferentially. the interface was located around the biotrophic primary hyphal neck. here, we showed that c. orbiculare also developed this interface in a biotrophic interaction with melon, which belongs to cucurb ...201626829249
pstha5a23, a candidate effector from the obligate biotrophic pathogen puccinia striiformis f. sp. tritici, is involved in plant defense suppression and rust pathogenicity.during the infection of host plants, pathogens can deliver virulence-associated 'effector' proteins to promote plant susceptibility. however, little is known about effector function in the obligate biotrophic pathogen puccinia striiformis f. sp. tritici (pst) that is an important fungal pathogen in wheat production worldwide. here, they report their findings on an in planta highly induced candidate effector from pst, pstha5a23. the pstha5a23 gene is unique to pst and shows a low level of intra-s ...201627871149
tasyp71, a qc-snare, contributes to wheat resistance against puccinia striiformis f. sp. tritici.n-ethylmaleimide-sensitive factor attachment protein receptors (snares) are involved in plant resistance; however, the role of syp71 in the regulation of plant-pathogen interactions is not well known. in this study, we characterized a plant-specific snare in wheat, tasyp71, which contains a qc-snare domain. three homologs are localized on chromosome 1al, 1bl, and 1dl. using agrobacterium-mediated transient expression, tasyp71 was localized to the plasma membrane in nicotiana benthamiana. quantit ...201627148348
identification and characterization of suppressors of plant cell death (spd) effectors from magnaporthe oryzae.phytopathogenic microorganisms, including the fungal pathogen magnaporthe oryzae, secrete a myriad of effector proteins to facilitate infection. utilizing the transient expression of candidate effectors in the leaves of the model plant nicotiana benthamiana, we identified 11 suppressors of plant cell death (spd) effectors from m. oryzae that were able to block the host cell death reaction induced by nep1. ten of these 11 were also able to suppress bax-mediated plant cell death. five of the 11 sp ...201627301772
an e3 ligase affects the nlr receptor stability and immunity to powdery mildew.following the detection of pathogen cognate effectors, plant nod-like receptors (nlrs) trigger isolate-specific immunity that is generally associated with cell death. the regulation of nlr stability is important to ensure effective immunity. in barley (hordeum vulgare), the allelic mildew locus a (mla) receptors mediate isolate-specific disease resistance against powdery mildew fungus (blumeria graminis f. sp. hordei). currently, how mla stability is controlled remains unknown. here, we identifi ...201627780896
heterologous expression screens in nicotiana benthamiana identify a candidate effector of the wheat yellow rust pathogen that associates with processing bodies.rust fungal pathogens of wheat (triticum spp.) affect crop yields worldwide. the molecular mechanisms underlying the virulence of these pathogens remain elusive, due to the limited availability of suitable molecular genetic research tools. notably, the inability to perform high-throughput analyses of candidate virulence proteins (also known as effectors) impairs progress. we previously established a pipeline for the fast-forward screens of rust fungal candidate effectors in the model plant nicot ...201626863009
magnaporthe oryzae effectors moheg13 and moheg16 interfere with host infection and moheg13 counteracts cell death caused by magnaporthe-nlps in tobacco.adapted pathogens are able to modulate cell responses of their hosts most likely due to the activity of secreted effector molecules thereby enabling colonisation by ostensible nonhost pathogens. it is postulated that host and nonhost pathogens of a given plant species differ in their repertoire of secreted effector molecules that are able to suppress plant resistance. we pursued the strategy of identifying novel effectors of magnaporthe oryzae, the causal agent of blast disease, by comparing the ...201626883226
ectopic expression of arabidopsis l-type lectin receptor kinase genes lecrk-i.9 and lecrk-ix.1 in nicotiana benthamiana confers phytophthora resistance.transgenic nicotiana benthamiana lines with constitutive expression of an arabidopsis lectin receptor kinase gene (lecrk - i.9 or lecrk - ix.1) show enhanced resistance to phytophthora pathogens, demonstrating conserved gene functionality after interfamily transfer. in plants, cell surface receptors mediate the first layer of innate immunity against pathogenic microbes. in arabidopsis several l-type lectin receptor kinases (lecrks) were previously found to function as phytophthora resistance com ...201626795144
transcriptional dynamics of phytophthora infestans during sequential stages of hemibiotrophic infection of tomato.hemibiotrophic plant pathogens, such as the oomycete phytophthora infestans, employ a biphasic infection strategy, initially behaving as biotrophs, where minimal symptoms are exhibited by the plant, and subsequently as necrotrophs, feeding on dead plant tissue. the regulation of this transition and the breadth of molecular mechanisms that modulate plant defences are not well understood, although effector proteins secreted by the pathogen are thought to play a key role. we examined the transcript ...201625845484
