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1.
EMBO J ; 42(18): e113378, 2023 09 18.
Article in English | MEDLINE | ID: mdl-37431920

ABSTRACT

In virus-host interactions, nucleic acid-directed first lines of defense that allow viral clearance without compromising growth are of paramount importance. Plants use the RNA interference pathway as a basal antiviral immune system, but additional RNA-based mechanisms of defense also exist. The infectivity of a plant positive-strand RNA virus, alfalfa mosaic virus (AMV), relies on the demethylation of viral RNA by the recruitment of the cellular N6-methyladenosine (m6 A) demethylase ALKBH9B, but how demethylation of viral RNA promotes AMV infection remains unknown. Here, we show that inactivation of the Arabidopsis cytoplasmic YT521-B homology domain (YTH)-containing m6 A-binding proteins ECT2, ECT3, and ECT5 is sufficient to restore AMV infectivity in partially resistant alkbh9b mutants. We further show that the antiviral function of ECT2 is distinct from its previously demonstrated function in the promotion of primordial cell proliferation: an ect2 mutant carrying a small deletion in its intrinsically disordered region is partially compromised for antiviral defense but not for developmental functions. These results indicate that the m6 A-YTHDF axis constitutes a novel branch of basal antiviral immunity in plants.


Subject(s)
Arabidopsis Proteins , Arabidopsis , RNA Viruses , Antiviral Agents , Plant Proteins/metabolism , RNA-Binding Proteins/metabolism , Arabidopsis/metabolism , RNA, Viral/genetics , Arabidopsis Proteins/genetics , Arabidopsis Proteins/metabolism , Intracellular Signaling Peptides and Proteins/metabolism
2.
Proc Natl Acad Sci U S A ; 121(21): e2401748121, 2024 May 21.
Article in English | MEDLINE | ID: mdl-38739789

ABSTRACT

Potyviridae, the largest family of plant RNA viruses, includes many important pathogens that significantly reduce the yields of many crops worldwide. In this study, we report that the 6-kilodalton peptide 1 (6K1), one of the least characterized potyviral proteins, is an endoplasmic reticulum-localized protein. AI-assisted structure modeling and biochemical assays suggest that 6K1 forms pentamers with a central hydrophobic tunnel, can increase the cell membrane permeability of Escherichia coli and Nicotiana benthamiana, and can conduct potassium in Saccharomyces cerevisiae. An infectivity assay showed that viral proliferation is inhibited by mutations that affect 6K1 multimerization. Moreover, the 6K1 or its homologous 7K proteins from other viruses of the Potyviridae family also have the ability to increase cell membrane permeability and transmembrane potassium conductance. Taken together, these data reveal that 6K1 and its homologous 7K proteins function as viroporins in viral infected cells.


Subject(s)
Nicotiana , Nicotiana/virology , Nicotiana/metabolism , Potyviridae/genetics , Potyviridae/metabolism , Viral Proteins/metabolism , Viral Proteins/genetics , Cell Membrane Permeability , Endoplasmic Reticulum/metabolism , Endoplasmic Reticulum/virology , Saccharomyces cerevisiae/genetics , Saccharomyces cerevisiae/metabolism , Viroporin Proteins/metabolism , Viroporin Proteins/genetics , Escherichia coli/genetics , Escherichia coli/metabolism , Plant Viruses/genetics , Plant Viruses/physiology , Plant Diseases/virology , Potassium/metabolism
3.
Proc Natl Acad Sci U S A ; 120(42): e2302069120, 2023 10 17.
Article in English | MEDLINE | ID: mdl-37824524

ABSTRACT

Stem cells are essential for the development and organ regeneration of multicellular organisms, so their infection by pathogenic viruses must be prevented. Accordingly, mammalian stem cells are highly resistant to viral infection due to dedicated antiviral pathways including RNA interference (RNAi). In plants, a small group of stem cells harbored within the shoot apical meristem generate all postembryonic above-ground tissues, including the germline cells. Many viruses do not proliferate in these cells, yet the molecular bases of this exclusion remain only partially understood. Here, we show that a plant-encoded RNA-dependent RNA polymerase, after activation by the plant hormone salicylic acid, amplifies antiviral RNAi in infected tissues. This provides stem cells with RNA-based virus sequence information, which prevents virus proliferation. Furthermore, we find RNAi to be necessary for stem cell exclusion of several unrelated RNA viruses, despite their ability to efficiently suppress RNAi in the rest of the plant. This work elucidates a molecular pathway of great biological and economic relevance and lays the foundations for our future understanding of the unique systems underlying stem cell immunity.


