Journal articles on the topic 'Nicotiana benthamiana Viruses'
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1
Ilgekbaeva,, G. D., E. Sh Makhashov, G. Tulepova, D. Yessimkhankyzy, and S. T. Sadiev. "EXPRESSION OF THE SURFACE ANTIGEN BRUCELLA ABORTUS OMR16 IN NICOTIANA BENTHAMIANA PLANT." REPORTS 5, no. 333 (October 15, 2020): 81–85. http://dx.doi.org/10.32014/2020.2518-1483.122.
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Brucellosis is one of the most contagious and infectious diseases with high incidence rates of cattle and humans in Kazakhstan. Using modern biotechnology techniques to develop vaccines that are reliable and affordable for farmers is an alternative solution to the problem. Plant viruses are often used as a vector for obtaining the expression of antigens of the pathogen. The grape virus A (BAB) is widely used among plant viruses. Brucella membrane proteins are the main objects of this research for futher development of vaccines or diagnostic texts against brucellosis, Membrane proteins (OMPs) are cell specific surface antigens that are immunogenic. OMPs are ideal candidates for the production of recombinant brucellosis vaccines. The object of the study was the outer membrane protein (Omp16), which plays an important role in the suppression of TNF-α production in macrophages. In this study, molecular cloning and analysis of the expression of the Omp16 gene, which was used to express the recombinant protein in plants, was carried out. We selected brucella from the vaccine strain of Brucella abortus 19, and the plant Nicotiana benthamiana, as the subjects for our research, since they widely used for the production of recombinant proteins, and they both appropriate for molecular genetic research. A viral vector was constructed to express the brucellosis antigen Omp16 in Nicotiana benthamiana plants. Nineteen explants were used for the regeneration of transgenic plants. As a result of this studies, the introduced gene of Omp16 was under the subgenomic promoter control of the ORF4 and was successfully expressed while maintaining the efficiency of expression in transgenic plants. The efficiency of viral vectors was evaluated at the level of transcription during expression of the protein Omp16 with viral proteins. The entire leaf blade was infiltrated; the density of Agrobacteria was 0.7. We were able to obtained transgenic plants Nicotiana benthamiana carrying the gene of capsid protein BAB, and the expression of the membrane antigen Omp16 in the viral vector was achieved by replacing the ORF4 with the Omp16 gene. The development of transgenic plants was carried out using agrobacterial transformation.
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Ruocco, Valentina, and Richard Strasser. "Transient Expression of Glycosylated SARS-CoV-2 Antigens in Nicotiana benthamiana." Plants 11, no. 8 (April 18, 2022): 1093. http://dx.doi.org/10.3390/plants11081093.
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The current COVID-19 pandemic very dramatically shows that the world lacks preparedness for novel viral diseases. In addition to newly emerging viruses, many known pathogenic viruses such as influenza are constantly evolving, leading to frequent outbreaks with severe diseases and deaths. Hence, infectious viruses are a recurrent burden to our daily life, and powerful strategies to stop the spread of human pathogens and disease progression are of utmost importance. Transient plant-based protein expression is a technology that allows fast and highly flexible manufacturing of recombinant viral proteins and, thus, can contribute to infectious disease detection and prevention. This review highlights recent progress in the transient production of viral glycoproteins in N. benthamiana with a focus on SARS-CoV-2-derived viral antigens.
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Bally, Julia, Hyungtaek Jung, Cara Mortimer, Fatima Naim, Joshua G. Philips, Roger Hellens, Aureliano Bombarely, Michael M. Goodin, and Peter M. Waterhouse. "The Rise and Rise of Nicotiana benthamiana: A Plant for All Reasons." Annual Review of Phytopathology 56, no. 1 (August 25, 2018): 405–26. http://dx.doi.org/10.1146/annurev-phyto-080417-050141.
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A decade ago, the value of Nicotiana benthamiana as a tool for plant molecular biologists was beginning to be appreciated. Scientists were using it to study plant-microbe and protein-protein interactions, and it was the species of choice with which to activate plasmid-encoded viruses, screen for gene functions with virus-induced gene silencing (VIGS), and transiently express genes by leaf agroinfiltration. However, little information about the species’ origin, diversity, genetics, and genomics was available, and biologists were asking the question of whether N. benthamiana is a second fiddle or virtuoso. In this review, we look at the increased knowledge about the species and its applications over the past decade. Although N. benthamiana may still be the sidekick to Arabidopsis, it shines ever more brightly with realized and yet-to-be-exploited potential.
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Schneider, William L., and Marilyn J. Roossinck. "Evolutionarily Related Sindbis-Like Plant Viruses Maintain Different Levels of Population Diversity in a Common Host." Journal of Virology 74, no. 7 (April 1, 2000): 3130–34. http://dx.doi.org/10.1128/jvi.74.7.3130-3134.2000.
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ABSTRACT The levels of population diversity of three related Sindbis-like plant viruses, Tobacco mosaic virus (TMV), Cucumber mosaic virus (CMV), and Cowpea chlorotic mottle virus(CCMV), in infections of a common host, Nicotiana benthamiana, established from genetically identical viral RNA were examined. Despite probably having a common evolutionary ancestor, the three viruses maintained different levels of population diversity. CMV had the highest levels of diversity, TMV had an intermediate level of diversity, and CCMV had no measurable level of diversity in N. benthamiana. Interestingly, the levels of diversity were correlated to the relative host range sizes of the three viruses. The levels of diversity also remained relatively constant over the course of serial passage. Closer examination of the CMV and TMV populations revealed biases for particular types of substitutions and regions of the genome that may tolerate fewer mutations.
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Goodin, Michael M., David Zaitlin, Rayapati A. Naidu, and Steven A. Lommel. "Nicotiana benthamiana: Its History and Future as a Model for Plant–Pathogen Interactions." Molecular Plant-Microbe Interactions® 21, no. 8 (August 2008): 1015–26. http://dx.doi.org/10.1094/mpmi-21-8-1015.
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Nicotiana benthamiana is the most widely used experimental host in plant virology, due mainly to the large number of diverse plant viruses that can successfully infect it. Additionally, N. benthamiana is susceptible to a wide variety of other plant-pathogenic agents (such as bacteria, oomycetes, fungi, and so on), making this species a cornerstone of host–pathogen research, particularly in the context of innate immunity and defense signaling. Moreover, because it can be genetically transformed and regenerated with good efficiency and is amenable to facile methods for virus-induced gene silencing or transient protein expression, N. benthamiana is rapidly gaining popularity in plant biology, particularly in studies requiring protein localization, interaction, or plant-based systems for protein expression and purification. Paradoxically, despite being an indispensable research model, little is known about the origins, genetic variation, or ecology of the N. benthamiana accessions currently used by the research community. In addition to addressing these latter topics, the purpose of this review is to provide information regarding sources for tools and reagents that can be used to support research in N. benthamiana. Finally, we propose that N. benthamiana is well situated to become a premier plant cell biology model, particularly for the virology community, who as a group were the first to recognize the potential of this unique Australian native.
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Moreno, Marina, Belén Ojeda, Francisco J. Hernández-Walias, Eugenio Sanz-García, Tomás Canto, and Francisco Tenllado. "Water Deficit Improves Reproductive Fitness in Nicotiana benthamiana Plants Infected by Cucumber mosaic virus." Plants 11, no. 9 (May 4, 2022): 1240. http://dx.doi.org/10.3390/plants11091240.
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Plants are concurrently exposed to biotic and abiotic stresses, including infection by viruses and drought. Combined stresses result in plant responses that are different from those observed for each individual stress. We investigated compensatory effects induced by virus infection on the fitness of hosts grown under water deficit, and the hypothesis that water deficit improves tolerance, estimated as reproductive fitness, to virus infection. Our results show that infection by Turnip mosaic virus (TuMV) or Cucumber mosaic virus (CMV) promotes drought tolerance in Arabidopsis thaliana and Nicotiana benthamiana. However, neither CMV nor TuMV had a positive impact on host reproductive fitness following withdrawal of water, as determined by measuring the number of individuals producing seeds, seed grains, and seed germination rates. Importantly, infection by CMV but not by TuMV improved the reproductive fitness of N. benthamiana plants when exposed to drought compared to watered, virus-infected plants. However, no such conditional phenotype was found in Arabidopsis plants infected with CMV. Water deficit did not affect the capacity of infected plants to transmit CMV through seeds. These findings highlight a conditional improvement in biological efficacy of N. benthamiana plants infected with CMV under water deficit, and lead to the prediction that plants can exhibit increased tolerance to specific viruses under some of the projected climate change scenarios.
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Boissinot, Sylvaine, Marie Ducousso, Véronique Brault, and Martin Drucker. "Bioluminescence Production by Turnip Yellows Virus Infectious Clones: A New Way to Monitor Plant Virus Infection." International Journal of Molecular Sciences 23, no. 22 (November 8, 2022): 13685. http://dx.doi.org/10.3390/ijms232213685.
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We used the NanoLuc luciferase bioluminescent reporter system to detect turnip yellows virus (TuYV) in infected plants. For this, TuYV was genetically tagged by replacing the C-terminal part of the RT protein with full-length NanoLuc (TuYV-NL) or with the N-terminal domain of split NanoLuc (TuYV-N65-NL). Wild-type and recombinant viruses were agro-infiltrated in Nicotiana benthamiana, Montia perfoliata, and Arabidopsis thaliana. ELISA confirmed systemic infection and similar accumulation of the recombinant viruses in N. benthamiana and M. perfoliata but reduced systemic infection and lower accumulation in A. thaliana. RT-PCR analysis indicated that the recombinant sequences were stable in N. benthamiana and M. perfoliata but not in A. thaliana. Bioluminescence imaging detected TuYV-NL in inoculated and systemically infected leaves. For the detection of split NanoLuc, we constructed transgenic N. benthamiana plants expressing the C-terminal domain of split NanoLuc. Bioluminescence imaging of these plants after agro-infiltration with TuYV-N65-NL allowed the detection of the virus in systemically infected leaves. Taken together, our results show that NanoLuc luciferase can be used to monitor infection with TuYV.
