Publications

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post-embedding in situ hybridization for localization of viral nucleic acid in ultra-thin sections.we developed an in situ hybridization technique for localization of plant virus rna in ultra-thin sections of virus-infected, lowicryl hm20- or osmium-fixed, araldite-embedded plant leaf tissues. a digoxigenin-labeled in vitro transcript corresponding to 173 nucleotides at the 3' end of bamboo mosaic virus (bamv) rna was used as a riboprobe and the hybrids were detected by incubation with sheep antidigoxigenin antibody followed by gold-labeled rabbit anti-sheep igg. pre-treatment of ultra-thin s ...19938245409
nucleotide sequence of the genomic rna of bamboo mosaic potexvirus.the complete nucleotide sequence of the genomic rna of bamboo mosaic virus (bamv) was determined by sequencing a set of overlapping cdna clones and by direct sequencing of the viral rna. the rna genome of bamv is 6366 nucleotides long [excluding 3'poly(a) tail] and contains six open reading frames (orfs 1 to 6) coding for polypeptides with m(r) values of 155k, 28k, 13k, 6k, 25k and 14k, respectively. the genome organization and sizes of the encoded proteins are very similar to those of other pot ...19948077956
identification and characterization of the escherichia coli-expressed rna-dependent rna polymerase of bamboo mosaic virus.bamboo mosaic virus (bamv), a member of the potexvirus group, infects primarily members of the bambusoideae. the open reading frame 1 (orf1) of bamv encodes a 155-kda polypeptide that was postulated to be involved in the replication and the formation of cap structure at the 5' end of the viral genome. to characterize the activities associated with the 155-kda viral protein, it was expressed in escherichia coli bl21(de3) cells with thioredoxin, hexahistidine, and s. tag fused consecutively at its ...19989811749
evolution of bamboo mosaic virus in a nonsystemic host results in mutations in the helicase-like domain that cause reduced rna accumulation.a mutant population of bamboo mosaic potexvirus (bamv) was isolated after serial passage using chenopodium quinoa plants. while the wild type inoculum induced indistinct chlorotic lesions, the mutant produced obvious lesions on c. quinoa although rna accumulation of the mutant in nicotiana benthamiana protoplasts was significantly reduced compared to wild type. mutations were identified in the helicase-like domain. one rt-pcr-generated cdna clone (designated pl1-33) representing the helicase-lik ...19989879769
a defective rna associated with bamboo mosaic virus and the possible common mechanisms for rna recombination in potexviruses.a naturally occurring 1.1 kb rna was isolated from purified virions of bamboo mosaic potexvirus isolate s (bamv-s). this rna is a defective rna (d rna) derived from a single internal deletion of the bamv genome. a cdna clone representing the complete nucleotide sequence of the bamv-s d rna was generated and its nucleotide sequence was determined. the bamv d cdna is 1015 nts in length [excluding the poly(a) tail] and consists of two regions corresponding to 867 nts of the 5' terminus and 148 nts ...199910403698
characterization of the adomet-dependent guanylyltransferase activity that is associated with the n terminus of bamboo mosaic virus replicase.bamboo mosaic virus (bamv), a member of the potexvirus group, infects primarily members of the bambusoideae. open reading frame 1 (orf1) of bamv encodes a 155-kda polypeptide that has long been postulated to be a replicase involved in the replication and formation of the cap structure at the 5' end of the viral genome. to identify and characterize the enzymatic activities associated with the n-terminal domain of the bamv orf1 protein, the intact replicase and two c-terminally truncated proteins ...200111134291
virus-specific mrna capping enzyme encoded by hepatitis e virus.hepatitis e virus (hev), a positive-strand rna virus, is an important causative agent of waterborne hepatitis. expression of cdna (encoding amino acids 1 to 979 of hev nonstructural open reading frame 1) in insect cells resulted in synthesis of a 110-kda protein (p110), a fraction of which was proteolytically processed to an 80-kda protein. p110 was tightly bound to cytoplasmic membranes, from which it could be released by detergents. immunopurified p110 catalyzed transfer of a methyl group from ...200111413290
the helicase-like domain of plant potexvirus replicase participates in formation of rna 5' cap structure by exhibiting rna 5'-triphosphatase activity.open reading frame 1 (orf1) of potexviruses encodes a viral replicase comprising three functional domains: a capping enzyme at the n terminus, a putative helicase in the middle, and a polymerase at the c terminus. to verify the enzymatic activities associated with the putative helicase domain, the corresponding cdna fragment from bamboo mosaic virus (bamv) was cloned into vector pet32 and the protein was expressed in escherichia coli and purified by metal affinity chromatography. an activity ass ...200111711602
structural and mutational analyses of cis-acting sequences in the 5'-untranslated region of satellite rna of bamboo mosaic potexvirus.the satellite rna of bamboo mosaic virus (satbamv) contains on open reading frame for a 20-kda protein that is flanked by a 5'-untranslated region (utr) of 159 nucleotides (nt) and a 3'-utr of 129 nt. a secondary structure was predicted for the 5'-utr of satbamv rna, which folds into a large stem-loop (lsl) and a small stem-loop. enzymatic probing confirmed the existence of lsl (nt 8-138) in the 5'-utr. the essential cis-acting sequences in the 5'-utr required for satbamv rna replication were de ...200312832220
arg-16 and arg-21 in the n-terminal region of the triple-gene-block protein 1 of bamboo mosaic virus are essential for virus movement.the protein encoded by the first gene of the triple gene block (tgbp1) of potexviruses is required for movement of the viruses. it has been reported that single arg-->ala substitutions at position 11, 16 or 21 of tgbp1 of bamboo mosaic virus (bamv) eliminate its rna-binding activity, while substitutions at position 16 or 21 only affect its ntpase activity (liou et al., virology 277, 336-344, 2000). however, it remains unclear whether these arg-->ala substitutions also affect the movement of bamv ...200414718640