Subject(s)
RNA Viruses , Salicylic Acid , Animals , RNA Interference , RNA Viruses/genetics , Stem Cells/metabolism , Plant Stems/genetics , Plant Stems/metabolism , RNA, Small Interfering/genetics , RNA, Viral/genetics , Mammals/genetics
4.
Proc Natl Acad Sci U S A ; 120(43): e2221859120, 2023 10 24.
Article in English | MEDLINE | ID: mdl-37844250

ABSTRACT

Metastatic cancer accounts for 90% of all cancer-related deaths and continues to be one of the toughest challenges in cancer treatment. A growing body of data indicates that S100A9, a major regulator of inflammation, plays a central role in cancer progression and metastasis, particularly in the lungs, where S100A9 forms a premetastatic niche. Thus, we developed a vaccine against S100A9 derived from plant viruses and virus-like particles. Using multiple tumor mouse models, we demonstrate the effectiveness of the S100A9 vaccine candidates in preventing tumor seeding within the lungs and outgrowth of metastatic disease. The elicited antibodies showed high specificity toward S100A9 without cross-reactivity toward S100A8, another member of the S100A family. When tested in metastatic mouse models of breast cancer and melanoma, the vaccines significantly reduced lung tumor nodules after intravenous challenge or postsurgical removal of the primary tumor. Mechanistically, the vaccines reduce the levels of S100A9 within the lungs and sera, thereby increasing the expression of immunostimulatory cytokines with antitumor function [(interleukin) IL-12 and interferonγ] while reducing levels of immunosuppressive cytokines (IL-10 and transforming growth factorß). This also correlated with decreased myeloid-derived suppressor cell populations within the lungs. This work has wide-ranging impact, as S100A9 is overexpressed in multiple cancers and linked with poor prognosis in cancer patients. The data presented lay the foundation for the development of therapies and vaccines targeting S100A9 to prevent metastasis.


Subject(s)
Lung Neoplasms , Viral Vaccines , Humans , Mice , Animals , Calgranulin B/metabolism , Lung Neoplasms/prevention & control , Calgranulin A/metabolism , Lung/pathology , Cytokines/metabolism
5.
EMBO J ; 40(15): e108050, 2021 08 02.
Article in English | MEDLINE | ID: mdl-34155657

ABSTRACT

Selective autophagy mediates specific degradation of unwanted cytoplasmic components to maintain cellular homeostasis. The suppressor of gene silencing 3 (SGS3) and RNA-dependent RNA polymerase 6 (RDR6)-formed bodies (SGS3/RDR6 bodies) are essential for siRNA amplification in planta. However, whether autophagy receptors regulate selective turnover of SGS3/RDR6 bodies is unknown. By analyzing the transcriptomic response to virus infection in Arabidopsis, we identified a virus-induced small peptide 1 (VISP1) composed of 71 amino acids, which harbor a ubiquitin-interacting motif that mediates interaction with autophagy-related protein 8. Overexpression of VISP1 induced selective autophagy and compromised antiviral immunity by inhibiting SGS3/RDR6-dependent viral siRNA amplification, whereas visp1 mutants exhibited opposite effects. Biochemistry assays demonstrate that VISP1 interacted with SGS3 and mediated autophagic degradation of SGS3/RDR6 bodies. Further analyses revealed that overexpression of VISP1, mimicking the sgs3 mutant, impaired biogenesis of endogenous trans-acting siRNAs and up-regulated their targets. Collectively, we propose that VISP1 is a small peptide receptor functioning in the crosstalk between selective autophagy and RNA silencing.