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Zhong, Xueyan, Hesheng Hou, and Wenping Qiu. "Integrity of Nonviral Fragments in Recombinant Tomato bushy stunt virus and Defective Interfering RNA Is Influenced by Silencing and the Type of Inserts." Molecular Plant-Microbe Interactions® 18, no. 8 (August 2005): 800–807. http://dx.doi.org/10.1094/mpmi-18-0800.
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Recombinant plant viruses have the propensity to remove foreign inserts during replication. This process is virusspecific and occurs in a host-dependent manner. In the present study, we investigated the integrity of foreign inserts in recombinant plant viruses using a model system consisting of Tomato bushy stunt virus (TBSV) and its defective interfering RNA (DI). These were tested in Nicotiana benthamiana plants that were either wild type or transgenic for the green fluorescent protein (GFP) gene. GFP-derived inserts were retained in the recombinant TBSV and DI population that were inoculated onto GFPtransgenic N. benthamiana plants in which silencing of the GFP transgene was initiated, but they were removed from the virus and DIs that were maintained on wild-type plants. A foreign insert derived from an endogenous N. benthamiana gene encoding the H subunit of the magnesium chelatase (NbChlH) was deleted, whereas the fragment of an RNA-dependent RNA polymerase gene (NbRdRP1m) was retained in the recombinant TBSV population. These results demonstrate that the recombination of TBSV to remove nonviral fragments is influenced by silencing and the type of inserts.
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Siddiqui, Shahid Aslam, Cecilia Sarmiento, Mariliis Kiisma, Satu Koivumäki, Anne Lemmetty, Erkki Truve, and Kirsi Lehto. "Effects of viral silencing suppressors on tobacco ringspot virus infection in two Nicotiana species." Journal of General Virology 89, no. 6 (June 1, 2008): 1502–8. http://dx.doi.org/10.1099/vir.0.83621-0.
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This study investigated the effects of silencing suppressors derived from six different viruses (P1, P19, P25, HcPro, AC2 and 2b), expressed in transgenic Nicotiana tabacum and Nicotiana benthamiana plants, on the infection pattern of tobacco ringspot virus (TRSV) potato calico strain. In N. benthamiana, this virus produced an initial infection with severe systemic symptoms, but the infection was strongly reduced within a few weeks as the plant recovered from the infection. P25 and HcPro silencing suppressors effectively prevented recovery in this host, allowing continuous accumulation of the viral RNA as well as of the virus-specific small interfering RNAs, in the systemically infected leaves. In the P1-, P19-, AC2- or 2b-expressing transgenic N. benthamiana, the recovery was not complete. Susceptibility of N. tabacum to this virus was temperature sensitive. At lower temperatures, up to 25 °C, the plants became systemically infected, but at higher temperatures, the infections were limited to the inoculated leaves. In these preventative conditions, all silencing suppressor transgenes (except P25, which was expressed at very low levels) allowed the establishment of systemic infections. Very strong and consistent systemic infections were observed in HcPro- and AC2-expressing plants.
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Canto, Tomas, Stuart A. MacFarlane, and Peter Palukaitis. "ORF6 of Tobacco mosaic virus is a determinant of viral pathogenicity in Nicotiana benthamiana." Journal of General Virology 85, no. 10 (October 1, 2004): 3123–33. http://dx.doi.org/10.1099/vir.0.80270-0.
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Tobacco mosaic virus (TMV) contains a sixth open reading frame (ORF6) that potentially encodes a 4·8 kDa protein. Elimination of ORF6 from TMV attenuated host responses in Nicotiana benthamiana without alteration in virus accumulation. Furthermore, heterologous expression of TMV ORF6 from either potato virus X (PVX) or tobacco rattle virus (TRV) vectors enhanced the virulence of both viruses in N. benthamiana, also without effects on their accumulation. By contrast, the presence or absence of TMV ORF6 had no effect on host response or virus accumulation in N. tabacum plants infected with TMV or PVX. TMV ORF6 also had no effect on the synergism between TMV and PVX in N. tabacum. However, the presence of the TMV ORF6 did have an effect on the pathogenicity of a TRV vector in N. tabacum. In three different types of assay carried out in N. benthamiana plants, expression of TMV ORF6 failed to suppress gene silencing. Expression in N. benthamiana epidermal cells of the encoded 4·8 kDa protein fused to the green fluorescent protein at either end showed, in addition to widespread cytosolic fluorescence, plasmodesmatal targeting specific to both fusion constructs. The role of the ORF6 in host responses is discussed.
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Chung, Bong-Nam, Tomas Canto, and Peter Palukaitis. "Stability of recombinant plant viruses containing genes of unrelated plant viruses." Journal of General Virology 88, no. 4 (April 1, 2007): 1347–55. http://dx.doi.org/10.1099/vir.0.82477-0.
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The stability of hybrid plant viruses that might arise by recombination in transgenic plants was examined using hybrid viruses derived from the viral expression vectors potato virus X (PVX) and tobacco rattle virus (TRV). The potato virus Y (PVY) NIb and HCPro open reading frames (ORFs) were introduced into PVX to generate PVX-NIb and PVX-HCPro, while the PVY NIb ORF was introduced into a vector derived from TRV RNA2 to generate TRV-NIb. All three viruses were unstable and most of the progeny viruses had lost the inserted sequences between 2 and 4 weeks post-inoculation. There was some variation in the rate of loss of part or all of the inserted sequence and the number of plants containing the deleted viruses, depending on the sequence, the host (Nicotiana tabacum vs Nicotiana benthamiana) or the vector, although none of these factors was associated consistently with the preferential loss of the inserted sequences. PVX-NIb was unable to accumulate in NIb-transgenic tobacco resistant to infection by PVY and also showed loss of the NIb insert from PVX-NIb in some NIb-transgenic tobacco plants susceptible to infection by PVY. These data indicate that such hybrid viruses, formed in resistant transgenic plants from a transgene and an unrelated virus, would be at a selective disadvantage, first by being targeted by the resistance mechanism and second by not being competitive with the parental virus.
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Vigne, Emmanuelle, John Gottula, Corinne Schmitt-Keichinger, Véronique Komar, Léa Ackerer, Lorène Belval, Lalaina Rakotomalala, Olivier Lemaire, Christophe Ritzenthaler, and Marc Fuchs. "A strain-specific segment of the RNA-dependent RNA polymerase of grapevine fanleaf virus determines symptoms in Nicotiana species." Journal of General Virology 94, no. 12 (December 1, 2013): 2803–13. http://dx.doi.org/10.1099/vir.0.057646-0.
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Factors involved in symptom expression of viruses from the genus Nepovirus in the family Secoviridae such as grapevine fanleaf virus (GFLV) are poorly characterized. To identify symptom determinants encoded by GFLV, infectious cDNA clones of RNA1 and RNA2 of strain GHu were developed and used alongside existing infectious cDNA clones of strain F13 in a reverse genetics approach. In vitro transcripts of homologous combinations of RNA1 and RNA2 induced systemic infection in Nicotiana benthamiana and Nicotiana clevelandii with identical phenotypes to WT virus strains, i.e. vein clearing and chlorotic spots on N. benthamiana and N. clevelandii for GHu, respectively, and lack of symptoms on both hosts for F13. The use of assorted transcripts mapped symptom determinants on RNA1 of GFLV strain GHu, in particular within the distal 408 nt of the RNA-dependent RNA polymerase (1EPol), as shown by RNA1 transcripts for which coding regions or fragments derived thereof were swapped. Semi-quantitative analyses indicated no significant differences in virus titre between symptomatic and asymptomatic plants infected with various recombinants. Also, unlike the nepovirus tomato ringspot virus, no apparent proteolytic cleavage of GFLV protein 1EPol was detected upon virus infection or transient expression in N. benthamiana. In addition, GFLV protein 1EPol failed to suppress silencing of EGFP in transgenic N. benthamiana expressing EGFP or to enhance GFP expression in patch assays in WT N. benthamiana. Together, our results suggest the existence of strain-specific functional domains, including a symptom determinant module, on the RNA-dependent RNA polymerase of GFLV.
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Dardick, Christopher. "Comparative Expression Profiling of Nicotiana benthamiana Leaves Systemically Infected with Three Fruit Tree Viruses." Molecular Plant-Microbe Interactions® 20, no. 8 (August 2007): 1004–17. http://dx.doi.org/10.1094/mpmi-20-8-1004.
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Plant viruses cause a wide array of disease symptoms and cytopathic effects. Although some of these changes are virus specific, many appear to be common even among diverse viruses. Currently, little is known about the underlying molecular determinants. To identify gene expression changes that are concomitant with virus symptoms, we performed comparative expression profiling experiments on Nicotiana benthamiana leaves infected with one of three different fruit tree viruses that produce distinct symptoms: Plum pox potyvirus (PPV; leaf distortion and mosaic), Tomato ringspot nepovirus (ToRSV; tissue necrosis and general chlorosis), and Prunus necrotic ringspot ilarvirus (PNRSV; subtle chlorotic mottling). The numbers of statistically significant genes identified were consistent with the severity of the observed symptoms: 1,082 (ToRSV), 744 (PPV), and 89 (PNRSV). In all, 56% of the gene expression changes found in PPV-infected leaves also were altered by ToRSV, 87% of which changed in the same direction. Both PPV- and ToRSV-infected leaves showed widespread repression of genes associated with plastid functions. PPV uniquely induced the expression of large numbers of cytosolic ribosomal genes whereas ToRSV repressed the expression of plastidic ribosomal genes. How these and other observed expression changes might be associated with symptom development are discussed.
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Dietrich, Christof, and Edgar Maiss. "Fluorescent labelling reveals spatial separation of potyvirus populations in mixed infected Nicotiana benthamiana plants." Journal of General Virology 84, no. 10 (October 1, 2003): 2871–76. http://dx.doi.org/10.1099/vir.0.19245-0.