studying the mechanism of plasmopara viticola rxlr effectors on suppressing plant immunity.the rxlr effector family, produced by oomycete pathogens, may manipulate host physiological and biochemical events inside host cells. a group of putative rxlr effectors from plasmopara viticola have been recently identified by rna-seq analysis in our lab. however, their roles in pathogenesis are poorly understood. in this study, we attempted to characterize 23 pvrxlr effector candidates identified from a p. viticola isolate "zj-1-1." during host infection stages, expression patterns of the effec ...201627242731
comprehensive transcriptional and functional analyses of melatonin synthesis genes in cassava reveal their novel role in hypersensitive-like cell death.melatonin is a widely known hormone in animals. since melatonin was discovered in plants, more and more studies highlight its involvement in a wide range of physiological processes including plant development and stress responses. many advances have been made in the terms of melatonin-mediated abiotic stress resistance and innate immunity in plants, focusing on model plants such as rice and arabidopsis. in this study, 7 melatonin synthesis genes were systematically analyzed in cassava. quantitat ...201627739451
reduced susceptibility to xanthomonas citri in transgenic citrus expressing the fls2 receptor from nicotiana benthamiana.overexpression of plant pattern-recognition receptors by genetic engineering provides a novel approach to enhance plant immunity and broad-spectrum disease resistance. citrus canker disease associated with xanthomonas citri is one of the most important diseases damaging citrus production worldwide. in this study, we cloned the fls2 gene from nicotiana benthamiana cdna and inserted it into the binary vector pbinplus/ars to transform hamlin sweet orange and carrizo citrange. transgene presence was ...201626554734
comparison of helper component-protease rna silencing suppression activity, subcellular localization, and aggregation of three korean isolates of turnip mosaic virus.in 2014, we performed a nationwide survey in korean radish fields to investigate the distribution and variability of turnip mosaic virus (tumv). brassica rapa ssp. pekinensis sap-inoculated with three isolates of tumv from infected radish tissue showed different symptom severities, whereas symptoms in raphanus sativus were similar for each isolate. the helper component-protease (hc-pro) genes of each isolate were sequenced, and phylogenetic analysis showed that the three korean isolates were clu ...201627059238
host-specific accumulation and temperature effects on the generation of dimeric viral rna species derived from the s-rna of members of the tospovirus genus.polygonum ringspot virus (polrsv) is a recently characterized tospovirus reported in italy. northern blot analyses of polrsv infections in nicotiana benthamiana and tomato plants showed that a viral rna species with nearly twice the length of the small genomic rna (s-rna) accumulated abundantly in the former host, but was not detected in the latter. additional assays confirmed that biogenesis of this novel rna species was common to all polrsv isolates tested and also to an isolate of tomato spot ...201627600541
the p22 rna silencing suppressor of the crinivirus tomato chlorosis virus is dispensable for local viral replication but important for counteracting an antiviral rdr6-mediated response during systemic infection.among the components of the rna silencing pathway in plants, rna-dependent rna polymerases (rdrs) play fundamental roles in antiviral defence. here, we demonstrate that the nicotiana benthamiana rdr6 is involved in defence against the bipartite crinivirus (genus crinivirus, family closteroviridae) tomato chlorosis virus (tocv). additionally, by producing a p22-deficient tocv infectious mutant clone (tocvδp22), we studied the role of this viral suppressor of rna silencing in viral infection in bo ...201627367718
an in vitro reprogrammable antiviral risc with size-preferential ribonuclease activity.infection of nicotiana benthamiana plants with tomato bushy stunt virus (tbsv) mutants compromised for silencing suppression induces formation of an antiviral risc (vrisc) that can be isolated using chromatography procedures. the isolated vrisc sequence-specifically degrades tbsv rna in vitro, its activity can be down-regulated by removing sirnas, and re-stimulated by exogenous supply of sirnas. vrisc is most effective at hydrolyzing the ~4.8kb genomic rna, but less so for a ~2.2kb tbsv subgenom ...201626812224
transcriptomic changes in nicotiana benthamiana plants inoculated with the wild-type or an attenuated mutant of tobacco vein banding mosaic virus.tobacco vein banding mosaic virus (tvbmv) is a potyvirus which mainly infects solanaceous crops. the helper component proteinase (hcpro) of a potyvirus is an rna silencing suppressor protein and determines the severity of disease symptoms caused by different potyviruses, including tvbmv. it has been shown that substitution mutations introduced into the hcpro open reading frame (orf) in a tvbmv infectious clone result in changes of asp189 to lys or ile250 -gln251 to asp-glu (asp, aspartic acid; g ...201627539720
evaluation and identification of candidate genes for artificial microrna-mediated resistance to tomato spotted wilt virus.tomato spotted wilt virus (tswv) is an economically important viral pathogen of a wide range of field and horticultural crops. we developed an artificial microrna (amirna) strategy against tswv, targeting the nucleoprotein (n) and silencing suppressor (nss) genes. the amirna constructs replaced the natural mirna in a shortened arabidopsis 173-nucleotide (nt) mir159a precursor backbone (athmir159a) without the stem base extending beyond the mir/mir* duplex. further, each amirna was modified to co ...201626454192
