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Molecular Plant Pathology Aug 2023The tomato Tm-2 gene was considered to be one of the most durable resistance genes in agriculture, protecting against viruses of the Tobamovirus genus, such as tomato...
The tomato Tm-2 gene was considered to be one of the most durable resistance genes in agriculture, protecting against viruses of the Tobamovirus genus, such as tomato mosaic virus (ToMV) and tobacco mosaic virus (TMV). However, an emerging tobamovirus, tomato brown rugose fruit virus (ToBRFV), has overcome Tm-2 , damaging tomato production worldwide. Tm-2 encodes a nucleotide-binding leucine-rich repeat (NLR) class immune receptor that recognizes its effector, the tobamovirus movement protein (MP). Previously, we found that ToBRFV MP (MP ) enabled the virus to overcome Tm-2 -mediated resistance. Yet, it was unknown how Tm-2 remained durable against other tobamoviruses, such as TMV and ToMV, for over 60 years. Here, we show that a conserved cysteine (C68) in the MP of TMV (MP ) plays a dual role in Tm-2 activation and viral movement. Substitution of MP amino acid H67 with the corresponding amino acid in MP (C68) activated Tm-2 -mediated resistance. However, replacement of C68 in TMV and ToMV disabled the infectivity of both viruses. Phylogenetic and structural prediction analysis revealed that C68 is conserved among all Solanaceae-infecting tobamoviruses except ToBRFV and localizes to a predicted jelly-roll fold common to various MPs. Cell-to-cell and subcellular movement analysis showed that C68 is required for the movement of TMV by regulating the MP interaction with the endoplasmic reticulum and targeting it to plasmodesmata. The dual role of C68 in viral movement and Tm-2 immune activation could explain how TMV was unable to overcome this resistance for such a long period.
Topics: Tobacco Mosaic Virus; Cysteine; Phylogeny; Tobamovirus; Nicotiana; Plant Viral Movement Proteins
PubMed: 37086003
DOI: 10.1111/mpp.13318 -
Frontiers in Plant Science 2023Asparagus samples were examined from growing areas of Germany and selected European as well as North, Central and South American countries. Overall, 474 samples were...
Asparagus samples were examined from growing areas of Germany and selected European as well as North, Central and South American countries. Overall, 474 samples were analyzed for Asparagus virus 1 (AV1) using DAS-ELISA. In our survey, 19 AV1 isolates were further characterized. Experimental transmission to 11 species belonging to Aizoaceae, Amarantaceae, Asparagaceae, and Solanaceae succeeded. The ultrastructure of AV1 infection in asparagus has been revealed and has been compared with the one in indicator plants. The cylindrical inclusion (CI) protein, a core factor in viral replication, localized within the cytoplasm and in systemic infections adjacent to the plasmodesmata. The majority of isolates referred to pathotype I (PI). These triggered a hypersensitive resistance in inoculated leaves of spp. and were incapable of infecting spp. Only pathotype II (PII) and pathotype III (PIII) infected systemically but differed in their virulence when transmitted to spp. The newly identified PIII generated amorphous inclusion bodies and degraded chloroplasts during systemic infection but not in local lesions of infected spp. PIII probably evolved recombination in asparagus carrying a mixed infection by PI and PII. Phylogeny of the coat protein region recognized two clusters, which did not overlap with the CI-associated grouping of pathotypes. These results provide evidence for ongoing modular evolution of AV1.
PubMed: 37600206
DOI: 10.3389/fpls.2023.1187563 -
BioRxiv : the Preprint Server For... Oct 2023Callose, a beta-(1,3)-D-glucan polymer, is essential for regulating intercellular trafficking via plasmodesmata (PD). Pathogens manipulate PD-localized proteins to...