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The distribution of potyviruses in mixed infected Nicotiana benthamiana plants was investigated by using green and red fluorescent proteins (GFP, DsRed). Full-length cDNA clones of Plum pox virus (PPV-NAT-AgfpS; PPV-NAT-red), Tobacco vein mottling virus (TVMV-gfp; TVMV-red) and Clover yellow vein virus (ClYVV-GFP) expressing fluorescent proteins, referred to here as labelled viruses, were used to characterize the distribution of different potyviral populations (e.g. TVMV-gfp/PPV-NAT-red), as well as populations of identical, but differently labelled potyviruses (e.g. PPV-NAT-AgfpS/PPV-NAT-red) or in mixed infections of potyviruses with labelled Potato virus X (PVX). Plants infected by any of the PVX/potyvirus combinations exhibited synergistic symptoms and large numbers of cells were doubly infected. In contrast, co-infections of differently labelled potyvirus populations appeared non-synergistic and remained predominantly separate in the infected plants, independent of whether different viruses or identical but differently labelled viruses were co-infecting. Contact of differently labelled virus populations that exhibited spatial separation was restricted to a small number of cells at the border of different fluorescent cell clusters.
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Havelda, Zoltán, Csaba Hornyik, Aniello Crescenzi, and József Burgyán. "In Situ Characterization of Cymbidium Ringspot Tombusvirus Infection-Induced Posttranscriptional Gene Silencing in Nicotiana benthamiana." Journal of Virology 77, no. 10 (May 15, 2003): 6082–86. http://dx.doi.org/10.1128/jvi.77.10.6082-6086.2003.
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ABSTRACT In plants, posttranscriptional gene silencing (PTGS) is an ancient and effective defense mechanism against viral infection. A number of viruses encode proteins that suppress virus-activated PTGS. The p19 protein of tombusviruses is a potent PTGS suppressor which interferes with the onset of PTGS-generated systemic signaling and is not required for viral replication or for viral movement in Nicotiana benthamiana. This unique feature of p19 suppressor allowed us to analyze the mechanism of PTGS-based host defense and its viral suppression without interfering with other viral functions. In contrast to the necrotic symptoms caused by wild-type tombusvirus, the infection of p19-defective mutant virus results in the development of a typical PTGS-associated recovery phenotype in N. benthamiana. In this report we show the effect of PTGS on the viral infection process for N. benthamiana infected with either wild-type Cymbidium Ringspot Tombusvirus (CymRSV) or a p19-defective mutant (Cym19stop). In situ analyses of different virus-derived products revealed that PTGS is not able to reduce accumulation of virus in primary infected cells regardless of the presence of p19 PTGS suppressor. We also showed that both CymRSV and Cym19stop viruses move systemically in the vasculature, with similar efficiencies. However, in contrast to the uniform accumulation of CymRSV throughout systemically infected leaves, the presence of Cym19stop virus was confined to and around the vascular bundles. These results suggest that the role of p19 is to prevent the onset of mobile signal-induced systemic PTGS ahead of the viral infection front, leading to generalized infection.
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Andika, Ida Bagus, Liying Sun, Rong Xiang, Junmin Li, and Jianping Chen. "Root-Specific Role for Nicotiana benthamiana RDR6 in the Inhibition of Chinese wheat mosaic virus Accumulation at Higher Temperatures." Molecular Plant-Microbe Interactions® 26, no. 10 (October 2013): 1165–75. http://dx.doi.org/10.1094/mpmi-05-13-0137-r.
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Some viruses only infect plants at cool temperatures but the molecular mechanism underlying this low-temperature dependence remains unclear. Chinese wheat mosaic virus (CWMV, genus Furovirus) was able to infect wheat and Nicotiana benthamiana plants at 16 but not at 24°C. When CWMV-infected plants were transferred to 24°C for 2 weeks, the newly emerged leaves and roots became virus free. Co-infection with Potato virus Y rescued CWMV accumulation in N. benthamiana plants after a temperature shift to 24°C. In transgenic N. benthamiana plants silenced for the N. benthamiana RNA-dependent RNA polymerase 6 (NbRDR6), CWMV was able to accumulate in roots but not in leaves after a temperature shift to 24°C. Deep sequencing of small RNAs showed that, at 16°C, abundant CWMV small interfering (si)RNAs accumulated in infected N. benthamiana plants. Silencing of NbRDR6 increased the abundance of CWMV siRNAs and the generation of siRNAs from hotspots in the CWMV genome. In contrast, when shifted to 24°C for 1 week, CWMV siRNAs were markedly fewer in roots of NbRDR6-silenced than in roots of wild-type plants but were similar in the leaves of those plants. Our results demonstrate the root-specific role of NbRDR6 in the inhibition of CWMV accumulation and biogenesis of CWMV siRNAs at higher temperatures.
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Rajamäki, Minna-Liisa, Dehui Xi, Sidona Sikorskaite-Gudziuniene, Jari P. T. Valkonen, and Steven A. Whitham. "Differential Requirement of the Ribosomal Protein S6 and Ribosomal Protein S6 Kinase for Plant-Virus Accumulation and Interaction of S6 Kinase with Potyviral VPg." Molecular Plant-Microbe Interactions® 30, no. 5 (May 2017): 374–84. http://dx.doi.org/10.1094/mpmi-06-16-0122-r.
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Ribosomal protein S6 (RPS6) is an indispensable plant protein regulated, in part, by ribosomal protein S6 kinase (S6K) which, in turn, is a key regulator of plant responses to stresses and developmental cues. Increased expression of RPS6 was detected in Nicotiana benthamiana during infection by diverse plant viruses. Silencing of the RPS6 and S6K genes in N. benthamiana affected accumulation of Cucumber mosaic virus, Turnip mosaic virus (TuMV), and Potato virus A (PVA) in contrast to Turnip crinkle virus and Tobacco mosaic virus. In addition, the viral genome-linked protein (VPg) of TuMV and PVA interacted with S6K in plant cells, as detected by bimolecular fluorescence complementation assay. The VPg–S6K interaction was detected in cytoplasm, nucleus, and nucleolus, whereas the green fluorescent protein-tagged S6K alone showed cytoplasmic localization only. These results demonstrate that the requirement for RPS6 and S6K differs for diverse plant viruses with different translation initiation strategies and suggest that potyviral VPg–S6K interaction may affect S6K functions in both the cytoplasm and the nucleus.
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Li, Yi, Yi M. Bao, Chun H. Wei, Zhen S. Kang, Yong W. Zhong, Peng Mao, Gang Wu, Zhang L. Chen, Joachim Schiemann, and Richard S. Nelson. "Rice Dwarf Phytoreovirus Segment S6-Encoded Nonstructural Protein Has a Cell-to-Cell Movement Function." Journal of Virology 78, no. 10 (May 15, 2004): 5382–89. http://dx.doi.org/10.1128/jvi.78.10.5382-5389.2004.
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ABSTRACT Rice dwarf virus (RDV) is a member of the genus Phytoreovirus, which is composed of viruses with segmented double-stranded RNA genomes. Proteins that support the intercellular movement of these viruses in the host have not been identified. Microprojectile bombardment was used to determine which open reading frames (ORFs) support intercellular movement of a heterologous virus. A plasmid containing an infectious clone of Potato virus X (PVX) defective in cell-to-cell movement and expressing either β-glucuronidase or green fluorescent protein (GFP) was used for cobombardment with plasmids containing ORFs from RDV gene segments S1 through S12 onto leaves of Nicotiana benthamiana. Cell-to-cell movement of the movement-defective PVX was restored by cobombardment with a plasmid containing S6. In the absence of S6, no other gene segment supported movement. Identical results were obtained with Nicotiana tabacum, a host that allows fewer viruses to infect and spread within its tissue. S6 supported the cell-to-cell movement of the movement-defective PVX in sink and source leaves of N. benthamiana. A mutant S6 lacking the translation start codon did not complement the cell-to-cell movement of the movement-defective PVX. An S6 protein product (Pns6)-enhanced GFP fusion was observed near or within cell walls of epidermal cells from N. tabacum. By immunocytochemistry, unfused Pns6 was localized to plasmodesmata in rice leaves infected with RDV. S6 thus encodes a protein with characteristics identical to those of other viral proteins required for the cell-to-cell movement of their genome and therefore is likely required for the cell-to-cell movement of RDV.
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Qu, Feng, Xiaohong Ye, Guichuan Hou, Shirley Sato, Thomas E. Clemente, and T. Jack Morris. "RDR6 Has a Broad-Spectrum but Temperature-Dependent Antiviral Defense Role in Nicotiana benthamiana." Journal of Virology 79, no. 24 (December 15, 2005): 15209–17. http://dx.doi.org/10.1128/jvi.79.24.15209-15217.2005.
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ABSTRACT SDE1/SGS2/RDR6, a putative RNA-dependent RNA polymerase (RdRP) from Arabidopsis thaliana, has previously been found to be indispensable for maintaining the posttranscriptional silencing of transgenes, but it is seemingly redundant for antiviral defense. To elucidate the antiviral role of this RdRP in a different host plant and to evaluate whether plant growth conditions affect its role, we down-regulated expression of the Nicotiana benthamiana homolog, NbRDR6, and examined the plants for altered susceptibility to various viruses at different growth temperatures. The results we describe here clearly show that plants with reduced expression of NbRDR6 were more susceptible to all viruses tested and that this effect was more pronounced at higher growth temperatures. Diminished expression of NbRDR6 also permitted efficient multiplication of tobacco mosaic virus in the shoot apices, leading to serious disruption with microRNA-mediated developmental regulation. Based on these results, we propose that NbRDR6 participates in the antiviral RNA silencing pathway that is stimulated by rising temperatures but suppressed by virus-encoded silencing suppressors. The relative strengths of these two factors, along with other plant defense components, critically influence the outcome of virus infections.
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Liu, Junying, Huiyan Fan, Ying Wang, Chenggui Han, Xianbing Wang, Jialin Yu, Dawei Li, and Yongliang Zhang. "Genome-Wide microRNA Profiling Using Oligonucleotide Microarray Reveals Regulatory Networks of microRNAs in Nicotiana benthamiana During Beet Necrotic Yellow Vein Virus Infection." Viruses 12, no. 3 (March 12, 2020): 310. http://dx.doi.org/10.3390/v12030310.