comparison of small rna profiles in nicotiana benthamiana and solanum lycopersicum infected by polygonum ringspot tospovirus reveals host-specific responses to viral infection.viral small rnas (vsrnas) are one of the key elements involved in rna silencing-based defense against viruses in plants. we analyzed the vsrna profiles in nicotiana benthamiana and solanum lycopersicum infected by polygonum ringspot virus (polrsv) (tospovirus, bunyaviridae). vsrnas were abundant in both hosts, but a different size profile was observed, with an abundance peak at 21 in n. benthamiana and at 22 nt in tomato. vsrnas mapping to the polrsv l genomic segment were under-represented in b ...201626432447
the c2 protein of tomato yellow leaf curl sardinia virus acts as a pathogenicity determinant and a 16-amino acid domain is responsible for inducing a hypersensitive response in plants.the role of the c2 protein in the pathogenicity of tomato yellow leaf curl sardinia virus (tylcsv) was investigated. here we report that agrobacterium-mediated transient expression of tylcsv c2 resulted in a strong hypersensitive response (hr) in nicotiana benthamiana, n. tabacum, and arabidopsis thaliana, with induction of plant cell death and production of h2o2. since hr is not evident in plants infected by tylcsv, it is expected that tylcsv encodes a gene (or genes) that counters this respons ...201626826600
a single amino acid substitution in movement protein of tomato torrado virus influences totv infectivity in solanum lycopersicum.tomato torrado virus (totv), which is a tomato-infecting member of the genus torradovirus, induces severe systemic necrosis in solanum lycopersicum cv. beta lux as well as leaf malformation and chlorosis in nicotiana benthamiana. to date, neither the tomato gene conferring resistance to the pathogen nor the totv-encoded necrosis determinant have been characterized. we herein revealed that the phenylalanine 210 residue in the movement protein domain encoded by totv rna2 is a necrosis-inducing pat ...201626562559
tobacco rattle virus 16k silencing suppressor binds argonaute 4 and inhibits formation of rna silencing complexes.the cysteine-rich 16k protein of tobacco rattle virus (trv), the type member of the genus tobravirus, is known to suppress rna silencing. however, the mechanism of action of the 16k suppressor is not well understood. in this study, we used a gfp-based sensor strategy and an agrobacterium-mediated transient assay in nicotiana benthamiana to show that 16k was unable to inhibit the activity of existing small interfering rna (sirna)- and microrna (mirna)-programmed rna-induced silencing effector com ...201626498945
endoplasmic reticulum stress responses function in the hrt-mediated hypersensitive response in nicotiana benthamiana.hrt is a plant coiled-coil, nucleotide-binding and leucine-rich repeat (cc-nb-lrr) disease resistance protein that triggers the hypersensitive response (hr) on recognition of turnip crinkle virus (tcv) coat protein (cp). the molecular mechanism and significance of hr-mediated cell death for tcv resistance have not been fully elucidated. to identify the genes involved in hrt/tcv cp-mediated hr in nicotiana benthamiana, we performed virus-induced gene silencing (vigs) of 459 expressed sequence tag ...201626780303
the involvement of ros producing aldehyde oxidase in plant response to tombusvirus infection.the influence of tomato bushy stunt virus (tbsv) infection on the activity and isoformic composition of aldehyde oxidase and catalase in nicotiana benthamiana plants was investigated. it was shown that the infection of plants with tbsv results in enhancement of leaf aldehyde oxidase (ao) isoforms ao2 and ao3. significantly enhanced levels of superoxide radical producing activity of ao isoforms were also detected. this is the first demonstration of involvement of plant ao in defense mechanisms ag ...201627632242
the p1n-pispo trans-frame gene of sweet potato feathery mottle potyvirus is produced during virus infection and functions as an rna silencing suppressor.the positive-sense rna genome of sweet potato feathery mottle virus (spfmv) (genus potyvirus, family potyviridae) contains a large open reading frame (orf) of 3,494 codons translatable as a polyprotein and two embedded shorter orfs in the -1 frame: pispo, of 230 codons, and pipo, of 66 codons, located in the p1 and p3 regions, respectively. pispo is specific to some sweet potato-infecting potyviruses, while pipo is present in all potyvirids. in spfmv these two extra orfs are preceded by conserve ...201626792740
identification of amino acid residues of the coat protein of sri lankan cassava mosaic virus affecting symptom production and viral titer in nicotiana benthamiana.sri lankan cassava mosaic virus (slcmv) is bipartite begomovirus infecting cassava in india and sri lanka. interestingly, the dna-a component of the slcmv alone is able to infect nicotiana benthamiana causing symptoms of upward leaf rolling and stunting. one of the differences between monopartite and bipartite begomoviruses is the requirement of coat protein (cp) for infectivity; cp being essential for the former, but dispensable in the latter. this investigation was aimed to determine the impor ...201626948262