critical residues for gtp methylation and formation of the covalent m7gmp-enzyme intermediate in the capping enzyme domain of bamboo mosaic virus.open reading frame 1 of bamboo mosaic virus (bamv), a potexvirus in the alphavirus-like superfamily, encodes a 155-kda replicase responsible for the formation of the 5' cap structure and replication of the viral rna genome. the n-terminal domain of the viral replicase functions as an mrna capping enzyme, which exhibits both gtp methyltransferase and s-adenosylmethionine (adomet)-dependent guanylyltransferase activities. we mutated each of the four conserved amino acids among the capping enzymes ...200414722282
complete nucleotide sequence and genome organization of a cactus virus x strain from hylocereus undatus (cactaceae).the complete nucleotide sequence of a strain of cactus virus x (cvx-hu) isolated from hylocereus undatus (cactaceae) has been determined. excluding the poly(a) tail, the sequence is 6614 nucleotides in length and contains seven open reading frames (orfs). the genome organization of cvx is similar to that of other potexviruses. orf1 encodes the putative viral replicase with conserved methyltransferase, helicase, and polymerase motifs. within orf1, two other orfs were located separately in the +2 ...200415098117
a conserved secondary structure in the hypervariable region at the 5' end of bamboo mosaic virus satellite rna is functionally interchangeable.satellite rna (satrna) associated with bamboo mosaic virus (bamv) is dependent on bamv for replication and encapsidation. molecular analyses of total rna extracted from bamboo species collected worldwide revealed that 26 out of 61 bamv isolates harbored satbamv. among them, two phylogenetically distinguishable groups, a and b, with a genetic diversity of 6.9 +/- 0.7% were identified. greatest sequence diversity occurred in the 5' untranslated region (utr) that contained one hypervariable region ...200415527838
mrna guanylation catalyzed by the s-adenosylmethionine-dependent guanylyltransferase of bamboo mosaic virus.the s-adenosylmethionine-dependent guanylyltransferase of bamboo mosaic virus belongs to a novel class of mrna capping enzymes distantly conserved in alphavirus-like superfamily. the reaction sequence of the viral enzyme has been proposed comprising steps of 1) binding of gtp and s-adenosylmethionine, 2) formation of m7gtp and s-adenosylhomocysteine, 3) formation of the covalent (enzyme-m7gmp) intermediate, and 4) transfer of m7gmp from the intermediate to the rna acceptor. in this study the acc ...200515677480
characterization of the infectivity of bamboo mosaic virus with its correlation to the in vitro replicase activities in nicotiana benthamiana.in vitro rna-dependent rna polymerase (rdrp) transcription assay share an extremely useful system for studying the molecular mechanisms of replication of positive-sense rna viruses such as bamboo mosaic virus (bamv). however, the obstacle encountered in this system is the inconsistency in the enzyme activity and the template specificity among different batches of the rdrp extracts. in order to overcome this obstacle, we designed experiments to study the functional dynamics of the bamv rdrp in te ...200515904988
structural and functional analysis of the cis-acting elements required for plus-strand rna synthesis of bamboo mosaic virus.bamboo mosaic virus (bamv) has a single-stranded positive-sense rna genome. the secondary structure of the 3'-terminal sequence of the minus-strand rna has been predicted by mfold and confirmed by enzymatic structural probing to consist of a large, stable stem-loop and a small, unstable stem-loop. to identify the promoter for plus-strand rna synthesis in this region, transcripts of 39, 77, and 173 nucleotides (ba-39, ba-77, and ba-173, respectively) derived from the 3' terminus of the minus-stra ...200515994798
the aauaaa motif of bamboo mosaic virus rna is involved in minus-strand rna synthesis and plus-strand rna polyadenylation.bamboo mosaic virus (bamv) has a single-stranded positive-sense rna genome with a 5'-cap structure and a 3' poly(a) tail. deleting the internal loop that contains the putative polyadenylation signal (aauaaa) in the 3' untranslated region (utr) of bamv genomic rna appeared to diminish coat protein accumulation to 2% (c. p. cheng and c. h. tsai, j. mol. biol. 288:555-565, 1999). to investigate the function of the aauaaa motif, mutations were introduced into an infectious bamv cdna at each residue ...200516282455
crucial role of the 5' conserved structure of bamboo mosaic virus satellite rna in downregulation of helper viral rna replication.satellite rna of bamboo mosaic virus (satbamv), a single-stranded mrna type satellite encoding a protein of 20 kda (p20), depends on the helper bamv for replication and encapsidation. two satbamv isolates, bsf4 and bsl6, exhibit distinctly differential phenotypes in nicotiana benthamiana plants when coinoculated with bamv rna. bsl6 significantly reduces bamv rna replication and suppresses the bamv-induced symptoms, whereas bsf4 does not. by studies with chimeric satbamvs generated by exchanging ...200616474162
movement of potexviruses requires species-specific interactions among the cognate triple gene block proteins, as revealed by a trans-complementation assay based on the bamboo mosaic virus satellite rna-mediated expression system.the intra- and intercellular transport of potexviruses require interactions among viral rna, coat protein and elements of the triple gene block proteins (tgbps). in this study, the requirement of bamboo mosaic virus (bamv) tgbps for movement functions and the compatibilities with those of two potexviruses, potato virus x (pvx) and foxtail mosaic virus (fomv), were examined using a satellite rna-mediated trans-complementation assay system. single or multiple tgbps of bamv, pvx and fomv were expre ...200616603539
the arginine-rich motif of bamboo mosaic virus satellite rna-encoded p20 mediates self-interaction, intracellular targeting, and cell-to-cell movement.satellite rna of bamboo mosaic virus (satbamv) has a single open reading frame for a nonstructural, rna-binding protein, p20, which facilitates the long-distance movement of satbamv in nicotiana benthamiana. here, we elucidate various biological properties of p20 and the involvement of a single domain in its activities. p20 displayed a strong self-interaction in vitro and in vivo, and cross-linking assays demonstrated its oligomerization. domain mapping, using the bacterial two-hybrid system, in ...200616838788