Subject(s)
Arabidopsis Proteins/metabolism , Arabidopsis/genetics , Arabidopsis/immunology , Peptides/genetics , RNA-Dependent RNA Polymerase/metabolism , Arabidopsis/metabolism , Arabidopsis/virology , Arabidopsis Proteins/genetics , Autophagosomes/physiology , Autophagy/physiology , Autophagy-Related Protein 8 Family/metabolism , Gene Expression Regulation, Plant , Mutation , Peptides/metabolism , Plant Immunity , Plants, Genetically Modified , RNA, Small Interfering , RNA-Dependent RNA Polymerase/genetics , Nicotiana/genetics
6.
J Gen Virol ; 105(3)2024 03.
Article in English | MEDLINE | ID: mdl-38441560

ABSTRACT

Plant viruses are transmitted mechanically or by vegetative propagation, and by vectors such as arthropods, fungi, nematodes, or parasitic plants. Sources to access available information regarding plant virus transmissions are scattered and require extensive literature searches. Here, a recently created plant virus transmission database is described. This was developed to provide access to the modes of transmission and vectors of over 1600 plant viruses. The database was compiled using over 3500 publication records spanning the last 100 years. The information is publicly accessible via https://library.wur.nl/WebQuery/virus and fully searchable by virus name, taxonomic position, mode of transmission or vector.


Subject(s)
Arthropods , Plant Viruses , Animals , Plant Viruses/genetics , Databases, Factual
7.
Planta ; 259(2): 38, 2024 Jan 16.
Article in English | MEDLINE | ID: mdl-38227024

ABSTRACT

MAIN CONCLUSION: Silencing of an ascorbate oxidase (AO) gene in N. benthamiana enhanced disease severity from cucumber mosaic virus (CMV), showing higher accumulation and expansion of the spreading area of CMV. A Nicotiana benthamiana ascorbate oxidase (NbAO) gene was found to be induced upon cucumber mosaic virus (CMV) infection. Virus-induced gene silencing (VIGS) was employed to elucidate the function of AO in N. benthamiana. The tobacco rattle virus (TRV)-mediated VIGS resulted in an efficient silencing of the NbAO gene, i.e., 97.5% and 78.8% in relative quantification as compared to the control groups (TRV::eGFP- and the mock-inoculated plants), respectively. In addition, AO enzymatic activity decreased in the TRV::NtAO-silenced plants as compared to control. TRV::NtAO-mediated NbAO silencing induced a greater reduction in plant height by 15.2% upon CMV infection. CMV titer at 3 dpi was increased in the systemic leaves of NbAO-silenced plants (a 35-fold change difference as compared to the TRV::eGFP-treated group). Interestingly, CMV and TRV titers vary in different parts of systemically infected N. benthamiana leaves. In TRV::eGFP-treated plants, CMV accumulated only at the top half of the leaf, whereas the bottom half of the leaf was "occupied" by TRV. In contrast, in the NbAO-silenced plants, CMV accumulated in both the top and the bottom half of the leaf, suggesting that the silencing of the NbAO gene resulted in the expansion of the spreading area of CMV. Our data suggest that the AO gene might function as a resistant factor against CMV infection in N. benthamiana.


Subject(s)
Cucumovirus , Cytomegalovirus Infections , Nicotiana/genetics , Ascorbate Oxidase , Plant Leaves/genetics
8.
Plant Biotechnol J ; 2024 Jun 03.
Article in English | MEDLINE | ID: mdl-38831638

ABSTRACT

The foundation of most food production systems underpinning global food security is the careful management of soil resources. Embedded in the concept of soil health is the impact of diverse soil-borne pests and pathogens, and phytoparasitic nematodes represent a particular challenge. Root-knot nematodes and cyst nematodes are severe threats to agriculture, accounting for annual yield losses of US$157 billion. The control of soil-borne phytoparasitic nematodes conventionally relies on the use of chemical nematicides, which can have adverse effects on the environment and human health due to their persistence in soil, plants, and water. Nematode-resistant plants offer a promising alternative, but genetic resistance is species-dependent, limited to a few crops, and breeding and deploying resistant cultivars often takes years. Novel approaches for the control of phytoparasitic nematodes are therefore required, those that specifically target these parasites in the ground whilst minimizing the impact on the environment, agricultural ecosystems, and human health. In addition to the development of next-generation, environmentally safer nematicides, promising biochemical strategies include the combination of RNA interference (RNAi) with nanomaterials that ensure the targeted delivery and controlled release of double-stranded RNA. Genome sequencing has identified more than 75 genes in root knot and cyst nematodes that have been targeted with RNAi so far. But despite encouraging results, the delivery of dsRNA to nematodes in the soil remains inefficient. In this review article, we describe the state-of-the-art RNAi approaches targeting phytoparasitic nematodes and consider the potential benefits of nanotechnology to improve dsRNA delivery.