transient expression of tetrameric recombinant human butyrylcholinesterase in nicotiana benthamiana.to optimize the expression, extraction and purification of plant-derived tetrameric recombinant human butyrylcholinesterase (prbche), we describe the development and use of plant viral amplicon-based gene expression system; tobacco mosaic virus (tmv) rna-based overexpression vector (trbo) to express enzymatically active flag-tagged plant made recombinant butyrylcholinesterase (rbche) in nicotiana benthamiana leaves using transient agroinfiltration. two gene expression cassettes were designed to ...201627379103
production of recombinant cholera toxin b subunit in nicotiana benthamiana using geneware® tobacco mosaic virus vector.here, we describe a method to produce a recombinant cholera toxin b subunit in nicotiana benthamiana plants (ctbp) using the geneware(®) tobacco mosaic virus vector system. infectious transcripts of the vector rna are generated in vitro and inoculated on n. benthamiana seedlings. after 11 days, ctbp is extracted in a simple tris buffer at room temperature. no protease inhibitor is required. the leaf homogenate is treated with mild heat and a ph shift to selectively precipitate host-derived prote ...201626614286
production of bioactive wheat puroindoline proteins in nicotiana benthamiana using a virus-based transient expression system.the emergence of antibiotic resistant pathogenic strains of bacteria has necessitated the development of novel antimicrobial agents. the puroindoline a and b (pina and pinb) proteins of wheat, well-known for their roles in determining the important phenotype of grain texture, are also antimicrobial, making them attractive as natural bio-control agents. however, the biochemical basis of pin functionality remains unclear due to limitations in expressing them at the required yield and purity and la ...201626363114
soil-borne wheat mosaic virus infectious clone and manipulation for gene-carrying capacity.a full-length infectious cdna clone of soil-borne wheat mosaic virus (sbwmv; genus furovirus; family virgaviridae) was developed for agrobacterium delivery. the cloned virus can be agroinfiltrated to nicotiana benthamiana for subsequent infection of wheat (triticum aestivum, l.). the utility of the virus as a vector for gene silencing and expression was assessed through sequence insertions in multiple sites of rna2. virus-induced photobleaching was observed in n. benthamiana but not in wheat, de ...201627236459
an extracellular zn-only superoxide dismutase from puccinia striiformis confers enhanced resistance to host-derived oxidative stress.accumulation of reactive oxygen species (ros) following plant-pathogen interactions can trigger plant defence responses and directly damage pathogens. thus, it is essential for pathogens to scavenge host-derived ros to establish a parasitic relationship. however, the mechanisms protecting pathogens from host-derived oxidative stress remain unclear. in this study, a superoxide dismutase (sod) gene, pssod1, was cloned from a wheat-puccinia striiformis f. sp. tritici (pst) interaction cdna library. ...201627399209
the aquaporin splice variant nbxip1;1α is permeable to boric acid and is phosphorylated in the n-terminal domain.aquaporins (aqps) are membrane channel proteins that transport water and uncharged solutes across different membranes in organisms in all kingdoms of life. in plants, the aqps can be divided into seven different subfamilies and five of these are present in higher plants. the most recently characterized of these subfamilies is the xip subfamily, which is found in most dicots but not in monocots. in this article, we present data on two different splice variants (α and β) of nbxip1;1 from nicotiana ...201627379142
14-3-3 protein mediates plant seed oil biosynthesis through interaction with atwri1.plant 14-3-3 proteins are phosphopeptide-binding proteins, belonging to a large family of proteins involved in numerous physiological processes including primary metabolism, although knowledge about the function of 14-3-3s in plant lipid metabolism is sparse. wrinkled1 (wri1) is a key transcription factor that governs plant oil biosynthesis. at present, atwri1-interacting partners remain largely unknown. here, we show that 14-3-3 proteins are able to interact with atwri1, both in yeast and plant ...201627322486
a novel rice protein family of oshigds may be involved in early signalling of hypoxia-promoted stem growth in deepwater rice.oshigds was identified as a novel hypoxia-responsive protein family. among them, oshigd2 is characterized as a mitochondrial protein and is related to hypoxia signalling through interacting with mitochondrial proteins of critical functions in reducing cell damages caused by hypoxia. recent evidence supports ethylene as a key factor in modulating plant responses to submergence stress. meanwhile, there has been general consent that ethylene is not the only signal for the submergence-induced stem g ...201627306456
overexpression of a stress-responsive nac transcription factor gene onac022 improves drought and salt tolerance in rice.the nac transcription factors play critical roles in regulating stress responses in plants. however, the functions for many of the nac family members in rice are yet to be identified. in the present study, a novel stress-responsive rice nac gene, onac022, was identified. expression of onac022 was induced by drought, high salinity, and abscisic acid (aba). the onac022 protein was found to bind specifically to a canonical nac recognition cis-element sequence and showed transactivation activity at ...201626834774