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Beet necrotic yellow vein virus (BNYVV) infections induce stunting and leaf curling, as well as root and floral developmental defects and leaf senescence in Nicotiana benthamiana. A microarray analysis with probes capable of detecting 1596 candidate microRNAs (miRNAs) was conducted to investigate differentially expressed miRNAs and their targets upon BNYVV infection of N. benthamiana plants. Eight species-specific miRNAs of N. benthamiana were identified. Comprehensive characterization of the N. benthamiana microRNA profile in response to the BNYVV infection revealed that 129 miRNAs were altered, including four species-specific miRNAs. The targets of the differentially expressed miRNAs were predicted accordingly. The expressions of miR164, 160, and 393 were up-regulated by BNYVV infection, and those of their target genes, NAC21/22, ARF17/18, and TIR, were down-regulated. GRF1, which is a target of miR396, was also down-regulated. Further genetic analysis of GRF1, by Tobacco rattle virus-induced gene silencing, assay confirmed the involvement of GRF1 in the symptom development during BNYVV infection. BNYVV infection also induced the up-regulation of miR168 and miR398. The miR398 was predicted to target umecyanin, and silencing of umecyanin could enhance plant resistance against viruses, suggesting the activation of primary defense response to BNYVV infection in N. benthamiana. These results provide a global profile of miRNA changes induced by BNYVV infection and enhance our understanding of the mechanisms underlying BNYVV pathogenesis.
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Ge, Linmei, Jiangtao Zhang, Xueping Zhou, and Hongye Li. "Genetic Structure and Population Variability of Tomato Yellow Leaf Curl China Virus." Journal of Virology 81, no. 11 (March 21, 2007): 5902–7. http://dx.doi.org/10.1128/jvi.02431-06.
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ABSTRACT Geminiviruses have circular single-stranded DNA genomes and are important pathogens in tropical and subtropical regions, but their population diversity and variability are poorly understood. Here, we have investigated variations accumulating in Tomato yellow leaf curl China virus (TYLCCNV), a geminivirus in the genus Begomovirus of the family Geminiviridae. The population variation was analyzed in a naturally infected tomato (Solanum lycopersicom) plant and in Nicotiana benthamiana and tomato plants experimentally infected with a swarm of TYLCCNV DNA clones to provide an identical sequence for initiation of infection. Our results demonstrate that the population of TYLCCNV in a naturally infected tomato plant was genetically heterogeneous and that rapid mutation occurred in the populations amplified from N. benthamiana and tomato plants that had been infected with cloned DNA. This feature of the population of TYLCCNV in these plants consisted of the consensus sequence and a pool of mutants that are not identical but are closely related to the consensus sequence, and it coincides with the quasispecies concept described for many RNA viruses. The mutation frequency was circa 10−4 in N. benthamiana and tomato at 60 days postinoculation, a value comparable to that reported for plant RNA viruses. The quasispecies-like nature of the TYLCCNV populations suggested that TYLCCNV is capable of rapid evolution and adaptation in response to changing agricultural practices.
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Chiu, Yi-Shu, Pi-Yu Chen, Tung Kuan, Po-Chuan Wang, Ying-Ju Chen, Yu-Liang Yang, and Hsin-Hung Yeh. "A Polysaccharide Derived from a Trichosporon sp. Culture Strongly Primes Plant Resistance to Viruses." Molecular Plant-Microbe Interactions® 31, no. 12 (December 2018): 1257–70. http://dx.doi.org/10.1094/mpmi-01-18-0012-r.
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Plant viruses cause devastating diseases in plants, yet no effective viricide is available for agricultural application. We screened cultured filtrates derived from various soil microorganisms cultured in vegetable broth that enhanced plant viral resistance. A cultured filtrate, designated F8 culture filtrate, derived from a fungus belonging to the genus Trichosporon, induced strong resistance to various viruses on different plants. Our inoculation assay found the infection rate of Tobacco mosaic virus (TMV)-inoculated Nicotiana benthamiana with F8 culture filtrate pretreatment may decrease to 0%, whereas salicylic acid (SA)-pretreated N. benthamiana attenuated TMV-caused symptoms but remained 100% infected. Tracking Tobacco mosaic virus tagged with green fluorescence protein in plants revealed pretreatment with F8 culture filtrate affected the initial establishment of the virus infection. From F8 culture filtrate, we identified a previously unknown polysaccharide composed of D-mannose, D-galactose, and D-glucose in the ratio 1.0:1.2:10.0 with a α-D-1,4-glucan linkage to be responsible for the induction of plant resistance against viruses through priming of SA-governed immune-responsive genes. Notably, F8 culture filtrate only triggered local defense but was much more effective than conventional SA-mediated systematic acquired resistance. Our finding revealed that microbial cultured metabolites provided a rich source for identification of potent elicitors in plant defense.
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Qian, Xin, Qing Xiang, Tongqing Yang, Hongyu Ma, Xin Ding, and Xiaorong Tao. "Molecular Co-Chaperone SGT1 Is Critical for Cell-to-Cell Movement and Systemic Infection of Tomato Spotted Wilt Virus in Nicotiana benthamiana." Viruses 10, no. 11 (November 17, 2018): 647. http://dx.doi.org/10.3390/v10110647.
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Tospovirus is a tripartite negative stranded RNA virus and is considered as one of the most devastating plant viruses. Successful virus infection in plant requires many host factors. To date, very few host factors have been identified as important in Tospovirus infection in plants. We reported earlier that NSm protein encoded by Tomato spotted wilt virus (TSWV), a type species of the genus Orthotospovirus, plays critical roles in viral cell-to-cell and long-distance movement. In this study, we determined that molecular co-chaperone NbSGT1 interacted with TSWV NSm in Nicotiana benthamiana. TSWV infection significantly upregulated the expression of NbSGT1 gene and transient overexpression of NbSGT1 in N. benthamiana leaves accelerated TSWV infection. In contrast, silencing the NbSGT1 gene expression using a virus-induced gene silencing (VIGS) approach strongly inhibited TSWV NSm cell-to-cell movement, as well as TSWV local and systemic infection in N. benthamiana plants. Furthermore, NbSGT1 was found to regulate the infection of both American and Euro/Asia type tospoviruses in N. benthamiana plant. Collectively, our findings presented in this paper and the results published previously indicated that molecular co-chaperone NbSGT1 plays important roles in modulating both positive stranded and tripartite negative stranded RNA virus infection in plants.
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Moura, Monika Fecury, Marcelo Soman, Tatiana Mituti, Marcelo Agenor Pavan, and Renate Krause-Sakate. "Solanum americanum: reservoir for Potato virus Y and Cucumber mosaic virus in sweet pepper crops." Summa Phytopathologica 40, no. 1 (March 2014): 78–80. http://dx.doi.org/10.1590/s0100-54052014000100012.
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Weeds can act as important reservoirs for viruses. Solanum americanum (Black nightshade) is a common weed in Brazil and samples showing mosaic were collected from sweet pepper crops to verify the presence of viruses. One sample showed mixed infection between Cucumber mosaic virus (CMV) and Potato virus Y (PVY) and one sample showed simple infection by PVY. Both virus species were transmitted by plant extract and caused mosaic in tomato (Solanum lycopersicum cv. Santa Clara), sweet pepper (Capsicum annuum cv. Magda), Nicotiana benthamiana and N. tabaccum TNN, and local lesions on Chenopodium quinoa, C. murale and C. amaranticolor. The coat protein sequences for CMV and PVY found in S. americanum are phylogenetically more related to isolates from tomato. We conclude that S. americanum can act as a reservoir for different viruses during and between sweet pepper crop seasons.
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Huang, Ying Wen, Chung Chi Hu, Ching Hsiu Tsai, Na Sheng Lin, and Yau Heiu Hsu. "Chloroplast Hsp70 Isoform Is Required for Age-Dependent Tissue Preference of Bamboo mosaic virus in Mature Nicotiana benthamiana Leaves." Molecular Plant-Microbe Interactions® 30, no. 8 (August 2017): 631–45. http://dx.doi.org/10.1094/mpmi-01-17-0012-r.
Full textAbstract:
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 isoforms of NbcpHsp70, only the specific silencing of NbcpHsp70-2 resulted in the significant decrease of BaMV RNA in N. benthamiana protopalsts, indicating that NbcpHsp70-2 is involved in the efficient replication of BaMV RNA. We further identified the age-dependent import regulation signal contained in the transit peptide of NbcpHsp70-2. Deletion, overexpression, and substitution experiments revealed that the signal in the transit peptide of NbcpHsp70-2 is crucial for both the import of NbcpHsp70-2 into older chloroplasts and the preference of BaMV for infecting older leaves of N. benthamiana. Together, these data demonstrated that BaMV may exploit a cellular age-dependent transportation mechanism to target a suitable environment for viral replication.
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Wylie, Steve, and Hua Li. "Historical and Scientific Evidence for the Origin and Cultural Importance to Australia’s First-Nations Peoples of the Laboratory Accession of Nicotiana benthamiana, a Model for Plant Virology." Viruses 14, no. 4 (April 8, 2022): 771. http://dx.doi.org/10.3390/v14040771.
Full textAbstract:
Nicotiana benthamiana is an indigenous plant species distributed across northern Australia. The laboratory accession (LAB) of N. benthamiana has become widely adopted as a model host for plant viruses, and it is distinct from other accessions morphologically, physiologically, and by having an attenuation-of-function mutation in the RNA-dependent RNA polymerase 1 (NbRdr1) gene, referred to as NbRdr1m. Recent historical evidence suggested LAB was derived from a 1936 collection by John Cleland at The Granites of the Northern Territory, although no scientific evidence was provided. We provide scientific evidence and further historical evidence supporting the origin of LAB as The Granites. Analysis of a herbarium specimen of N. benthamiana collected by Cleland in 1936 revealed that The Granites population contains plants heterozygous for the NbRdr1 locus, having both the functional NbRdr1 and the mutant NbRdr1m alleles. N. benthamiana was an important cultural asset actively utilised as the narcotic Pituri (chewing tobacco) by the Warlpiri Aboriginal people at the site, who prevented women of child-bearing age from consuming it. We propose that Aboriginal people selected some of the unique traits of LAB that have subsequently facilitated its adoption as a model plant, such as lack of seed dormancy, fast maturity, low nornicotine content, and gracility.