chloroplast phosphoglycerate kinase, a gluconeogenetic enzyme, is required for efficient accumulation of bamboo mosaic virus.the tertiary structure in the 3'-untranslated region (3'-utr) of bamboo mosaic virus (bamv) rna is known to be involved in minus-strand rna synthesis. proteins found in the rna-dependent rna polymerase (rdrp) fraction of bamv-infected leaves interact with the radio labeled 3'-utr probe in electrophoretic mobility shift assays (emsa). results derived from the ultraviolet (uv) cross-linking competition assays suggested that two cellular factors, p43 and p51, interact specifically with the 3'-utr o ...200717169994
downregulation of bamboo mosaic virus replication requires the 5' apical hairpin stem loop structure and sequence of satellite rna.satellite rnas associated with bamboo mosaic virus (satbamv) exhibit different phenotypes. some isolates could reduce the accumulation of bamv rna and attenuate the bamv-induced symptoms in co-inoculated plants. the determinants of the downregulation of bamv replication were mapped in the 5' hypervariable region of satbamv, which folds into a conserved apical hairpin stem loop (ahsl) structure comprising an apical loop and two internal loops, as evidenced by enzymatic probing. we also demonstrat ...200717482233
mutational analysis of a helicase motif-based rna 5'-triphosphatase/ntpase from bamboo mosaic virus.the helicase-like domain of bamv replicase possesses ntpase and rna 5'-triphosphatase activities. in this study, mutational effects of the helicase signature motifs and residue l543 on the two activities were investigated. either activity was inactivated by k643a-s644a, d702a, d730a, r855a, or l543p mutations. on the other hand, q826a, d858a and l543a had activities, in terms of k(cat)/k(m), reduced by 5- to 15-fold. amppnp, a nonhydrolyzable atp analogue, competitively inhibited rna 5'-triphosp ...200717585982
induction of protective immunity in swine by recombinant bamboo mosaic virus expressing foot-and-mouth disease virus epitopes.plant viruses can be employed as versatile vectors for the production of vaccines by expressing immunogenic epitopes on the surface of chimeric viral particles. although several viruses, including tobacco mosaic virus, potato virus x and cowpea mosaic virus, have been developed as vectors, we aimed to develop a new viral vaccine delivery system, a bamboo mosaic virus (bamv), that would carry larger transgene loads, and generate better immunity in the target animals with fewer adverse environment ...200717900346
in vitro replication of bamboo mosaic virus satellite rna.an in vitro system was applied to analyze the replication of a satellite rna of bamboo mosaic virus (bamv), designated satbamv rna, using solubilized membrane-bound rna-dependent rna polymerase (rdrp) complexes isolated from bamv-infected nicotiana benthamiana. after removal of endogenous templates, the rdrp complexes of bamv catalyzed rna synthesis upon the addition of the full-length positive (+)- or negative (-)-strand satbamv rna transcripts used as templates. both (+)- and (-)-satbamv rna p ...200818538884
topological properties of the triple gene block protein 2 of bamboo mosaic virus.the triple gene block protein 2 (tgbp2) of bamboo mosaic virus (bamv) has been proposed to be a transmembrane protein; however, its features remain unclear. here, we used biochemical approaches to determine its topological properties. our data reveal that tgbp2 is mainly associated with the endoplasmic reticulum membrane. the resistance of tgbp2 in proteoliposomes, prepared from both the bamv-infected tissues and in vitro reconstitution system, to both chemical extraction and trypsin digestion c ...200818639913
subcellular localization and expression of bamboo mosaic virus satellite rna-encoded protein.the satellite rna of bamboo mosaic virus (satbamv) has a single open reading frame encoding a non-structural protein, p20, which facilitates long-distance movement of satbamv in bamv and satbamv co-infected plants. immunohistochemistry and immunoelectron microscopy revealed that the p20 protein accumulated in the cytoplasm and nuclei in co-infected cells. p20 and the helper virus coat protein (cp) were highly similar in their subcellular localization, except that aggregates of bamv virions were ...200919141462
structural and functional analyses of the 3' untranslated region of bamboo mosaic virus satellite rna.the 3'-untranslated region (utr) of rna genomes of viruses and satellite rnas plays essential roles in viral replication and transcription. the structural features of the 3'-utr of the satellite rna of bamboo mosaic virus (satbamv) involved in its replication were analyzed in this study. by the use of enzymatic probing, the secondary structure of satbamv 3'-utr was confirmed to comprise two small stem-loops (sla and slb), one large stem-loop (slc), and a poly(a) tail of mainly 75-200 adenylate r ...200919201437
suppression of bamboo mosaic virus accumulation by a putative methyltransferase in nicotiana benthamiana.bamboo mosaic virus (bamv) is a 6.4-kb positive-sense rna virus belonging to the genus potexvirus of the family flexiviridae. the 155-kda viral replicase, the product of orf1, comprises an n-terminal s-adenosyl-l-methionine (adomet)-dependent guanylyltransferase, a nucleoside triphosphatase/rna 5'-triphosphatase, and a c-terminal rna-dependent rna polymerase (rdrp). to search for cellular factors potentially involved in the regulation of replication and/or transcription of bamv, the viral rdrp d ...200919297487
characterization of the rna-binding properties of the triple-gene-block protein 2 of bamboo mosaic virus.the triple-gene-block protein 2 (tgbp2) of bamboo mosaic virus (bamv) is a transmembrane protein which was proposed to be involved in viral rna binding during virus transport. here, we report on the rna-binding properties of tgbp2. using tyrosine fluorescence spectroscopy and uv-crosslinking assays, the tgbp2 solubilized with triton x-100 was found to interact with viral rna in a non-specific manner. these results raise the possibility that tgbp2 facilitates intracellular delivery of viral rna t ...200919422690