9.
J Virol ; 97(6): e0022123, 2023 06 29.
Article in English | MEDLINE | ID: mdl-37199623

ABSTRACT

Plant viruses depend on a number of host factors for successful infection. Deficiency of critical host factors confers recessively inherited viral resistance in plants. For example, loss of Essential for poteXvirus Accumulation 1 (EXA1) in Arabidopsis thaliana confers resistance to potexviruses. However, the molecular mechanism of how EXA1 assists potexvirus infection remains largely unknown. Previous studies reported that the salicylic acid (SA) pathway is upregulated in exa1 mutants, and EXA1 modulates hypersensitive response-related cell death during EDS1-dependent effector-triggered immunity. Here, we show that exa1-mediated viral resistance is mostly independent of SA and EDS1 pathways. We demonstrate that Arabidopsis EXA1 interacts with three members of the eukaryotic translation initiation factor 4E (eIF4E) family, eIF4E1, eIFiso4E, and novel cap-binding protein (nCBP), through the eIF4E-binding motif (4EBM). Expression of EXA1 in exa1 mutants restored infection by the potexvirus Plantago asiatica mosaic virus (PlAMV), but EXA1 with mutations in 4EBM only partially restored infection. In virus inoculation experiments using Arabidopsis knockout mutants, EXA1 promoted PlAMV infection in concert with nCBP, but the functions of eIFiso4E and nCBP in promoting PlAMV infection were redundant. By contrast, the promotion of PlAMV infection by eIF4E1 was, at least partially, EXA1 independent. Taken together, our results imply that the interaction of EXA1-eIF4E family members is essential for efficient PlAMV multiplication, although specific roles of three eIF4E family members in PlAMV infection differ. IMPORTANCE The genus Potexvirus comprises a group of plant RNA viruses, including viruses that cause serious damage to agricultural crops. We previously showed that loss of Essential for poteXvirus Accumulation 1 (EXA1) in Arabidopsis thaliana confers resistance to potexviruses. EXA1 may thus play a critical role in the success of potexvirus infection; hence, elucidation of its mechanism of action is crucial for understanding the infection process of potexviruses and for effective viral control. Previous studies reported that loss of EXA1 enhances plant immune responses, but our results indicate that this is not the primary mechanism of exa1-mediated viral resistance. Here, we show that Arabidopsis EXA1 assists infection by the potexvirus Plantago asiatica mosaic virus (PlAMV) by interacting with the eukaryotic translation initiation factor 4E family. Our results imply that EXA1 contributes to PlAMV multiplication by regulating translation.


Subject(s)
Arabidopsis Proteins , Arabidopsis , Eukaryotic Initiation Factor-4E , Plant Diseases , Potexvirus , Arabidopsis/metabolism , Arabidopsis/virology , Eukaryotic Initiation Factor-4E/genetics , Eukaryotic Initiation Factor-4E/metabolism , Plant Diseases/genetics , Potexvirus/physiology , Arabidopsis Proteins/metabolism , Disease Resistance/genetics , Protein Binding , Amino Acid Motifs , Gene Deletion , Plant Cells/virology , Protein Biosynthesis/genetics
10.
J Virol ; 97(9): e0046323, 2023 09 28.
Article in English | MEDLINE | ID: mdl-37668368