elucidation of the first committed step in betalain biosynthesis enables the heterologous engineering of betalain pigments in plants.betalains are tyrosine-derived red-violet and yellow pigments, found in plants only of the caryophyllales order. although much progress has been made in recent years in the understanding of the betalain biosynthetic process, many questions remain open with regards to several of the proposed steps in the pathway. most conspicuous by its absence is the characterization of the first committed step in the pathway, namely the 3-hydroxylation of tyrosine to form l-3,4-dihydroxyphenylalanine (l-dopa). ...201626683006
identification and characterization of plant cell death-inducing secreted proteins from ustilaginoidea virens.ustilaginoidea virens (cooke) takah (telemorph villosiclava virens) is an ascomycetous fungus that causes rice false smut, one of the most important rice diseases. fungal effectors often play essential roles in host-pathogen coevolutionary interactions. however, little is known about the functions of u. virens effectors. here, we performed functional studies on putative effectors in u. virens and demonstrated that 13 of 119 putative effectors caused necrosis or necrosis-like phenotypes in nicoti ...201626927000
a r2r3-myb transcription factor regulates the flavonol biosynthetic pathway in a traditional chinese medicinal plant, epimedium sagittatum.flavonols as plant secondary metabolites with vital roles in plant development and defense against uv light, have been demonstrated to be the main bioactive components (bcs) in the genus epimedium plants, several species of which are used as materials for herba epimedii, an important traditional chinese medicine. the flavonol biosynthetic pathway genes had been already isolated from epimedium sagittatum, but a r2r3-myb transcription factor regulating the flavonol synthesis has not been functiona ...201627493658
two subclasses of differentially expressed tps1 genes and biochemically active tps1 proteins may contribute to sugar signalling in kiwifruit actinidia chinensis.trehalose metabolism and its intermediate trehalose-6-phosphate (t6p) are implicated in sensing and signalling sucrose availability. four class i trehalose-6-phosphate synthase (tps1) genes were identified in kiwifruit, three of which have both the tps and trehalose-6-phosphate phosphatase (tpp) domain, while the fourth gene gives rise to a truncated transcript. the transcript with highest sequence homology to arabidopsis tps1, designated tps1.1a was the most highly abundant tps1 transcript in a ...201627992562
a member of the phosphate transporter 1 (pht1) family from the arsenic-hyperaccumulating fern pteris vittata is a high-affinity arsenate transporter.pteris vittata exhibits enhanced arsenic uptake, but the corresponding mechanisms are not well known. the prevalent form of arsenic in most soils is arsenate, which is a phosphate analog and a substrate for phosphate transporter 1 (pht1) transporters. herein we identify and characterize three p. vittata pht1 transporters. pteris vittata pht1 cdnas were isolated and characterized via heterologous expression in saccharomyces cerevisiae (yeast) and nicotiana benthamiana leaves. expression of the pv ...201626010225
comparative analyses of ubiquitin-like atg8 and cysteine protease atg4 autophagy genes in the plant lineage and cross-kingdom processing of atg8 by atg4.autophagy is important for degradation and recycling of intracellular components. in a diversity of genera and species, orthologs and paralogs of the yeast atg4 and atg8 proteins are crucial in the biogenesis of double-membrane autophagosomes that carry the cellular cargoes to vacuoles and lysosomes. although many plant genome sequences are available, the atg4 and atg8 sequence analysis is limited to some model plants. we identified 28 atg4 and 116 atg8 genes from the available 18 different plan ...201627540766
aldo-keto reductase enzymes detoxify glyphosate and improve herbicide resistance in plants.in recent years, concerns about the use of glyphosate-resistant (gr) crops have increased because of glyphosate residual levels in plants and development of herbicide-resistant weeds. in spite of identifying glyphosate detoxifying genes from microorganisms, the plant mechanism to detoxify glyphosate has not been studied. we characterized an aldo-keto reductase gene from pseudomonas (psakr1) and rice (osakr1) and showed, by docking studies, both psakr1 and osakr1 can efficiently bind to glyphosat ...201627611904
soybean golgi snare 12 protein interacts with soybean mosaic virus encoded p3n-pipo protein.soybean mosaic virus (smv), a member of the potyvirus genus, is one of the most prevalent and devastating viral pathogens in soybean-growing regions worldwide. it is generally accepted that symptom development of a viral plant disease results from molecular interactions between the virus and its host plant. p3n-pipo, as a trans-frame protein consisting of the amino-terminal half of p3 fused to pipo of the potyvirus, plays a key role of viral cell-to-cell movement. this study provides evidence th ...201627553272
analysis of magnaporthe oryzae genome reveals a fungal effector, which is able to induce resistance response in transgenic rice line containing resistance gene, pi54.rice blast caused by magnaporthe oryzae is one of the most important diseases of rice. pi54, a rice gene that imparts resistance to m. oryzae isolates prevalent in india, was already cloned but its avirulent counterpart in the pathogen was not known. after decoding the whole genome of an avirulent isolate of m. oryzae, we predicted 11440 protein coding genes and then identified four candidate effector proteins which are exclusively expressed in the infectious structure, appresoria. in silico pro ...201627551285