identification and functional characterization of regions that can be crosslinked to rna in the helicase-like domain of bamv replicase.the helicase-like domain of the bamboo mosaic virus replicase catalyzes the release of 5'-gamma-phosphate from both atp and 5'-triphosphated rna by an identical set of catalytic residues with a presumably larger binding pocket for rna. in this study, the peptidyl regions involved in rna binding were mapped by reversible formaldehyde crosslinking and mass spectrometry. eleven residues within these regions were examined by mutational analysis. h636a, y704a, and k706a greatly diminished the enzymat ...200919443005
the two conserved cysteine residues of the triple gene block protein 2 are critical for both cell-to-cell and systemic movement of bamboo mosaic virus.the triple gene block protein 2 (tgbp2) of bamboo mosaic virus (bamv) is a transmembrane protein which is known to be required for the cell-to-cell movement of potexviruses. this protein has two conserved cys residues, cys-109 and cys-112, at its c-terminal tail, which is supposed to be exposed on the outer surface of the endoplasmic reticulum (er) membrane and er-derived granular vesicles. in this study, we investigated the importance of these two cys residues on the cell-to-cell and systemic m ...200919810807
structural homology between bamboo mosaic virus and its satellite rnas in the 5'untranslated region.a structural element was identified in the 5'-proximal sequence of the bamboo mosaic virus (bamv) rna. mutational analysis of the hairpin showed that disruptions of the secondary structure or substitutions of the loop sequences resulted in reduced accumulation of bamv genomic rna. phylogenetic analysis further suggested the presence of structural homologues of this hairpin in all other potexviruses. in addition, remarkable structural homology was discovered between the bamv hairpin and a stem-lo ...201019906937
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 ...201020447270
a unique glycine-rich motif at the n-terminal region of bamboo mosaic virus coat protein is required for symptom expression.the coat proteins (cp) of many plant viruses are multifunctional proteins. we used n-terminal sequencing and mass spectrometry/mass spectrometry analysis to identify a truncated form of the bamboo mosaic virus (bamv) cp missing the n-terminal 35 amino acids (n35). the n35 region is unique in the potexviruses by its containing a glycine-rich motif (grm) not present in databases but highly conserved among bamv isolates. results from site-directed mutagenesis and deletion mutational analysis showed ...201020521953
global analyses of small interfering rnas derived from bamboo mosaic virus and its associated satellite rnas in different plants.satellite rnas (satrnas), virus parasites, are exclusively associated with plant virus infection and have attracted much interest over the last 3 decades. upon virus infection, virus-specific small interfering rnas (vsirnas) are produced by dicer-like (dcl) endoribonucleases for anti-viral defense. the composition of vsirnas has been studied extensively; however, studies of satrna-derived sirnas (satsirnas) or sirna profiles after satrna co-infection are limited. here, we report on the small rna ...201020689857
the silencing suppressor p25 of potato virus x interacts with argonaute1 and mediates its degradation through the proteasome pathway.previous evidence has indicated that the p25 protein encoded by potato virus x (pvx) inhibits either the assembly or function of the effector complexes in the rna silencing-based antiviral defence system (bayne et al., cell-to-cell movement of potato potexvirus x is dependent on suppression of rna silencing. plant j.44, 471-482). this finding prompted us to investigate the possibility that p25 targets the argonaute (ago) effector nuclease of rna silencing. co-immunoprecipitation and western blot ...201020696002
identification of differentially expressed genes induced by bamboo mosaic virus infection in nicotiana benthamiana by cdna-amplified fragment length polymorphism.the genes of plants can be up- or down-regulated during viral infection to influence the replication of viruses. identification of these differentially expressed genes could shed light on the defense systems employed by plants and the mechanisms involved in the adaption of viruses to plant cells. differential gene expression in nicotiana benthamiana plants in response to infection with bamboo mosaic virus (bamv) was revealed using cdna-amplified fragment length polymorphism (aflp).201021184690
mutational effects of the consensus aromatic residues in the mrna capping domain of bamboo mosaic virus on gtp methylation and virus accumulation.rna viruses classified in the alphavirus-like superfamily possess a distinct capping domain, catalyzing gtp methylation and subsequent transfer of the m(7)gmp moiety from m(7)gtp to the 5'-diphosphate end of viral rna. the h68a mutation in the capping domain of bamboo mosaic virus enhanced gtp methylation but disabled the following transguanylation, making it possible to characterize the enzyme's methyltransferase activity separately. to explore the involvement of aromatic amino acids in substra ...201121227477
glyceraldehyde 3-phosphate dehydrogenase negatively regulates the replication of bamboo mosaic virus and its associated satellite rna.the identification of cellular proteins associated with virus replicase complexes is crucial to our understanding of virus-host interactions, influencing the host range, replication, and virulence of viruses. a previous in vitro study has demonstrated that partially purified bamboo mosaic virus (bamv) replicase complexes can be employed for the replication of both bamv genomic and satellite bamv (satbamv) rnas. in this study, we investigated the bamv and satbamv 3' untranslated region (utr) bind ...201121715476
the interaction between bamboo mosaic virus replication protein and coat protein is critical for virus movement in plant hosts.bamboo mosaic virus (bamv) is a positive-sense rna virus belonging to the genus potexvirus. open reading frame 1 (orf1) encodes the viral replication protein that consists of a capping enzyme domain, a helicase-like domain (hld), and an rna-dependent rna polymerase domain from the n to c terminus. orf5 encodes the viral coat protein (cp) required for genome encapsidation and the virus movement in plants. in this study, application of a yeast-two hybrid assay detected an interaction between the v ...201121917973