ABSTRACT

Plant viruses induce various disease symptoms that substantially impact agriculture, but the underlying mechanisms of viral disease in plants are poorly understood. Kobu-sho is a disease in gentian that shows gall formation with ectopic development of lignified cells and vascular tissues such as xylem. Here, we show that a gene fragment of gentian Kobu-sho-associated virus, which is designated as Kobu-sho-inducing factor (KOBU), induces gall formation accompanied by ectopic development of lignified cells and xylem-like tissue in Nicotiana benthamiana. Transgenic gentian expressing KOBU exhibited tumorous symptoms, confirming the gall-forming activity of KOBU. Surprisingly, KOBU expression can also induce differentiation of an additional leaf-like tissue on the abaxial side of veins in normal N. benthamiana and gentian leaves. Transcriptome analysis with Arabidopsis thaliana expressing KOBU revealed that KOBU activates signaling pathways that regulate xylem development. KOBU protein forms granules and plate-like structures and co-localizes with mRNA splicing factors within the nucleus. Our findings suggest that KOBU is a novel pleiotropic virulence factor that stimulates vascular and leaf development. IMPORTANCE While various mechanisms determine disease symptoms in plants depending on virus-host combinations, the details of how plant viruses induce symptoms remain largely unknown in most plant species. Kobu-sho is a disease in gentian that shows gall formation with ectopic development of lignified cells and vascular tissues such as xylem. Our findings demonstrate that a gene fragment of gentian Kobu-sho-associated virus (GKaV), which is designated as Kobu-sho-inducing factor, induces the gall formation accompanied by the ectopic development of lignified cells and xylem-like tissue in Nicotiana benthamiana. The molecular mechanism by which gentian Kobu-sho-associated virus induces the Kobu-sho symptoms will provide new insight into not only plant-virus interactions but also the regulatory mechanisms underlying vascular and leaf development.


Subject(s)
Gentiana , Nicotiana , Plant Tumors , Plant Viruses , Virulence Factors , Xylem , Arabidopsis/genetics , Arabidopsis/metabolism , Arabidopsis/virology , Gene Expression Profiling , Gene Expression Regulation, Plant , Gentiana/virology , Plant Viruses/genetics , Plant Viruses/pathogenicity , Nicotiana/metabolism , Nicotiana/virology , Xylem/metabolism , Virulence Factors/genetics , Virulence Factors/metabolism , Plant Leaves , Plant Tumors/virology , Signal Transduction , RNA Splicing Factors
11.
New Phytol ; 241(4): 1415-1420, 2024 Feb.
Article in English | MEDLINE | ID: mdl-38058221

ABSTRACT

Stem cells are vital for plant development and reproduction. The stem cells within shoot apical meristems are known to possess exceptionally effective antiviral defenses against pathogenic viruses which preclude their infection, yet how this is achieved remains poorly understood and scarcely investigated. In this Tansley Insight, we connect very recent experimental results with previous work to summarize the known molecular mechanisms determining stem cell antiviral immunity. More broadly, we attempt to define the viral features triggering immunity and the global consequences of virus infection in these essential cells. This brief article will highlight how these phenomena are fascinating, complex and often crucial for virus-host interactions, while emphasizing the potential for discovery in their investigation.


Subject(s)
Meristem , Plants , Plant Development , Stem Cells
12.
New Phytol ; 241(2): 845-860, 2024 Jan.
Article in English | MEDLINE | ID: mdl-37920100

ABSTRACT

Specificity in plant-pathogen gene-for-gene (GFG) interactions is determined by the recognition of pathogen proteins by the products of plant resistance (R) genes. The evolutionary dynamics of R genes in plant-virus systems is poorly understood. We analyse the evolution of the L resistance locus to tobamoviruses in the wild pepper Capsicum annuum var. glabriusculum (chiltepin), a crop relative undergoing incipient domestication. The frequency, and the genetic and phenotypic diversity, of the L locus was analysed in 41 chiltepin populations under different levels of human management over its distribution range in Mexico. The frequency of resistance was lower in Cultivated than in Wild populations. L-locus genetic diversity showed a strong spatial structure with no isolation-by-distance pattern, suggesting environment-specific selection, possibly associated with infection by the highly virulent tobamoviruses found in the surveyed regions. L alleles differed in recognition specificity and in the expression of resistance at different temperatures, broad-spectrum recognition of P0 + P1 pathotypes and expression above 32°C being ancestral traits that were repeatedly lost along L-locus evolution. Overall, loss of resistance co-occurs with incipient domestication and broad-spectrum resistance expressed at high temperatures has apparent fitness costs. These findings contribute to understand the role of fitness trade-offs in plant-virus coevolution.