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Senshu, Hiroko, Johji Ozeki, Ken Komatsu, Masayoshi Hashimoto, Kouji Hatada, Michiko Aoyama, Satoshi Kagiwada, Yasuyuki Yamaji, and Shigetou Namba. "Variability in the level of RNA silencing suppression caused by triple gene block protein 1 (TGBp1) from various potexviruses during infection." Journal of General Virology 90, no. 4 (April 1, 2009): 1014–24. http://dx.doi.org/10.1099/vir.0.008243-0.
Full textAbstract:
RNA silencing is an important defence mechanism against virus infection, and many plant viruses encode RNA silencing suppressors as a counter defence. In this study, we analysed the RNA silencing suppression ability of multiple virus species of the genus Potexvirus. Nicotiana benthamiana plants exhibiting RNA silencing of a green fluorescent protein (GFP) transgene showed reversal of GFP fluorescence when systemically infected with potexviruses. However, the degree of GFP fluorescence varied among potexviruses. Agrobacterium-mediated transient expression assay in N. benthamiana leaves demonstrated that the triple gene block protein 1 (TGBp1) encoded by these potexviruses has drastically different levels of silencing suppressor activity, and these differences were directly related to variations in the silencing suppression ability during virus infection. These results suggest that suppressor activities differ even among homologous proteins encoded by viruses of the same genus, and that TGBp1 contributes to the variation in the level of RNA silencing suppression by potexviruses. Moreover, we investigated the effect of TGBp1 encoded by Plantago asiatica mosaic virus (PlAMV), which exhibited a strong suppressor activity, on the accumulation of microRNA, virus genomic RNA and virus-derived small interfering RNAs.
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Haikonen, Tuuli, Minna-Liisa Rajamäki, and Jari P. T. Valkonen. "Interaction of the Microtubule-Associated Host Protein HIP2 with Viral Helper Component Proteinase Is Important in Infection with Potato virus A." Molecular Plant-Microbe Interactions® 26, no. 7 (July 2013): 734–44. http://dx.doi.org/10.1094/mpmi-01-13-0023-r.
Full textAbstract:
Microtubules (MT) outline and maintain the overall shape of cells and can reorganize cellular membranes to serve as sites of RNA virus replication. Here, we provide data on involvement of an MT-associated protein in infection of plants with a potyvirus, Potato virus A (PVA), representing the largest family of plant-infecting RNA viruses. Our results showed that helper-component proteinase (HCpro)-interacting protein 2 (HIP2) of potato (Solanum tuberosum) is an MT-associated protein similar to Arabidopsis SPR2. Virus-induced silencing of HIP2 in Nicotiana benthamiana resulted in a spiral-like growth phenotype, similar to the Arabidopsis spr2 mutant, and the spr2 phenotype in Arabidopsis was complemented with potato HIP2. HCpro of PVA interacted with HIP2 of potato and tobacco (Nicotiana tabacum). The interaction was detected by bimolecular fluorescence complementation in PVA-infected leaves on MT and MT intersections at the cell cortex. HIP2-HCpro interaction was determined by the C-proximal α-helix-rich domain of HIP2, whereas the N-proximal putative TOG domain and the central coiled-coil domain of HIP2 controlled HIP2 dimerization and binding to MT. Accumulation of PVA was significantly reduced in the HIP2-silenced leaves of N. benthamiana, which indicates that HIP2-HCpro interactions are important for virus infection.
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Pohl, Diana, and Christina Wege. "Synergistic pathogenicity of a phloem-limited begomovirus and tobamoviruses, despite negative interference." Journal of General Virology 88, no. 3 (March 1, 2007): 1034–40. http://dx.doi.org/10.1099/vir.0.82653-0.
Full textAbstract:
In contrast to previous observations on phloem-limited geminiviruses supported in movement andaccumulation by RNA viruses such as cucumo- and tobamoviruses, tissue infiltration by Abutilon mosaic virus (AbMV) was enhanced by neither Tobacco mosaic virus nor Tomato mosaic virus (ToMV) in two different hosts, Nicotiana benthamiana and tomato. Both tobamoviruses exerted a negative effect on the DNA virus, resulting in a decrease in AbMV accumulation and significantly reduced infectivity in N. benthamiana. Despite these unexpected molecular observations, a striking synergistic enhancement in pathogenicity occurred with respectto stunting and necrosis. In situ hybridization revealed that this was not due to any alteration of tissue infiltration by AbMV, which also remained limited to the phloem in the mixed infections. Transgenically expressed ToMV 30K movement protein was not able to induce phloemescape of AbMV in tomato plants and did not lead to any obvious change in begomovirus symptomatology.
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Brault, V., S. Pfeffer, M. Erdinger, J. Mutterer, and V. Ziegler-Graff. "Virus-Induced Gene Silencing in Transgenic Plants Expressing the Minor Capsid Protein of Beet western yellows virus." Molecular Plant-Microbe Interactions® 15, no. 8 (August 2002): 799–807. http://dx.doi.org/10.1094/mpmi.2002.15.8.799.
Full textAbstract:
Transgenic Nicotiana benthamiana expressing the minor coat protein P74 of the phloem-limited Beet western yellows virus (BWYV) exhibited an unusual spatial pattern of post-transcriptional gene silencing (PTGS) when infected with BWYV or related viruses. Following infection, transgenic P74 and its mRNA accumulated to only low levels, 21 to 23 nucleotide RNAs homologous to the transgene appeared, and the transgene DNA underwent methylation. The infecting viral RNA, however, was not subject to significant silencing but multiplied readily and produced P74 in the phloem tissues, although the P74 encoded by the transgene disappeared from the phloem as well as the nonvascular tissues.
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Luna, Ana P., Gabriel Morilla, Olivier Voinnet, and Eduardo R. Bejarano. "Functional Analysis of Gene-Silencing Suppressors from Tomato Yellow Leaf Curl Disease Viruses." Molecular Plant-Microbe Interactions® 25, no. 10 (October 2012): 1294–306. http://dx.doi.org/10.1094/mpmi-04-12-0094-r.
Full textAbstract:
Tomato yellow leaf curl disease (TYLCD) is caused by a complex of phylogenetically related Begomovirus spp. that produce similar symptoms when they infect tomato plants but have different host ranges. In this work, we have evaluated the gene-silencing-suppression activity of C2, C4, and V2 viral proteins isolated from the four main TYLCD-causing strains in Spain in Nicotiana benthamiana. We observed varying degrees of local silencing suppression for each viral protein tested, with V2 proteins from all four viruses exhibiting the strongest suppression activity. None of the suppressors were able to avoid the spread of the systemic silencing, although most produced a delay. In order to test the silencing-suppression activity of Tomato yellow leaf curl virus (TYLCV) and Tomato yellow leaf curl Sardinia virus (TYLCSV) proteins in a shared (tomato) and nonshared (bean) host, we established novel patch assays. Using these tools, we found that viral proteins from TYLCV were able to suppress silencing in both hosts, whereas TYLCSV proteins were only effective in tomato. This is the first time that viral suppressors from a complex of disease-causing geminiviruses have been subject to a comprehensive analysis using two economically important crop hosts, as well as the established N. benthamiana plant model.
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Ribeiro, Simone G., Hendrikus Lohuis, Rob Goldbach, and Marcel Prins. "Tomato Chlorotic Mottle Virus Is a Target of RNA Silencing but the Presence of Specific Short Interfering RNAs Does Not Guarantee Resistance in Transgenic Plants." Journal of Virology 81, no. 4 (November 29, 2006): 1563–73. http://dx.doi.org/10.1128/jvi.01238-06.
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ABSTRACT Tomato chlorotic mottle virus (ToCMoV) is a begomovirus found widespread in tomato fields in Brazil. ToCMoV isolate BA-Se1 (ToCMoV-[BA-Se1]) was shown to trigger the plant RNA silencing surveillance in different host plants and, coinciding with a decrease in viral DNA levels, small interfering RNAs (siRNAs) specific to ToCMoV-[BA-Se1] accumulated in infected plants. Although not homogeneously distributed, the siRNA population in both infected Nicotiana benthamiana and tomato plants represented the entire DNA-A and DNA-B genomes. We determined that in N. benthamiana, the primary targets corresponded to the 5′ end of AC1 and the embedded AC4, the intergenic region and 5′ end of AV1 and overlapping central part of AC5. Subsequently, transgenic N. benthamiana plants were generated that were preprogrammed to express double-stranded RNA corresponding to this most targeted portion of the virus genome by using an intron-hairpin construct. These plants were shown to indeed produce ToCMoV-specific siRNAs. When challenge inoculated, most transgenic lines showed significant delays in symptom development, and two lines had immune plants. Interestingly, the levels of transgene-produced siRNAs were similar in resistant and susceptible siblings of the same line. This indicates that, in contrast to RNA viruses, the mere presence of transgene siRNAs corresponding to DNA virus sequences does not guarantee virus resistance and that other factors may play a role in determining RNA-mediated resistance to DNA viruses.
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Ferro, Camila G., F. Murilo Zerbini, Jesús Navas-Castillo, and Elvira Fiallo-Olivé. "Revealing the Complexity of Sweepovirus-Deltasatellite–Plant Host Interactions: Expanded Natural and Experimental Helper Virus Range and Effect Dependence on Virus-Host Combination." Microorganisms 9, no. 5 (May 10, 2021): 1018. http://dx.doi.org/10.3390/microorganisms9051018.