phosphorylation of bamboo mosaic virus satellite rna (satbamv)-encoded protein p20 downregulates the formation of satbamv-p20 ribonucleoprotein complex.bamboo mosaic virus (bamv) satellite rna (satbamv) depends on bamv for its replication and encapsidation. satbamv-encoded p20 protein is an rna-binding protein that facilitates satbamv systemic movement in co-infected plants. here, we examined phosphorylation of p20 and its regulatory functions. recombinant p20 (rp20) was phosphorylated by host cellular kinase(s) in vitro, and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry and mutational analyses revealed ser-11 as ...201121965537
hsp90 interacts specifically with viral rna and differentially regulates replication initiation of bamboo mosaic virus and associated satellite rna.host factors play crucial roles in the replication of plus-strand rna viruses. in this report, a heat shock protein 90 homologue of nicotiana benthamiana, nbhsp90, was identified in association with partially purified replicase complexes from bamv-infected tissue, and shown to specifically interact with the 3' untranslated region (3' utr) of bamv genomic rna, but not with the 3' utr of bamv-associated satellite rna (satbamv rna) or that of genomic rna of other viruses, such as potato virus x (pv ...201222654666
functional analysis of the conserved histidine residue of bamboo mosaic virus capping enzyme in the activity for the formation of the covalent enzyme-m7gmp intermediate.the alphavirus-like mrna capping enzyme of bamboo mosaic virus (bamv) exhibits an adomet-dependent guanylyltransferase activity by which the methyl group of adomet is transferred to gtp, leading to the formation of m(7)gtp, and the m(7)gmp moiety is next transferred to the 5' end of pprna via a covalent enzyme-m(7)gmp intermediate. the function of the conserved h68 of the bamv capping enzyme in the intermediate formation was analyzed by mutagenesis in this study. the nature of the bond linking t ...201222641040
induction of protective immunity in chickens immunized with plant-made chimeric bamboo mosaic virus particles expressing very virulent infectious bursal disease virus antigen.very virulent infectious bursal disease virus (vvibdv) causes a highly contagious disease in young chickens and leads to significant economic loss in the poultry industry. effective new vaccines are urgently needed. autonomously replicating plant virus-based vector provides attractive means for producing chimeric virus particles (cvps) in plants that can be developed into vaccines. in this study, we demonstrate the potential for vaccine development of bamboo mosaic virus (bamv) epitope-presentat ...201222406128
the conserved 5' apical hairpin stem loops of bamboo mosaic virus and its satellite rna contribute to replication competence.satellite rnas associated with bamboo mosaic virus (satbamvs) depend on bamv for replication and encapsidation. certain satbamvs isolated from natural fields significantly interfere with bamv replication. the 5' apical hairpin stem loop (ahsl) of satbamv is the major determinant in interference with bamv replication. in this study, by in vivo competition assay, we revealed that the sequence and structure of ahsl, along with specific nucleotides (c(60) and c(83)) required for interference with ba ...201222278884
a putative rab-gtpase activation protein from nicotiana benthamiana is important for bamboo mosaic virus intercellular movement.the cdna-amplified fragment length polymorphism technique was applied to isolate the differentially expressed genes during bamboo mosaic virus (bamv) infection on nicotiana benthamiana plants. one of the upregulated genes was cloned and predicted to contain a tbc domain designated as nbrabgap1 (rab gtpase activation protein 1). no significant difference was observed in bamv accumulation in the nbrabgap1-knockdown and the control protoplasts. however, bamv accumulation was 50% and 2% in the inocu ...201324210126
chloroplast phosphoglycerate kinase is involved in the targeting of bamboo mosaic virus to chloroplasts in nicotiana benthamiana plants.the bamboo mosaic virus (bamv) is a positive-sense, single-stranded rna virus. previously, we identified that the chloroplast phosphoglycerate kinase (chl-pgk) from nicotiana benthamiana is one of the viral rna binding proteins involved in the bamv infection cycle. because chl-pgk is transported to the chloroplast, we hypothesized that chl-pgk might be involved in viral rna localization in the chloroplasts. to test this hypothesis, we constructed two green fluorescent protein (gfp)-fused misloca ...201324154620
maintaining the structural integrity of the bamboo mosaic virus 3' untranslated region is necessary for retaining the catalytic constant for minus-strand rna synthesis.bamboo mosaic virus (bamv) and the potato virus x (pvx) are members of the genus potexvirus and have a single-stranded positive-sense rna genome. the 3'-untranslated region (utr) of the bamv rna genome was mapped structurally into abc (a cloverleaf-like), d (a stem-loop), and e (pseudoknot) domains. the bamv replicase complex that was isolated from the infected plants was able to recognize the 3' utr of pvx rna to initiate minus-strand rna synthesis in vitro.201323800142
characterization of the polyadenylation activity in a replicase complex from bamboo mosaic virus-infected nicotiana benthamiana plants.bamboo mosaic virus (bamv) has a positive-sense single-stranded rna genome with a 5' cap and a 3' poly(a) tail. to characterize polyadenylation activity in the bamv replicase complex, we performed the in vitro polyadenylation with various bamv templates. we conducted a polyadenylation activity assay for bamv rna by using a partially purified bamv replicase complex. the results showed that approximately 200 adenylates at the 3' end of the rna were generated on the endogenous rna templates. specif ...201323768785
the stable association of virion with the triple-gene-block protein 3-based complex of bamboo mosaic virus.the triple-gene-block protein 3 (tgbp3) of bamboo mosaic virus (bamv) is an integral endoplasmic reticulum (er) membrane protein which is assumed to form a membrane complex to deliver the virus intracellularly. however, the virus entity that is delivered to plasmodesmata (pd) and its association with tgbp3-based complexes are not known. results from chemical extraction and partial proteolysis of tgbp3 in membrane vesicles revealed that tgbp3 has a right-side-out membrane topology; i.e., tgbp3 ha ...201323754943