identification, cloning and characterization of r2r3-myb gene family in canola (brassica napus l.) identify a novel member modulating ros accumulation and hypersensitive-like cell death.the r2r3-myb proteins comprise one of the largest families of transcription factors in plants. although genome-wide analysis of this family has been carried out in some plant species, little is known about r2r3-myb genes in canola (brassica napus l.). in this study, we have identified 76 r2r3-myb genes in the canola genome through mining of expressed sequence tags (ests). the cdna sequences of 44 myb genes were successfully cloned. the transcriptional activities of bnamyb proteins encoded by the ...201626800702
efficient in vitro and in vivo activity of glyco-engineered plant-produced rabies monoclonal antibodies e559 and 62-71-3.rabies is a neglected zoonotic disease that has no effective treatment after onset of illness. however the disease can be prevented effectively by prompt administration of post exposure prophylaxis which includes administration of passive immunizing antibodies (rabies immune globulin, rig). currently, human rig suffers from many restrictions including limited availability, batch-to batch inconsistencies and potential for contamination with blood-borne pathogens. anti-rabies monoclonal antibodies ...201627427976
a conserved puccinia striiformis protein interacts with wheat npr1 and reduces induction of pathogenesis-related genes in response to pathogens.in arabidopsis, npr1 is a key transcriptional coregulator of systemic acquired resistance. upon pathogen challenge, npr1 translocates from the cytoplasm to the nucleus, in which it interacts with tga-bzip transcription factors to activate the expression of several pathogenesis-related (pr) genes. in a screen of a yeast two-hybrid library from wheat leaves infected with puccinia striiformis f. sp. tritici, we identified a conserved rust protein that interacts with wheat npr1 and named it pnpi (fo ...201627898286
mir169 and its target paghap2-6 regulated by aba are involved in poplar cambium dormancy.dormancy is an effective strategy for perennial plants in temperate zones to survive the winter stress. micrornas (mirnas) have been well known as important regulators for various biological processes. in this study, we checked the expression of mir169 members in the cambium zone during dormancy and active growth in poplar and found that they had distinct expression patterns. we identified and characterized a dormancy-specific target gene of mir169, paghap2-6. 5' race assays confirmed the direct ...201627111502
tobacco ring e3 ligase ntrfp1 mediates ubiquitination and proteasomal degradation of a geminivirus-encoded βc1.the βc1 protein encoded by the tomato yellow leaf curl china virus-associated betasatellite functions as a pathogenicity determinant. to better understand the molecular basis whereby βc1 functions in pathogenicity, a yeast two-hybrid screen of a tobacco cdna library was carried out using βc1 as the bait. the screen revealed that βc1 interacts with a tobacco ring-finger protein designated ntrfp1, which was further confirmed by the bimolecular fluorescence complementation and co-immunoprecipitatio ...201627018391
potato nph3/rpt2-like protein stnrl1, targeted by a phytophthora infestans rxlr effector, is a susceptibility factor.plant pathogens deliver effectors to manipulate host processes. we know little about how fungal and oomycete effectors target host proteins to promote susceptibility, yet such knowledge is vital to understand crop disease. we show that either transient expression in nicotiana benthamiana, or stable transgenic expression in potato (solanum tuberosum), of the phytophthora infestans rxlr effector pi02860 enhances leaf colonization by the pathogen. expression of pi02860 also attenuates cell death tr ...201626966171
the grapevine uncharacterized intrinsic protein 1 (vvxip1) is regulated by drought stress and transports glycerol, hydrogen peroxide, heavy metals but not water.a mip (major intrinsic protein) subfamily called uncharacterized intrinsic proteins (xip) was recently described in several fungi and eudicot plants. in this work, we cloned a xip from grapevine, vvxip1, and agrobacterium-mediated transformation studies in nicotiana benthamiana revealed that the encoded aquaporin shows a preferential localization at the endoplasmic reticulum membrane. stopped-flow spectrometry in vesicles from the aqy-null yeast strain ysh1172 overexpressing vvxip1 showed that v ...201627504956
identification and regulation of host genes related to rice stripe virus symptom production.viral infections cause plant chlorosis, stunting, necrosis or other symptoms. the down-regulation of chloroplast-related genes (chrgs) is assumed to be responsible for chlorosis. we identified the differentially expressed genes (degs) in rice stripe virus (rsv)-infected nicotiana benthamiana, and examined the contribution of 75 down-regulated degs to rsv symptoms by silencing them one by one using tobacco rattle virus (trv)-induced gene silencing. silencing of 11 of the 75 down-regulated degs ca ...201626487490