Full textAbstract:
Sweepoviruses are begomoviruses (genus Begomovirus, family Geminiviridae) with ssDNA genomes infecting sweet potato and other species of the family Convolvulaceae. Deltasatellites (genus Deltasatellite, family Tolecusatellitidae) are small-size non-coding DNA satellites associated with begomoviruses. In this study, the genetic diversity of deltasatellites associated with sweepoviruses infecting Ipomoea indica plants was analyzed by further sampling the populations where the deltasatellite sweet potato leaf curl deltasatellite 1 (SPLCD1) was initially found, expanding the search to other geographical areas in southern continental Spain and the Canary Islands. The sweepoviruses present in the samples coinfected with deltasatellites were also fully characterized by sequencing in order to define the range of viruses that could act as helper viruses in nature. Additionally, experiments were performed to assess the ability of a number of geminivirids (the monopartite tomato leaf deformation virus and the bipartite NW begomovirus Sida golden yellow vein virus, the bipartite OW begomovirus tomato leaf curl New Delhi virus, and the curtovirus beet curly top virus) to transreplicate SPLCD1 in their natural plant hosts or the experimental host Nicotiana benthamiana. The results show that SPLCD1 can be transreplicated by all the geminivirids assayed in N. benthamiana and by tomato leaf curl New Delhi virus in zucchini. The presence of SPLCD1 did not affect the symptomatology caused by the helper viruses, and its effect on viral DNA accumulation depended on the helper virus–host plant combination.
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Matsuo, Yuki, Fawzia Novianti, Miki Takehara, Toshiyuki Fukuhara, Tsutomu Arie, and Ken Komatsu. "Acibenzolar-S-Methyl Restricts Infection of Nicotiana benthamiana by Plantago Asiatica Mosaic Virus at Two Distinct Stages." Molecular Plant-Microbe Interactions® 32, no. 11 (November 2019): 1475–86. http://dx.doi.org/10.1094/mpmi-03-19-0087-r.
Full textAbstract:
Plant activators, including acibenzolar-S-methyl (ASM), are chemical compounds that stimulate plant defense responses to pathogens. ASM treatment inhibits infection by a variety of plant viruses, however, the mechanisms of this broad-spectrum and strong effect remain poorly understood. We employed green fluorescent protein (GFP)-expressing viruses and Nicotiana benthamiana plants to identify the infection stages that are restricted by ASM. ASM suppressed infection by three viral species, plantago asiatica mosaic virus (PlAMV), potato virus X (PVX), and turnip mosaic virus (TuMV), in inoculated cells. Furthermore, ASM delayed the long-distance movement of PlAMV and PVX, and the cell-to-cell (short range) movement of TuMV. The ASM-mediated delay of long-distance movement of PlAMV was not due to the suppression of viral accumulation in the inoculated leaves, indicating that ASM restricts PlAMV infection in at least two independent steps. We used Arabidopsis thaliana mutants to show that the ASM-mediated restriction of PlAMV infection requires the NPR1 gene but was independent of the dicer-like genes essential for RNA silencing. Furthermore, experiments using protoplasts showed that ASM treatment inhibited PlAMV replication without cell death. Our approach, using GFP-expressing viruses, will be useful for the analysis of mechanisms underlying plant activator–mediated virus restriction.
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Liu, Zongrang, Ralph Scorza, Jean-Michel Hily, Simon W. Scott, and Delano James. "Engineering Resistance to Multiple Prunus Fruit Viruses Through Expression of Chimeric Hairpins." Journal of the American Society for Horticultural Science 132, no. 3 (May 2007): 407–14. http://dx.doi.org/10.21273/jashs.132.3.407.
Full textAbstract:
Prunus L. fruit production is seriously affected by several predominant viruses. The development of new cultivars resistant to these viruses is challenging but highly desired by breeders and growers. We report a posttranscriptional gene silencing-based approach for engineering multivirus resistance in plants. A single chimeric transgene, PTRAP6, was created by the fusion of 400 to 500-base pair (bp) gene fragments from six major Prunus fruit viruses, including american plum line pattern virus, peach mosaic virus, plum pox virus (PPV), prune dwarf virus (PDV), prunus necrotic ringspot virus, and tomato ringspot virus (ToRSV). Both strands of PTRAP6 were found being transcribed as an ≈2.5-kilobp transcript in planta without splicing interruption. To induce gene silencing/virus resistance, we placed two copies of PTRAP6 in an inverted repeat under the control of the cauliflower mosaic virus 35S promoter and separated by an intron spacer fragment to create PTRAP6i. Inoculation of the resulting transgenic Nicotiana benthamiana Domin. plants revealed that 12 of 28 R0 PTRAP6i transgenic lines (43%) were resistant to ToRSV ranging from mild symptoms to symptom-free phenotypes. Detailed analysis of two of three highly resistant homozygous R3 generation lines demonstrated that they were resistant to all three viruses tested, including PDV, PPV, and ToRSV. The remaining three viruses targeted by PTRAP6i were either unavailable for this study or were unable to systemically infect N. benthamiana. Transgene-wide and -specific small interfering RNA species were detected along with disappearance of transgene transcript in the resistant lines, indicating that posttranscriptional gene silencing underlies the mechanism of resistance. This work presents evidence that PTRAP6i is able to confer gene silencing-based resistance to multiple Prunus fruit viruses.
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Liu, Yu, Chenyang Li, Yaqin Wang, Yi Xu, Jianxiang Wu, and Xueping Zhou. "Comprehensive Analysis of Ubiquitome Changes in Nicotiana benthamiana after Rice Stripe Virus Infection." Viruses 14, no. 11 (October 26, 2022): 2349. http://dx.doi.org/10.3390/v14112349.
Full textAbstract:
Rice stripe virus (RSV) is one of the most devastating viruses affecting rice production. During virus infection, ubiquitination plays an important role in the dynamic regulation of host defenses. We combined the ubiquitomics approach with the label-free quantitation proteomics approach to investigate potential ubiquitination status changes of Nicotiana benthamiana infected with RSV. Bioinformatics analyses were performed to elucidate potential associations between proteins with differentially ubiquitinated sites (DUSs) and various cellular components/pathways during virus infection. In total, 399 DUSs in 313 proteins were identified and quantified, among them 244 ubiquitinated lysine (Kub) sites in 186 proteins were up-regulated and 155 Kub sites in 127 proteins were down-regulated at 10 days after RSV infection. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analyses indicated that proteins with up-regulated Kub sites were significantly enriched in the ribosome. Silencing of 3-isopropylmalate dehydratase large subunit through virus-induced gene silencing delayed RSV infection, while silencing of mRNA-decapping enzyme-like protein promoted RSV symptom in the late stage of infection. Moreover, ubiquitination was observed in all seven RSV-encoded proteins. Our study supplied the comprehensive analysis of the ubiquitination changes in N. benthamiana after RSV infection, which is helpful for understanding RSV pathogenesis and RSV-host interactions.
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Lee, Shu-Chuan, Hsuan Pai, Ying-Wen Huang, Meng-Hsun He, Yun-Lin Song, Song-Yi Kuo, Wen-Chi Chang, Yau-Heiu Hsu, and Na-Sheng Lin. "Exploring the Multifunctional Roles of Odontoglossum Ringspot Virus P126 in Facilitating Cymbidium Mosaic Virus Cell-to-Cell Movement during Mixed Infection." Viruses 13, no. 8 (August 6, 2021): 1552. http://dx.doi.org/10.3390/v13081552.
Full textAbstract:
Synergistic interactions among viruses, hosts and/or transmission vectors during mixed infection can alter viral titers, symptom severity or host range. Viral suppressors of RNA silencing (VSRs) are considered one of such factors contributing to synergistic responses. Odontoglossum ringspot virus (ORSV) and cymbidium mosaic virus (CymMV), which are two of the most significant orchid viruses, exhibit synergistic symptom intensification in Phalaenopsis orchids with unilaterally enhanced CymMV movement by ORSV. In order to reveal the underlying mechanisms, we generated infectious cDNA clones of ORSV and CymMV isolated from Phalaenopsis that exerted similar unilateral synergism in both Phalaenopsis orchid and Nicotiana benthamiana. Moreover, we show that the ORSV replicase P126 is a VSR. Mutagenesis analysis revealed that mutation of the methionine in the carboxyl terminus of ORSV P126 abolished ORSV replication even though some P126 mutants preserved VSR activity, indicating that the VSR function of P126 alone is not sufficient for viral replication. Thus, P126 functions in both ORSV replication and as a VSR. Furthermore, P126 expression enhanced cell-to-cell movement and viral titers of CymMV in infected Phalaenopsis flowers and N. benthamiana leaves. Taking together, both the VSR and protein function of P126 might be prerequisites for unilaterally enhancing CymMV cell-to-cell movement by ORSV.
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Osterbaan, Larissa J., Jiyeong Choi, Jaimie Kenney, Madison Flasco, Emmanuelle Vigne, Corinne Schmitt-Keichinger, Ana Rita Rebelo, Michelle Heck, and Marc Fuchs. "The Identity of a Single Residue of the RNA-Dependent RNA Polymerase of Grapevine Fanleaf Virus Modulates Vein Clearing in Nicotiana benthamiana." Molecular Plant-Microbe Interactions® 32, no. 7 (July 2019): 790–801. http://dx.doi.org/10.1094/mpmi-12-18-0337-r.
Full textAbstract:
The mechanisms underlying host plant symptom development upon infection by viruses of the genus Nepovirus in the family Secoviridae, including grapevine fanleaf virus (GFLV), are poorly understood. In the systemic host Nicotiana benthamiana, GFLV strain GHu produces characteristic symptoms of vein clearing in apical leaves, unlike other GFLV strains such as F13, which cause an asymptomatic infection. In this study, we expanded on earlier findings and used reverse genetics to identify residue 802 (lysine, K) of the GFLV-GHu RNA1-encoded RNA-dependent RNA polymerase (1EPol) as a modulator of vein-clearing symptom development in N. benthamiana. Mutations to this site abolished (K to G, A, or Q) or attenuated (K to N or P) symptom expression. Noteworthy, residue 802 is necessary but not sufficient for vein clearing, as GFLV-F13 RNA1 carrying K802 remained asymptomatic in N. benthamiana. No correlation was found between symptom expression and RNA1 accumulation, as shown by reverse transcription-quantitative polymerase chain reaction. Additionally, the involvement of RNA silencing of vein clearing was ruled out by virus-induced gene silencing experiments and structure predictions for protein 1EPol suggested that residue 802 is flanked by strongly predicted stable secondary structures, including a conserved motif of unknown function (805LLKT/AHLK/RT/ALR814). Together, these results reveal the protein nature of the GFLV-GHu symptom determinant in N. benthamiana and provide a solid basis for probing and determining the virus-host proteome network for symptoms of vein clearing.