the glutathione transferase of nicotiana benthamiana nbgstu4 plays a role in regulating the early replication of bamboo mosaic virus.bamboo mosaic virus (bamv) is a single-stranded positive-sense rna virus. one of the plant glutathione s-transferase (gst) genes, nbgstu4, responds as an upregulated gene in nicotiana benthamiana post bamv infection. in order to identify the role of nbgstu4 in bamv infection, the expression of nbgstu4 was knocked down using a virus-induced gene silencing technique or was transiently expressed in n. benthamiana in bamv inoculation. the results show a significant decrease in bamv rna accumulation ...201323701112
transgenic resistance to bamboo mosaic virus by expression of interfering satellite rna.plant genetic engineering has broadened the options for plant virus resistance and is mostly based on pathogen-derived resistance. previously, we have shown that interfering satellite rna (satrna) of bamboo mosaic virus (satbamv) greatly reduces bamboo mosaic virus (bamv) accumulation and bamv-induced symptoms in co-inoculated plants. here, we generated a nonviral source of virus-resistant transgenic nicotiana benthamiana and arabidopsis thaliana by introducing interfering satbamv. asymptomatic ...201323675895
ser/thr kinase-like protein of nicotiana benthamiana is involved in the cell-to-cell movement of bamboo mosaic virus.to investigate the plant genes affected by bamboo mosaic virus (bamv) infection, we applied a cdna-amplified fragment length polymorphism technique to screen genes with differential expression. a serine/threonine kinase-like (nbstkl) gene of nicotiana benthamiana is upregulated after bamv infection. nbstkl contains the homologous domain of ser/thr kinase. knocking down the expression of nbstkl by virus-induced gene silencing reduced the accumulation of bamv in the inoculated leaves but not in th ...201323646157
genetic diversity and evolution of satellite rnas associated with the bamboo mosaic virus.satellite rnas (satrnas) are subviral agents that depend on cognate helper viruses for genome replication and encapsidation. their negative impacts on helper viruses have been exploited to control plant viral diseases. satbamv is a commonly found satrna associated with bamboo mosaic virus (bamv) that infects diverse bamboo species in the field. to investigate the genetic diversity and evolution of satrnas, we examined seven satbamv populations derived from five bamboo species and cultivars from ...201425275532
potato virus x-based expression vectors are stabilized for long-term production of proteins and larger inserts.plus-strand rna viruses such as potato virus x (pvx) are often used as high-yielding expression vectors in plants, because they tolerate extra transgene insertion and expression without disrupting normal virus functions. however, sequence redundancy due to promoter duplication often leads to genetic instability. although heterologous subgenomic promoter-like sequences (sgps) have been successfully used in tobacco mosaic virus vectors, only homologous sgp duplications have been used in pvx vector ...201425171768
phosphorylation of coat protein by protein kinase ck2 regulates cell-to-cell movement of bamboo mosaic virus through modulating rna binding.in this study, we investigated the fine regulation of cell-to-cell movement of bamboo mosaic virus (bamv). we report that the coat protein (cp) of bamv is phosphorylated in planta at position serine 241 (s241), in a process involving nicotiana benthamiana casein kinase 2α (nbck2α). bamv cp and nbck2α colocalize at the plasmodesmata, suggesting that phosphorylation of bamv may be involved in its movement. s241 was mutated to examine the effects of temporal and spatial dysregulation of phosphoryla ...201425025779
two key arginine residues in the coat protein of bamboo mosaic virus differentially affect the accumulation of viral genomic and subgenomic rnas.the interactions between viral rnas and coat proteins (cps) are critical for the efficient completion of infection cycles of rna viruses. however, the specificity of the interactions between cps and genomic or subgenomic rnas remains poorly understood. in this study, bamboo mosaic virus (bamv) was used to analyse such interactions. using reversible formaldehyde cross-linking and mass spectrometry, two regions in cp, each containing a basic amino acid (r99 and r227, respectively), were identified ...201424393453
a dual gene-silencing vector system for monocot and dicot plants.plant virus-based gene-silencing vectors have been extensively and successfully used to elucidate functional genomics in plants. however, only limited virus-induced gene-silencing (vigs) vectors can be used in both monocot and dicot plants. here, we established a dual gene-silencing vector system based on bamboo mosaic virus (bamv) and its satellite rna (satbamv). both bamv and satbamv vectors could effectively silence endogenous genes in nicotiana benthamiana and brachypodium distachyon. the sa ...201424283212
the abscisic acid pathway has multifaceted effects on the accumulation of bamboo mosaic virus.accepted 29 october 2013. abscisic acid (aba) plays a key role in modulating plant responses to different biotic and abiotic stresses. however, the effect of aba on virus infection is not fully understood. here, we describe the effects of the aba pathway on the accumulation of bamboo mosaic virus (bamv) and cucumber mosaic virus (cmv) in two different hosts: arabidopsis thaliana and nicotiana benthamiana. we report that aba2 plays a critical role in the accumulation of bamv and cmv. mutants down ...201424224533
the molecular basis for flexibility in the flexible filamentous plant viruses.flexible filamentous plant viruses cause more than half the viral crop damage in the world but are also potentially useful for biotechnology. structural studies began more than 75 years ago but have failed, owing to the virion's extreme flexibility. we have used cryo-em to generate an atomic model for bamboo mosaic virus, which reveals flexible n- and c-terminal extensions that allow deformation while still maintaining structural integrity.201526167882