Subject(s)
Capsicum , Disease Resistance , Humans , Disease Resistance/genetics , Temperature , Alleles , Mexico , Capsicum/genetics , Plant Diseases/genetics
13.
Plant Cell Environ ; 2024 Jan 02.
Article in English | MEDLINE | ID: mdl-38168864

ABSTRACT

Reactive oxygen species (ROS) are important signalling molecules that influence many aspects of plant biology. One way in which ROS influence plant growth and development is by modifying intercellular trafficking through plasmodesmata (PD). Viruses have evolved to use PD for their local cell-to-cell spread between plant cells, so it is therefore not surprising that they have found ways to modulate ROS and redox signalling to optimise PD function for their benefit. This review examines how intracellular signalling via ROS and redox pathways regulate intercellular trafficking via PD during development and stress. The relationship between viruses and ROS-redox systems, and the strategies viruses employ to control PD function by interfering with ROS-redox in plants is also discussed.

14.
J Exp Bot ; 75(5): 1265-1273, 2024 Feb 28.
Article in English | MEDLINE | ID: mdl-37940194

ABSTRACT

Calcium is a universal messenger in different kingdoms of living organisms and regulates most physiological processes, including defense against pathogens. The threat of viral infections in humans has become very clear in recent years, and this has triggered detailed research into all aspects of host-virus interactions, including the suppression of calcium signaling in infected cells. At the same time, however, the threat of plant viral infections is underestimated in society, and research in the field of calcium signaling during plant viral infections is scarce. Here we highlight an emerging role of calcium signaling for antiviral protection in plants, in parallel with the known evidence from studies of animal cells. Obtaining more knowledge in this domain might open up new perspectives for future crop protection and the improvement of food security.


Subject(s)
Plant Viruses , Virus Diseases , Humans , Animals , Calcium Signaling , Plants/genetics , Plant Viruses/physiology , Antiviral Agents , Plant Diseases , Plant Immunity
15.
Virol J ; 21(1): 86, 2024 04 15.
Article in English | MEDLINE | ID: mdl-38622686

ABSTRACT

BACKGROUND: Viruses have notable effects on agroecosystems, wherein they can adversely affect plant health and cause problems (e.g., increased biosecurity risks and economic losses). However, our knowledge of their diversity and interactions with specific host plants in ecosystems remains limited. To enhance our understanding of the roles that viruses play in agroecosystems, comprehensive analyses of the viromes of a wide range of plants are essential. High-throughput sequencing (HTS) techniques are useful for conducting impartial and unbiased investigations of plant viromes, ultimately forming a basis for generating further biological and ecological insights. This study was conducted to thoroughly characterize the viral community dynamics in individual plants. RESULTS: An HTS-based virome analysis in conjunction with proximity sampling and a tripartite network analysis were performed to investigate the viral diversity in chunkung (Cnidium officinale) plants. We identified 61 distinct chunkung plant-associated viruses (27 DNA and 34 RNA viruses) from 21 known genera and 6 unclassified genera in 14 known viral families. Notably, 12 persistent viruses (7 DNA and 5 RNA viruses) were exclusive to dwarfed chunkung plants. The detection of viruses from the families Partitiviridae, Picobirnaviridae, and Spinareoviridae only in the dwarfed plants suggested that they may contribute to the observed dwarfism. The co-infection of chunkung by multiple viruses is indicative of a dynamic and interactive viral ecosystem with significant sequence variability and evidence of recombination. CONCLUSIONS: We revealed the viral community involved in chunkung. Our findings suggest that chunkung serves as a significant reservoir for a variety of plant viruses. Moreover, the co-infection rate of individual plants was unexpectedly high. Future research will need to elucidate the mechanisms enabling several dozen viruses to co-exist in chunkung. Nevertheless, the important insights into the chunkung virome generated in this study may be relevant to developing effective plant viral disease management and control strategies.


Subject(s)
Coinfection , Dwarfism , Plant Viruses , RNA Viruses , Humans , Virome , Ecosystem , Cnidium/genetics , RNA, Viral/genetics , High-Throughput Nucleotide Sequencing/methods , Plant Viruses/genetics , DNA , Phylogeny
16.
Mol Pharm ; 21(6): 2727-2739, 2024 Jun 03.
Article in English | MEDLINE | ID: mdl-38709860