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Arif, Muhammad, Saif ul Islam, Saqer S. Alotaibi, Ahmed M. Elshehawi, Mohamed A. A. Ahmed, and Abdullah M. Al-Sadi. "Infectious clone construction and pathogenicity confirmation of Cotton leaf curl Multan virus (CLCuMuV), Ramie mosaic virus (RamV) and Corchorus yellow vein Vietnam virus (CoYVV) by southern blot analysis." PLOS ONE 16, no. 5 (May 14, 2021): e0251232. http://dx.doi.org/10.1371/journal.pone.0251232.
Full textAbstract:
Geminiviruses are insect-transmissible, economically vital group of plant viruses, which cause significant losses to crop production and ornamental plants across the world. During this study, infectious clones of three devastating begomoviruses, i.e., Cotton leaf curl Multan virus (CLCuMuV), Ramie mosaic virus (RamV) and Corchorus yellow vein Vietnam virus (CoYVV) were constructed by following novel protocol. All infectious clones were confirmed by cloning and sequencing. All of the infectious clones were agro-inoculated in Agrobacterium. After the agro-infiltrations, all clones were injected into Nicotiana benthamiana and jute plants under controlled condition. After 28 days of inoculation, plants exhibited typical symptoms of their corresponding viruses. All the symptomatic and asymptomatic leaves were collected from inoculated plants for further analysis. The southern blot analysis was used to confirm the infection of studied begomoviruses. At the end, all the products were sequenced and analyzed.
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Zhan, Jin, Huiping Shi, Weimin Li, Chao Zhang, and Yongqiang Zhang. "NbTMP14 Is Involved in Tomato Spotted Wilt Virus Infection and Symptom Development by Interaction with the Viral NSm Protein." Viruses 13, no. 3 (March 7, 2021): 427. http://dx.doi.org/10.3390/v13030427.
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Tomato spotted wilt virus (TSWV) is one of the most destructive plant viruses, causing severe losses in many important crops worldwide. The non-structural protein NSm of TSWV is a viral movement protein that induces viral symptoms. However, the molecular mechanisms by which NSm contributes to symptom development are unclear. Here, we present evidence that NSm directly interacts with Nicotiana benthamiana chloroplast thylakoid membrane protein TMP14 (NbTMP14) by yeast two-hybrid and bimolecular fluorescence complementation (BiFC) assays. The interaction between NSm and NbTMP14 led to the translocation of the NbTMP14 protein from the chloroplast to the cytoplasm in TSWV-infected plants, and overexpressing NSm decreased NbTMP14 mRNA accumulation. In addition, abnormal chloroplasts and starch accumulation were observed in TSWV-infected plants. Silencing of NbTMP14 by TRV VIGS also showed similar results to those of TSWV-infected plants. Overexpressing NbTMP14 in transgenic N. benthamiana plants impeded TSWV infection, and silencing NbTMP14 in N. benthamiana plants increased disease symptom severity and virus accumulation. To our knowledge, this is the first report showing that the plant chloroplast TMP14 protein is involved in viral infection. Knowledge of the interaction between NSm and NbTMP14 advances our understanding of the molecular mechanisms underlying TSWV symptom development and infection.
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Holcomb, G. E., R. A. Valverde, J. Sim, and J. Nuss. "First Report on Natural Occurrence of Tomato Spotted Wilt Tospovirus in Basil (Ocimum basilicum)." Plant Disease 83, no. 10 (October 1999): 966. http://dx.doi.org/10.1094/pdis.1999.83.10.966b.
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Virus-like symptoms were observed on basil plants (Ocimum basilicum L. ‘Mrs. Burns Lemon’ [MBL]) growing in containers and a demonstration plot at the Louisiana State University Burden Research Plantation, Baton Rouge, during July 1998. Symptoms consisted of ring spots, leaf distortion, and severe mosaic. Mechanical transmission of the suspect virus by sap inoculation from infected MBL to basil cvs. MBL, Aussie Sweet, Cinnamon, Siam Queen, and Sweet Dani was successful. Symptoms were similar to those on infected MBL. Nicotiana benthamiana Domin. reacted with local chlorotic spots followed by severe yellows, necrosis, and death. Electron microscopy of thin sections of infected basil revealed virus inclusions but no virus particles. However, infected N. benthamiana revealed the presence of 82-nm membrane-bound particles in the cytoplasm. The virus was identified from basil and N. benthamiana as the common strain of tomato spotted wilt tospovirus (TSWV) by enzyme-linked immunosorbent assay (Agdia, Elkhart, IN). An outbreak of thrips insects during the summer drought in 1998 was probably responsible for the occurrence of TSWV in basil. This is the first report of the occurrence of TSWV in basil (1). Reference: (1) A. A. Brunt et al., eds. 1996. Plant Viruses Online: Descriptions and Lists from the VIDE Database. Published online by Australian National University, Canberra.
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Kubota, Kenji, and James C. K. Ng. "Lettuce chlorosis virus P23 Suppresses RNA Silencing and Induces Local Necrosis with Increased Severity at Raised Temperatures." Phytopathology® 106, no. 6 (June 2016): 653–62. http://dx.doi.org/10.1094/phyto-09-15-0219-r.
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RNA silencing functions as an antivirus defense strategy in plants, one that plant viruses counter by producing viral suppressors of RNA silencing (VSRs). VSRs have been identified in three members of the genus Crinivirus but they do not all share identical suppression mechanisms. Here, we used Agrobacterium co-infiltration assays to investigate the suppressor activity of proteins encoded by Lettuce chlorosis virus (LCV). Of 7 LCV proteins (1b, P23, HSP70 homolog, P60, CP, CPm, and P27) tested for the suppression of silencing of green fluorescent protein (GFP) expression in wild-type Nicotiana benthamiana plants, only P23 suppressed the onset of local silencing. Small-interfering (si)RNA accumulation was reduced in leaves co-infiltrated with P23, suggesting that P23 inhibited the accumulation or enhanced the degradation of siRNA. P23 also inhibited the cell-to-cell and systemic movement of RNA silencing in GFP-expressing transgenic N. benthamiana plants. Expression of P23 via agroinfiltration of N. benthamiana leaves induced local necrosis that increased in severity at elevated temperatures, a novelty given that a direct temperature effect on necrosis severity has not been reported for the other crinivirus VSRs. These results further affirm the sophistication of crinivirus VSRs in mediating the evasion of host’s antiviral defenses and in symptom modulation.
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Senthil, G., H. Liu, V. G. Puram, A. Clark, A. Stromberg, and M. M. Goodin. "Specific and common changes in Nicotiana benthamiana gene expression in response to infection by enveloped viruses." Journal of General Virology 86, no. 9 (September 1, 2005): 2615–25. http://dx.doi.org/10.1099/vir.0.81043-0.
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Microarrays derived from Solanum tuberosum expressed sequence tags were used to test the hypothesis that genetically distinct enveloped viruses elicit unique changes in Nicotiana benthamiana gene expression. The results of our study, which included Sonchus yellow net virus (SYNV), a plant rhabdovirus that replicates in the nucleus of infected cells, and Impatiens necrotic spot virus (INSV), a plant bunyavirus that replicates in the cytoplasm, were consistent with this hypothesis. Statistically significant changes (P⩽0·01) in the expression of 275, 2646 and 4165 genes were detected in response to INSV at 2, 4 and 5 days post-inoculation (d.p.i.), respectively. In contrast, 35, 665 and 1458 genes were expressed differentially in response to SYNV at 5, 11 and 14 d.p.i., respectively. The microarray results were verified by Northern hybridization using a subset of these genes as probes. Notably, INSV, but not SYNV, induced expression of small heat-shock protein genes to high levels. In contrast to SYNV, infection by INSV resulted in downregulation of all histone genes, of which the downregulation of histone 2b expression to very low levels was confirmed by Northern hybridization. The expression of a putative WRKY transcription factor at 11 d.p.i., but not at 5 or 14 d.p.i., in SYNV-infected tissue suggested that the temporal response to virus infection was identified readily using our experimental design. Overall, infection by INSV resulted in larger fold changes in host gene expression relative to infection by SYNV. Taken together, the present data demonstrate differential responses of a common host to two genetically distinct viruses.
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Cheng, De-Jie, Xiao-Jie Xu, Zhi-Yong Yan, Carlos Kwesi Tettey, Le Fang, Guang-Ling Yang, Chao Geng, Yan-Ping Tian, and Xiang-Dong Li. "The chloroplast ribosomal protein large subunit 1 interacts with viral polymerase and promotes virus infection." Plant Physiology 187, no. 1 (May 31, 2021): 174–86. http://dx.doi.org/10.1093/plphys/kiab249.
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Abstract Chloroplasts play an indispensable role in the arms race between plant viruses and hosts. Chloroplast proteins are often recruited by plant viruses to support viral replication and movement. However, the mechanism by which chloroplast proteins regulate potyvirus infection remains largely unknown. In this study, we observed that Nicotiana benthamiana ribosomal protein large subunit 1 (NbRPL1), a chloroplast ribosomal protein, localized to the chloroplasts via its N-terminal 61 amino acids (transit peptide), and interacted with tobacco vein banding mosaic virus (TVBMV) nuclear inclusion protein b (NIb), an RNA-dependent RNA polymerase. Upon TVBMV infection, NbRPL1 was recruited into the 6K2-induced viral replication complexes in chloroplasts. Silencing of NbRPL1 expression reduced TVBMV replication. NbRPL1 competed with NbBeclin1 to bind NIb, and reduced the NbBeclin1-mediated degradation of NIb. Therefore, our results suggest that NbRPL1 interacts with NIb in the chloroplasts, reduces NbBeclin1-mediated NIb degradation, and enhances TVBMV infection.