viral elements and host cellular proteins in intercellular movement of bamboo mosaic virus.as a member of the genus potexvirus, bamboo mosaic virus (bamv) also belongs to the plant viruses that encode triple gene block proteins (tgbps) for intercellular movement within the host plants. recent studies of the movement mechanisms of bamv have revealed similarities and differences between bamv and other potexviruses. this review focuses on the general aspects of viral and host elements involved in bamv movement, the interactions among these elements, and the possible pathways for intra- a ...201525951346
a transgenic plant cell-suspension system for expression of epitopes on chimeric bamboo mosaic virus particles.we describe a novel strategy to produce vaccine antigens using a plant cell-suspension culture system in lieu of the conventional bacterial or animal cell-culture systems. we generated transgenic cell-suspension cultures from nicotiana benthamiana leaves carrying wild-type or chimeric bamboo mosaic virus (bamv) expression constructs encoding the viral protein 1 (vp1) epitope of foot-and-mouth disease virus (fmdv). antigens accumulated to high levels in bdt38 and bdt19 transgenic cell lines co-ex ...201625879277
molecular characterization and detection of a recombinant isolate of bamboo mosaic virus from china.the complete genome sequences of three isolates of bamboo mosaic virus (bamv) from mainland china were determined and compared to those of bamv isolates from taiwan. sequence analysis showed that isolate bamv-jxybz1 from fuzhou shares 98 % nucleotide sequence identity with bamv-ythsl14 from nucleotides 2586 to 6306, and more than 94 % nucleotide sequence identity with bamv-muzhubz2 in other regions. recombination and phylogenetic analyses indicate that bamv-jxybz1 is a recombinant with one recom ...201626923929
nbrabg3f, a member of rab gtpase, is involved in bamboo mosaic virus infection in nicotiana benthamiana.the screening of differentially expressed genes in plants after pathogen infection can uncover the potential host factors required for the pathogens. in this study, an up-regulated gene was identified and cloned from nicotiana benthamiana plants after bamboo mosaic virus (bamv) inoculation. the up-regulated gene was identified as a member of the rab small guanosine triphosphatase (gtpase) family, and was designated as nbrabg3f according to its in silico translated product with high identity to t ...201626416342
promotion of bamboo mosaic virus accumulation in nicotiana benthamiana by 5'→3' exonuclease nbxrn4.bamboo mosaic virus (bamv) has a 6.4-kb (+) sense rna genome with a 5' cap and a 3' poly(a) tail. orf1 of this potexvirus encodes a 155-kda replication protein responsible for the viral rna replication/transcription and 5' cap formation. to learn more about the replication complex of bamv, a protein preparation enriched in the 155-kda replication protein was obtained from nicotiana benthamiana by a protocol involving agroinfiltration and immunoprecipitation. subsequent analysis by sds-page and m ...201626779163
structural properties of potexvirus coat proteins detected by optical methods.it has been shown by x-ray analysis that cores of coat proteins (cps) from three potexviruses, flexible helical rna-containing plant viruses, have similar α-helical structure. however, this similarity cannot explain structural lability of potexvirus virions, which is believed to determine their biological activity. here, we used circular dichroism (cd) spectroscopy in the far uv region to compare optical properties of cps from three potexviruses with the same morphology and similar structure. cp ...201628259129
host factors in the infection cycle of bamboo mosaic virus.to complete the infection cycle efficiently, the virus must hijack the host systems in order to benefit for all the steps and has to face all the defense mechanisms from the host. this review involves a discussion of how these positive and negative factors regulate the viral rna accumulation identified for the bamboo mosaic virus (bamv), a single-stranded rna virus. the genome of bamv is approximately 6.4 kb in length, encoding five functional polypeptides. to reveal the host factors involved in ...201728360904
abscisic acid induces resistance against bamboo mosaic virus through argonaute 2 and 3.plant resistance to pathogens is tuned by defense-related hormones. of which, aba is well documented to moderate resistance against fungi and bacteria. however, aba's contribution to resistance against viruses is pleiotropic. aba affects callose deposition at plasmodesmata (therefore hindering the viral cell-to-cell movement), but here we show that when callose synthase is downregulated, aba still induces resistance against bamboo mosaic virus (bamv) infection. by examining the potential connect ...201728270624
a thioredoxin nbtrxh2 from nicotiana benthamiana negatively regulates the movement of bamboo mosaic virus.an up-regulated gene derived from bamboo mosaic virus (bamv)-infected nicotiana benthamiana plants was cloned and characterized in this study. bamv is a single-stranded, positive-sense rna virus. this gene product, designated as nbtrxh2, was matched with sequences of thioredoxin h proteins, a group of small proteins with a conserved active-site motif wcxpc conferring disulfide reductase activity. to examine how nbtrxh2 is involved in the infection cycle of bamv, we used the virus-induced gene si ...201728052479
identification and characterization of bamboo mosaic virus isolates from a naturally occurring coinfection in bambusa xiashanensis.bamboo mosaic virus (bamv) is a well-characterized virus and a model of virus-host interaction in plants. here, we identified naturally occurring bamv isolates from fujian province, china and furthermore describe a naturally occurring bamv coinfection in bamboo (bambusa xiashanensis) plants. two different types of bamv were identified, represented by isolates bamv-xsnzha7 (x7) and bamv-xsnzha10 (x10). the phylogenetic relationships between x7- and x10-like isolates and published bamv isolates we ...201728050737
the cysteine residues at the c-terminal tail of bamboo mosaic virus triple gene block protein 2 are critical for efficient plasmodesmata localization of protein 1 in the same block.the movement of some plant viruses are accomplished by three proteins encoded by a triple gene block (tgb). the second protein (tgbp2) in the block is a transmembrane protein. this study was aimed to unravel the mechanism underlying the relatively inefficient cell-to-cell movement of bamboo mosaic virus (bamv) caused by amino acid substitutions for the three cys residues, cys-109, cys-112 and cys-119, at the c-terminal tail of tgbp2. results from confocal microscopy revealed that substitutions o ...201727863274