ABSTRACT

The dramatic effectiveness of recent mRNA (mRNA)-based COVID vaccines delivered in lipid nanoparticles has highlighted the promise of mRNA therapeutics in general. In this report, we extend our earlier work on self-amplifying mRNAs delivered in spherical in vitro reconstituted virus-like particles (VLPs), and on drug delivery using cylindrical virus particles. In particular, we carry out separate in vitro assemblies of a self-amplifying mRNA gene in two different virus-like particles: one spherical, formed with the capsid protein of cowpea chlorotic mottle virus (CCMV), and the other cylindrical, formed from the capsid protein of tobacco mosaic virus (TMV). The mRNA gene is rendered self-amplifying by genetically fusing it to the RNA-dependent RNA polymerase (RdRp) of Nodamura virus, and the relative efficacies of cell uptake and downstream protein expression resulting from their CCMV- and TMV-packaged forms are compared directly. This comparison is carried out by their transfections into cells in culture: expressions of two self-amplifying genes, enhanced yellow fluorescent protein (EYFP) and Renilla luciferase (Luc), packaged alternately in CCMV and TMV VLPs, are quantified by fluorescence and chemiluminescence levels, respectively, and relative numbers of the delivered mRNAs are measured by quantitative real-time PCR. The cellular uptake of both forms of these VLPs is further confirmed by confocal microscopy of transfected cells. Finally, VLP-mediated delivery of the self-amplifying-mRNA in mice following footpad injection is shown by in vivo fluorescence imaging to result in robust expression of EYFP in the draining lymph nodes, suggesting the potential of these plant virus-like particles as a promising mRNA gene and vaccine delivery modality. These results establish that both CCMV and TMV VLPs can deliver their in vitro packaged mRNA genes to immune cells and that their self-amplifying forms significantly enhance in situ expression. Choice of one VLP (CCMV or TMV) over the other will depend on which geometry of nucleocapsid is self-assembled more efficiently for a given length and sequence of RNA, and suggests that these plant VLP gene delivery systems will prove useful in a wide variety of medical applications, both preventive and therapeutic.


Subject(s)
Capsid Proteins , RNA, Messenger , Tobacco Mosaic Virus , Animals , RNA, Messenger/genetics , RNA, Messenger/metabolism , Mice , Tobacco Mosaic Virus/genetics , Capsid Proteins/genetics , Bromovirus/genetics , Nanoparticles/chemistry , Humans , Female , COVID-19 Vaccines/administration & dosage , Virion/genetics , RNA-Dependent RNA Polymerase/metabolism , RNA-Dependent RNA Polymerase/genetics , Vaccines, Virus-Like Particle/administration & dosage , Liposomes
17.
Arch Virol ; 169(3): 58, 2024 Mar 01.
Article in English | MEDLINE | ID: mdl-38424260

ABSTRACT

In this study, we determined the complete genome sequence of a novel totivirus, tentatively named "Mangifera indica totivirus 1" (MiTV1), identified in 'Apple' mango in China. The double-stranded RNA genome of MiTV1 is 4800 base pairs (bp) in length and contains two open reading frames (ORFs) encoding a putative coat protein (CP) and an RNA-dependent RNA polymerase (RdRp). Phylogenetic analysis based on RdRp and CP amino acid sequences showed that MiTV1 is closely related to members of the genus Totivirus in the family Totiviridae. To our knowledge, this is the first report of a totivirus found in Mangifera indica.


Subject(s)
Mangifera , Totivirus , Totivirus/genetics , Mangifera/genetics , Phylogeny , Amino Acid Sequence , RNA, Double-Stranded , RNA-Dependent RNA Polymerase/genetics , Open Reading Frames , Genome, Viral , RNA, Viral/genetics
18.
Arch Virol ; 169(7): 151, 2024 Jun 20.
Article in English | MEDLINE | ID: mdl-38902586

ABSTRACT

A new fusagra-like virus infecting papaya (Carica papaya L.) was genetically characterized. The genome of the virus, provisionally named "papaya sticky fruit-associated virus" (PSFaV), is a single molecule of double-stranded RNA, 9,199 nucleotides (nt) in length, containing two discontinuous open reading frames. Pairwise sequence comparisons based on complete RNA-dependent-RNA-polymerase (RdRp) sequences revealed identity of 79.4% and 83.3% at the nt and amino acid (aa) level, respectively, to babaco meleira-like virus (BabMelV), an uncharacterized virus sequence discovered in babaco (Vasconcellea x heilbornii) in Ecuador. Additional plant-associated viruses with sequence identity in the 50% range included papaya meleira virus (PMeV) isolates from Brazil. Phylogenetic analysis based on the amino acid sequences of the capsid protein (CP), RdRp, and CP-RdRp fusion protein genes placed PSFaV in a group within a well-supported clade that shares a recent ancestor with Sclerotium rolfsii RNA virus 2 and Phlebiopsis gigantea mycovirus dsRNA 2, two fungus-associated fusagraviruses. Genomic features and phylogenetic relatedness suggest that PSFaV, along with its closest relative BabMelV, represent a species of novel plant-associated virus classified within the recently established family Fusagraviridae.