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Butenko, Konstantin O., Inna A. Chaban, Eugene V. Skurat, Olga A. Kondakova, and Yuri F. Drygin. "Chimeric Virus Made from crTMV RNA and the Coat Protein of Potato Leafroll Virus is Targeted to the Nucleolus and Can Infect Nicotiana benthamiana Mechanically." High-Throughput 9, no. 2 (April 26, 2020): 11. http://dx.doi.org/10.3390/ht9020011.
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A genetically engineered chimeric virus crTMV-CP-PLRV composed of the crucifer-infecting tobacco mosaic virus (crTMV) RNA and the potato leafroll virus (PLRV) coat protein (CP) was obtained by agroinfiltration of Nicotiana benthamiana with the binary vector pCambia-crTMV-CPPLRV. The significant levels of the chimeric virus enabled direct visualization of crTMV-CP-PLRV in the cell and to investigate the mechanism of the pathogenesis. Localization of the crTMV-CP-PLRV in plant cells was examined by immunoblot techniques, as well as light, and transmission electron microscopy. The chimera can transfer between vascular and nonvascular tissues. The chimeric virus inoculum is capable to infect N. benthamiana mechanically. The distinguishing feature of the chimeric virus, the RNA virus with the positive genome, was found to localize in the nucleolus. We also investigated the role of the N-terminal sequence of the PLRV P3 coat protein in the cellular localization of the virus. We believe that the gene of the PLRV CP can be substituted with genes from other challenging-to-study plant pathogens to produce other useful recombinant viruses.
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Kawamura, Ryusuke, Hanako Shimura, Tomofumi Mochizuki, Satoshi T. Ohki, and Chikara Masuta. "Pollen Transmission of Asparagus virus 2 (AV-2) May Facilitate Mixed Infection by Two AV-2 Isolates in Asparagus Plants." Phytopathology® 104, no. 9 (September 2014): 1001–6. http://dx.doi.org/10.1094/phyto-12-13-0348-r.
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Asparagus virus 2 (AV-2) is a member of the genus Ilarvirus and thought to induce the asparagus decline syndrome. AV-2 is known to be transmitted by seed, and the possibility of pollen transmission was proposed 25 years ago but not verified. In AV-2 sequence analyses, we have unexpectedly found mixed infection by two distinct AV-2 isolates in two asparagus plants. Because mixed infections by two related viruses are normally prevented by cross protection, we suspected that pollen transmission of AV-2 is involved in mixed infection. Immunohistochemical analyses and in situ hybridization using AV-2-infected tobacco plants revealed that AV-2 was localized in the meristem and associated with pollen grains. To experimentally produce a mixed infection via pollen transmission, two Nicotiana benthamiana plants that were infected with each of two AV-2 isolates were crossed. Derived cleaved-amplified polymorphic sequence analysis identified each AV-2 isolate in the progeny seedlings, suggesting that pollen transmission could indeed result in a mixed infection, at least in N. benthamiana.
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Castillo, A. G., L. J. Kong, L. Hanley-Bowdoin, and E. R. Bejarano. "Interaction between a Geminivirus Replication Protein and the Plant Sumoylation System." Journal of Virology 78, no. 6 (March 15, 2004): 2758–69. http://dx.doi.org/10.1128/jvi.78.6.2758-2769.2004.
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ABSTRACT Geminiviruses are small DNA viruses that replicate in nuclei of infected plant cells after accumulation of host replication machinery. Tomato golden mosaic virus (TGMV) and Tomato yellow leaf curl Sardinia virus (TYLCSV) encode a protein, RepAC1 (or Rep), that is essential for viral replication. Rep/RepAC1 is an oligomeric protein that binds to double-stranded DNA, catalyzes cleavage and ligation of single-stranded DNA, and is sufficient for host induction. It also interacts with several host proteins, including the cell cycle regulator, retinoblastoma, and essential components of the cell DNA replication machinery, like proliferating nuclear cell antigen (PCNA) and RFC-1. To identify other cellular proteins that interact with Rep/RepAC1 protein, a Nicotiana benthamiana cDNA library was screened with a yeast two-hybrid assay. The host cell sumoylation enzyme, NbSCE1 (N. benthamiana SUMO-conjugating enzyme, homolog to Saccharomyces cerevisiae UBC9), was found to interact specifically with RepAC1. Mapping studies localized the interaction to the N-terminal half of RepAC1. Effects on geminivirus replication were observed in transgenic plants with altered levels of SUMO, the substrate for UBC9.
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Wintermantel, William M., Arturo A. Cortez, Amy G. Anchieta, Anju Gulati-Sakhuja, and Laura L. Hladky. "Co-Infection by Two Criniviruses Alters Accumulation of Each Virus in a Host-Specific Manner and Influences Efficiency of Virus Transmission." Phytopathology® 98, no. 12 (December 2008): 1340–45. http://dx.doi.org/10.1094/phyto-98-12-1340.
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Tomato chlorosis virus (ToCV), and Tomato infectious chlorosis virus (TICV), family Closteroviridae, genus Crinivirus, cause interveinal chlorosis, leaf brittleness, and limited necrotic flecking or bronzing on tomato leaves. Both viruses cause a decline in plant vigor and reduce fruit yield, and are emerging as serious production problems for field and greenhouse tomato growers in many parts of the world. The viruses have been found together in tomato, indicating that infection by one Crinivirus sp. does not prevent infection by a second. Transmission efficiency and virus persistence in the vector varies significantly among the four different whitefly vectors of ToCV; Bemisia tabaci biotypes A and B, Trialeurodes abutilonea, and T. vaporariorum. Only T. vaporariorum can transmit TICV. In order to elucidate the effects of co-infection on Crinivirus sp. accumulation and transmission efficiency, we established Physalis wrightii and Nicotiana benthamiana source plants, containing either TICV or ToCV alone or both viruses together. Vectors were allowed to feed separately on all virus sources, as well as virus-free plants, then were transferred to young plants of both host species. Plants were tested by quantitative reverse-transcription polymerase chain reaction, and results indicated host-specific differences in accumulation by TICV and ToCV and alteration of accumulation patterns during co-infection compared with single infection. In N. benthamiana, TICV titers increased during co-infection compared with levels in single infection, while ToCV titers decreased. However, in P. wrightii, titers of both TICV and ToCV decreased during mixed infection compared with single infection, although to different degrees. Vector transmission efficiency of both viruses corresponded with virus concentration in the host in both single and mixed infections. This illustrates that Crinivirus epidemiology is impacted not only by vector transmission specificity and incidence of hosts but also by interactions between viruses and efficiency of accumulation in host plants.
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Sharma, Monika, Zsuzsanna Sasvari, and Peter D. Nagy. "Inhibition of Sterol Biosynthesis Reduces Tombusvirus Replication in Yeast and Plants." Journal of Virology 84, no. 5 (December 16, 2009): 2270–81. http://dx.doi.org/10.1128/jvi.02003-09.
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ABSTRACT The replication of plus-strand RNA viruses depends on subcellular membranes. Recent genome-wide screens have revealed that the sterol biosynthesis genes ERG25 and ERG4 affected the replication of Tomato bushy stunt virus (TBSV) in a yeast model host. To further our understanding of the role of sterols in TBSV replication, we demonstrate that the downregulation of ERG25 or the inhibition of the activity of Erg25p with an inhibitor (6-amino-2-n-pentylthiobenzothiazole; APB) leads to a 3- to 5-fold reduction in TBSV replication in yeast. In addition, the sterol biosynthesis inhibitor lovastatin reduced TBSV replication by 4-fold, confirming the importance of sterols in viral replication. We also show reduced stability for the p92pol viral replication protein as well as a decrease in the in vitro activity of the tombusvirus replicase when isolated from APB-treated yeast. Moreover, APB treatment inhibits TBSV RNA accumulation in plant protoplasts and in Nicotiana benthamiana leaves. The inhibitory effect of APB on TBSV replication can be complemented by exogenous stigmasterol, the main plant sterol, suggesting that sterols are required for TBSV replication. The silencing of SMO1 and SMO2 genes, which are orthologs of ERG25, in N. benthamiana reduced TBSV RNA accumulation but had a lesser inhibitory effect on the unrelated Tobacco mosaic virus, suggesting that various viruses show different levels of dependence on sterol biosynthesis for their replication.
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Were, H. K., S. Winter, and E. Maiss. "Viruses Infecting Cassava in Kenya." Plant Disease 88, no. 1 (January 2004): 17–22. http://dx.doi.org/10.1094/pdis.2004.88.1.17.
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A survey of cassava viruses was conducted in major cassava-growing regions of Kenya. A total of 185 leaf samples and 62 stem cuttings from plants with viral disease symptoms were collected and analyzed by biological, electron microscopy, enzyme-linked immunosorbent assay, and polymerase chain reaction. All samples from western Kenya had cassava begomoviruses (African cassava mosaic virus [ACMV], East African cassava mosaic virus [EACMV], and Uganda variant [EACMV-UG]) in either single or in mixed infection. However, all samples from the Coast region were infected with only EACMV, a begomovirus. In addition, 15 samples had mixed infections of EACMV and three other hitherto unidentified filamentous viruses. The viruses observed were 200, 500, 650, and 750 nm long, respectively. In addition to rod-shaped and some flexuous viruses, as seen in a crude sap preparation, pinwheels also were observed, indicating a possible association of some of the viruses with the Potyviridae family. The symptoms induced by these viruses in Nicotiana benthamiana were very severe and often caused about 50% death of the test plants. Back inoculation onto cassava resulted in 100% infections. This finding provides evidence that, other than begomoviruses that cause serious diseases of cassava in Africa, filamentous viruses also are present and, despite their limited distribution, they could reach local significance and, most probably, be as serious as begomoviruses. The implications of these findings are discussed and recommendations for future work suggested.