characterisation of sirnas derived from new isolates of bamboo mosaic virus and their associated satellites in infected ma bamboo (dendrocalamus latiflorus).we characterised the virus-derived small interfering rnas (vsirna) of bamboo mosaic virus (ba-vsirnas) and its associated satellite rna (satrna)-derived sirnas (satsirnas) in a bamboo plant (dendrocalamus latiflorus) by deep sequencing. ba-vsirnas and satsirnas of 21-22 nt in length, with both (+) and (-) polarity, predominated. the 5'-terminal base of ba-vsirna was biased towards a, whereas a bias towards c/u was observed in sense satsirnas, and towards a in antisense satsirnas. a large set of ...201727743256
function and structural organization of the replication protein of bamboo mosaic virus.the genus potexvirus is one of the eight genera belonging to the family alphaflexiviridae according to the virus taxonomy 2015 released by international committee on taxonomy of viruses (www.ictvonline.org/index.asp). currently, the genus contains 35 known species including many agricultural important viruses, e.g., potato virus x (pvx). members of this genus are characterized by flexuous, filamentous virions of 13 nm in diameter and 470-580 nm in length. a potexvirus has a monopartite positive- ...201728400766
a versatile papaya mosaic virus (papmv) vaccine platform based on sortase-mediated antigen coupling.flexuous rod-shaped nanoparticles made of the coat protein (cp) of papaya mosaic virus (papmv) have been shown to trigger innate immunity through engagement of toll-like receptor 7 (tlr7). papmv nanoparticles can also serve as a vaccine platform as they can increase the immune response to fused peptide antigens. although this approach shows great potential, fusion of antigens directly to the cp open reading frame (orf) is challenging because the fused peptides can alter the structure of the cp a ...201728720097
phylogeography and coevolution of bamboo mosaic virus and its associated satellite rna.bamboo mosaic virus (bamv), a plant potexvirus, has been found only in infected bamboo species. it is frequently associated with a large, linear single-stranded satellite rna (satbamv) that encodes a non-structural protein. decades of collecting across a wide geographic area in asia have accumulated a sizable number of bamv and satbamv isolates. in this study, we reconstructed the bamv phylogeny and satbamv phylogeny with partial coat protein gene sequences and partial genomic sequences, respect ...201728588562
transmission of bamboo mosaic virus in bamboos mediated by insects in the order diptera.bamboo mosaic virus (bamv), a member of the genus potexvirus, is the major threat to bamboo cultivation. similar to most potexviruses, the transmission of bamv by insect vectors has not been documented previously. however, field observations of bamv disease incidences suggested that insect vectors might be involved. in this study, we aimed to investigate the possibility of insect-mediated transmission of bamv among bamboo clumps, in order to provide further insights into the infection cycles of ...201728559888
interfering satellite rnas of bamboo mosaic virus.satellite rnas (satrnas) are sub-viral agents that may interact with their cognate helper virus (hv) and host plant synergistically and/or antagonistically. satrnas totally depend on the hv for replication, so satrnas and hv usually evolve similar secondary or tertiary rna structures that are recognized by a replication complex, although satrnas and hv do not share an appreciable sequence homology. the satrnas of bamboo mosaic virus (satbamv), the only satrnas of the genus potexvirus, have becom ...201728522996
production of japanese encephalitis virus antigens in plants using bamboo mosaic virus-based vector.japanese encephalitis virus (jev) is among the major threats to public health in asia. for disease control and prevention, the efficient production of safe and effective vaccines against jev is in urgent need. in this study, we produced a plant-made jev vaccine candidate using a chimeric virus particle (cvp) strategy based on bamboo mosaic virus (bamv) for epitope presentation. the chimeric virus, designated bj2a, was constructed by fusing jev envelope protein domain iii (ediii) at the n-terminu ...201728515719
host factors involved in the intracellular movement of bamboo mosaic virus.viruses move intracellularly to their replication compartments, and the newly synthesized viral complexes are transported to neighboring cells through hijacking of the host endomembrane systems. during these processes, numerous interactions occur among viral proteins, host proteins, and the cytoskeleton system. this review mainly focuses on the plant endomembrane network, which may be utilized by bamboo mosaic virus (bamv) to move to its replication compartment, and summarizes the host factors t ...201728487692
chloroplast hsp70 isoform is required for age-dependent tissue preference of bamboo mosaic virus in mature nicotiana benthamiana leaves.plant viruses may exhibit age-dependent tissue preference in their hosts but the underlying mechanisms are not well understood. in this study, we provide several lines of evidence to reveal the determining role of a protein of the nicotiana benthamiana chloroplast hsp70 (nbcphsp70) family, nbcphsp70-2, involved in the preference of bamboo mosaic virus (bamv) to infect older tissues. nbcphsp70 family proteins were identified in complexes pulled down with bamv replicase as the bait. among the isof ...201728459172
the functional roles of the cis-acting elements in bamboo mosaic virus rna genome.bamboo mosaic virus (bamv), which belongs to the genus potexvirus in the family alphaflexiviridae, has a single-stranded positive-sense rna genome that is approximately 6400 nucleotides (nts) in length. positive-sense rna viruses can use genomic rna as a template for translation and replication after entering a suitable host cell. furthermore, such viral rna is recognized by capsid protein for packaging and by viral movement protein(s) or the movement protein complex for cell-to-cell and systemi ...201728450857
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