Subject(s)
Carica , Genome, Viral , Open Reading Frames , Phylogeny , Plant Diseases , RNA, Viral , Carica/virology , Genome, Viral/genetics , Ecuador , Plant Diseases/virology , RNA, Viral/genetics , Whole Genome Sequencing , RNA Viruses/genetics , RNA Viruses/classification , RNA Viruses/isolation & purification , RNA-Dependent RNA Polymerase/genetics , Capsid Proteins/genetics
19.
Arch Virol ; 169(8): 160, 2024 Jul 09.
Article in English | MEDLINE | ID: mdl-38981875

ABSTRACT

A novel monopartite dsRNA virus, tentatively named "sponge gourd amalgavirus 1" (SGAV1), was discovered by high-throughput sequencing in sponge gourd (Luffa cylindrica) displaying mosaic symptoms in Jiashan County, Zhejiang Province, China. The genome of SGAV1 is 3,447 nucleotides in length and contains partially overlapping open reading frames (ORFs) encoding a putative replication factory matrix-like protein and a fusion protein, respectively. The fusion protein of SGAV1 shares 57.07% identity with the homologous protein of salvia miltiorrhiza amalgavirus 1 (accession no. DAZ91057.1). Phylogenetic analysis based on the RNA-dependent RNA polymerase (RdRp) protein suggests that SGAV1 belongs to the genus Amalgavirus of the family Amalgaviridae. Moreover, analysis of SGAV1-derived small interfering RNAs indicated that SGAV1 was actively replicating in the host plant. Semi-quantitative RT-PCR showed higher levels of SGAV1 expression in leaves than in flowers and fruits. This is the first report of a novel amalgavirus found in sponge gourd in China.


Subject(s)
Genome, Viral , Luffa , Open Reading Frames , Phylogeny , Genome, Viral/genetics , Luffa/virology , Animals , China , Double Stranded RNA Viruses/genetics , Double Stranded RNA Viruses/classification , Double Stranded RNA Viruses/isolation & purification , Whole Genome Sequencing , Viral Proteins/genetics , RNA, Viral/genetics , RNA-Dependent RNA Polymerase/genetics
20.
Arch Insect Biochem Physiol ; 115(3): e22102, 2024 Mar.
Article in English | MEDLINE | ID: mdl-38500452

ABSTRACT

The structural cuticle proteins (CPs) play important roles in the development and fitness of insects. However, knowledge about CP gene superfamily is limited in virus-transmitting insect vectors, although its importance on transmission of plant virus has been gradually emphasized. In this study, the genome-wide identification of CP superfamily was conducted in western flower thrips Frankliniella occidentalis that is the globally invasive pest and plant virus vector pest. The pest transmits notorious tomato spotted wilt virus (TSWV) around the world, causing large damage to a wide array of plants. One hundred and twenty-eight F. occidentalis CP genes (FoCPs) were annotated in this study and they were classified into 10 distinct families, including 68 CPRs, 16 CPAP1s, 6 CPAP3s, 2 CPCFCs, 10 Tweedles, 4 CPFs, 16 CPLCPs, and 6 CPGs. The comprehensive analysis was performed including phylogenetic relationship, gene location and gene expression profiles during different development stages of F. occidentalis. Transcriptome analysis revealed more than 30% FoCPs were upregulated at least 1.5-fold when F. occidentalis was infected by TSWV, indicating their potential involvement in TSWV interactions. Our study provided an overview of F. occidentalis CP superfamily. The study gave a better understand of CP's role in development and virus transmission, which provided clues for reducing viral damages through silencing CP genes in insect vectors.


Subject(s)
Thysanoptera , Tospovirus , Animals , Insect Vectors/genetics , Insecta , Phylogeny , Thysanoptera/genetics , Tospovirus/genetics
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