the ralstonia solanacearum type iii effector ripay targets plant redox regulators to suppress immune responses.the subversion of plant cellular functions is essential for bacterial pathogens to proliferate in host plants and cause disease. most bacterial plant pathogens employ a type iii secretion system to inject type iii effector (t3e) proteins inside plant cells, where they contribute to the pathogen-induced alteration of plant physiology. in this work, we found that the ralstonia solanacearum t3e ripay suppresses plant immune responses triggered by bacterial elicitors and by the phytohormone salicyli ...201627768829
identification and characterization of privet leaf blotch-associated virus, a novel idaeovirus.a novel virus has been identified by next-generation sequencing (ngs) in privet (ligustrum japonicum l.) affected by a graft-transmissible disease characterized by leaf blotch symptoms resembling infectious variegation, a virus-like privet disease with an unclear aetiology. this virus, which has been tentatively named 'privet leaf blotch-associated virus' (prlbav), was absent in non-symptomatic privet plants, as revealed by ngs and reverse transcription-polymerase chain reaction (rt-pcr). molecu ...201627349357
root-mediated signal transmission of systemic acquired resistance against above-ground and below-ground pathogens.plants modulate defence signalling networks in response to various biotic stresses via inter-organ communications. the root-mediated transmission of systemic acquired resistance (sar) against soil-borne and air-borne plant pathogens from sar-induced plants to neighbouring plants subjected to local chemical and pathogen treatments was evaluated.201627555496
differential modulation of plant immune responses by diverse members of the pseudomonas savastanoi pv. savastanoi hopaf type iii effector family.the pseudomonas savastanoi pv. savastanoi ncppb 3335 type iii secretion system (t3ss) effector repertoire includes 33 candidates, seven of which translocate into host cells and interfere with plant defences. the present study was performed to investigate the co-existence of both plasmid- and chromosomal-encoded members of the hopaf effector family, hopaf1-1 and hopaf1-2, respectively, in the genome of ncppb 3335. here, we show that the hopaf1 paralogues are widely distributed in the pseudomonas ...201627116193
solanum venturii, a suitable model system for virus-induced gene silencing studies in potato reveals stmkk6 as an important player in plant immunity.virus-induced gene silencing (vigs) is an optimal tool for functional analysis of genes in plants, as the viral vector spreads throughout the plant and causes reduced expression of selected gene over the whole plant. potato (solanum tuberosum) is one of the most important food crops, therefore studies performing functional analysis of its genes are very important. however, the majority of potato cultivars used in laboratory experimental setups are not well amenable to available vigs systems, thu ...201627213007
autophagy induction in tobacco leaves infected by potato virus y(o) and its putative roles.autophagy plays a critical role in the innate immune response of plants to pathogen infection. in the present study, we examined autophagy induced by potato virus y ordinary strain (pvy(o)) infection in tobacco (nicotiana benthamiana). enzyme-linked immunosorbent assays revealed that the number of virus particles in the plant peaked at 2 weeks post-inoculation and then gradually decreased. additionally, the amount of virus increased significantly in the 3rd and 4th leaves distal to the inoculate ...201627137843
activation of senescence-associated dark-inducible (din) genes during infection contributes to enhanced susceptibility to plant viruses.virus infections in plants cause changes in host gene expression that are common to other environmental stresses. in this work, we found extensive overlap in the transcriptional responses between arabidopsis thaliana plants infected with tobacco rattle virus (trv) and plants undergoing senescence. this is exemplified by the up-regulation during infection of several senescence-associated dark-inducible (din) genes, including atdin1 (senescence 1, sen1), atdin6 (asparagine synthetase 1, atasn1) an ...201625787925
an efficient potato virus x -based microrna silencing in nicotiana benthamiana.plant micrornas (mirnas) play pivotal roles in many biological processes. although many mirnas have been identified in various plant species, the functions of these mirnas remain largely unknown due to the shortage of effective genetic tools to block their functional activity. recently, mirna target mimic (tm) technologies have been applied to perturb the activity of specific endogenous mirna or mirna families. we previously reported that tobacco rattle virus (trv)-based tm expression can succes ...201626837708
the stripe rust fungal effector pec6 suppresses pattern-triggered immunity in a host species-independent manner and interacts with adenosine kinases.we identified a wheat stripe rust (puccinia striiformis) effector candidate (pec6) with pattern-triggered immunity (pti) suppression function and its corresponding host target. pec6 compromised pti host species-independently. in nicotiana benthamiana, it hampers reactive oxygen species (ros) accumulation and callose deposition induced by pseudomonas fluorescens. in arabidopsis, plants expressing pec6 were more susceptible to pseudomonas syringae pv. tomato (pto) dc3000 δavrpto/δavrptob. in wheat ...201627252028
two homologous host proteins interact with potato virus x rnas and cps and affect viral replication and movement.because viruses encode only a small number of proteins, all steps of virus infection rely on specific interactions between viruses and hosts. we previously screened several nicotiana benthamiana (nb) proteins that interact with the stem-loop 1 (sl1) rna structure located at the 5' end of the potato virus x (pvx) genome. in this study, we characterized two of these proteins (nbcpip2a and nbcpip2b), which are homologous and are induced upon pvx infection. electrophoretic mobility shift assay confi ...201627353522
a novel block of plant virus movement genes.hibiscus green spot virus (hgsv) is a recently discovered and so far poorly characterized bacilliform plant virus with a positive-stranded rna genome consisting of three rna species. here, we demonstrate that the proteins encoded by the orf2 and orf3 in hgsv rna2 are necessary and sufficient to mediate cell-to-cell movement of transport-deficient potato virus x in nicotiana benthamiana. these two genes represent a specialized transport module called a 'binary movement block' (bmb), and orf2 and ...201627118327
a land plant-specific transcription factor directly enhances transcription of a pathogenic noncoding rna template by dna-dependent rna polymerase ii.some dna-dependent rna polymerases (ddrps) possess rna-dependent rna polymerase activity, as was first discovered in the replication of potato spindle tuber viroid (pstvd) rna genome in tomato (solanum lycopersicum). recent studies revealed that this activity in bacteria and mammals is important for transcriptional and posttranscriptional regulatory mechanisms. here, we used pstvd as a model to uncover auxiliary factors essential for rna-templated transcription by ddrp pstvd replication in the n ...201627113774
protein composition of 6k2-induced membrane structures formed during potato virus a infection.the definition of the precise molecular composition of membranous replication compartments is a key to understanding the mechanisms of virus multiplication. here, we set out to investigate the protein composition of the potyviral replication complexes. we purified the potyviral 6k2 protein-induced membranous structures from potato virus a (pva)-infected nicotiana benthamiana plants. for this purpose, the 6k2 protein, which is the main inducer of potyviral membrane rearrangements, was expressed i ...201626574906
the ire1/bzip60 pathway and bax inhibitor 1 suppress systemic accumulation of potyviruses and potexviruses in arabidopsis and nicotiana benthamiana plants.the inositol requiring enzyme (ire1) is an endoplasmic reticulum (er) stress sensor. when activated, it splices the bzip60 mrna, producing a truncated transcription factor that upregulates genes involved in the unfolded protein response. bax inhibitor 1 (bi-1) is another er stress sensor that regulates cell death in response to environmental assaults. the potyvirus 6k2 and potexvirus tgb3 proteins are known to reside in the er, serving, respectively, as anchors for the viral replicase and moveme ...201627578623
dissecting the secondary structure of the circular rna of a nuclear viroid in vivo: a "naked" rod-like conformation similar but not identical to that observed in vitro.with a minimal (250-400 nt), non-protein-coding, circular rna genome, viroids rely on sequence/structural motifs for replication and colonization of their host plants. these motifs are embedded in a compact secondary structure whose elucidation is crucial to understand how they function. viroid rna structure has been tackled in silico with algorithms searching for the conformation of minimal free energy, and in vitro by probing in solution with rnases, dimethyl sulphate and bisulphite, and with ...201627574720
artificial micrornas and synthetic trans-acting small interfering rnas interfere with viroid infection.artificial micrornas (amirnas) and synthetic trans-acting small interfering rnas (syn-tasirnas) are two classes of artificial small rnas (srnas) engineered to silence endogenous transcripts as well as viral rnas in plants. here, we explore the possibility of using amirnas and syn-tasirnas to specifically interfere with infections by viroids, small (250-400-nucleotide) non-coding circular rnas with compact secondary structure infecting a wide range of plant species. the combined use of recent hig ...201628026103
translationally controlled tumour protein (tctp) from tomato and nicotiana benthamiana is necessary for successful infection by a potyvirus.translationally controlled tumour protein (tctp) is a ubiquitously distributed protein in eukaryotes, involved in the regulation of several processes, including cell cycle progression, cell growth, stress protection, apoptosis and maintenance of genomic integrity. its expression is induced during the early stages of tomato (solanum lycopersicum) infection by the potyvirus pepper yellow mosaic virus (pepymv, a close relative of potato virus y). tomato tctp is a protein of 168 amino acids, which c ...201627159273
cell death triggering and effector recognition by sw-5 sd-cnl proteins from resistant and susceptible tomato isolines to tomato spotted wilt virus.only a limited number of dominant resistance genes acting against plant viruses have been cloned, and further functional studies of these have been almost entirely limited to the resistance genes rx against potato virus x (pvx) and n against tobacco mosaic virus (tmv). recently, the cell-to-cell movement protein (nsm ) of tomato spotted wilt virus (tswv) has been identified as the avirulence determinant (avr) of sw-5b-mediated resistance, a dominant resistance gene which belongs to the class of ...201627271212
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