Callose, a beta-(1,3)-D-glucan polymer, is essential for regulating intercellular trafficking via plasmodesmata (PD). Pathogens manipulate PD-localized proteins to enable intercellular trafficking by removing callose at PD, or conversely by increasing callose accumulation at PD to limit intercellular trafficking during infection. Plant defense hormones like salicylic acid regulate PD-localized proteins to control PD and intercellular trafficking during innate immune defense responses such as systemic acquired resistance. Measuring callose deposition at PD in plants has therefore emerged as a popular parameter for assessing the intercellular trafficking activity during plant immunity. Despite the popularity of this metric there is no standard for how these measurements should be made. In this study, three commonly used methods for identifying and quantifying PD callose by aniline blue staining were evaluated to determine the most effective in the leaf model. The results reveal that the most reliable method used aniline blue staining and fluorescent microscopy to measure callose deposition in fixed tissue. Manual or semi-automated workflows for image analysis were also compared and found to produce similar results although the semi-automated workflow produced a wider distribution of data points.
PubMed: 37873219
DOI: 10.1101/2023.09.30.560305 -
International Journal of Molecular... Sep 2023To move from cell to cell through plasmodesmata, many plant viruses require the concerted action of two or more movement proteins (MPs) encoded by transport gene modules...
To move from cell to cell through plasmodesmata, many plant viruses require the concerted action of two or more movement proteins (MPs) encoded by transport gene modules of virus genomes. A tetra-cistron movement block (TCMB) is a newly discovered transport module comprising four genes. TCMB encodes three proteins, which are similar to MPs of the transport module known as the "triple gene block", and a protein unrelated to known viral MPs and containing a double-stranded RNA (dsRNA)-binding domain similar to that found in a family of cell proteins, including AtDRB4 and AtHYL1. Here, the latter TCMB protein, named vDRB for virus dsRNA-binding protein, is shown to bind both dsRNA and single-stranded RNA in vitro. In a turnip crinkle virus-based assay, vDRB exhibits the properties of a viral suppressor of RNA silencing (VSR). In the context of potato virus X infection, vDRB significantly decreases the number and size of "dark green islands", regions of local antiviral silencing, supporting the VSR function of vDRB. Nevertheless, vDRB does not exhibit the VSR properties in non-viral transient expression assays. Taken together, the data presented here indicate that vDRB is an RNA-binding protein exhibiting VSR functions in the context of viral infection.
PubMed: 37762447
DOI: 10.3390/ijms241814144 -
Plant Cell Reports Dec 2023The leaf hyponasty response depends on tip-to-petiole auxin transport. This transport can happen through two parallel pathways: active trans-membrane transport mediated...
The leaf hyponasty response depends on tip-to-petiole auxin transport. This transport can happen through two parallel pathways: active trans-membrane transport mediated by PIN proteins and passive diffusion through plasmodesmata. A plant's ability to counteract potential shading by neighboring plants depends on transport of the hormone auxin. Neighbor sensing at the leaf tip triggers auxin production. Once this auxin reaches the abaxial petiole epidermis, it causes cell elongation, which leads to leaf hyponasty. Two pathways are known to contribute to this intercellular tip-to-petiole auxin movement: (i) transport facilitated by plasma membrane-localized PIN auxin transporters and (ii) diffusion enabled by plasmodesmata. We tested if these two modes of transport are arranged sequentially or in parallel. Moreover, we investigated if they are functionally linked. Mutants in which one of the two pathways is disrupted indicated that both pathways are necessary for a full hyponasty response. Visualization of PIN3-GFP and PIN7-GFP localization indicated PIN-mediated transport in parallel to plasmodesmata-mediated transport along abaxial midrib epidermis cells. We found plasmodesmata-mediated cell coupling in the pin3pin4pin7 mutant to match wild-type levels, indicating no redundancy between pathways. Similarly, PIN3, PIN4 and PIN7 mRNA levels were unaffected in a mutant with disrupted plasmodesmata pathway. Our results provide mechanistic insight on leaf hyponasty, which might facilitate the manipulation of the shade avoidance response in crops.
Topics: Arabidopsis; Plasmodesmata; Biological Transport; Membrane Transport Proteins; Indoleacetic Acids
PubMed: 38117314
DOI: 10.1007/s00299-023-03119-1 -
Plant Biotechnology Journal May 2024Viral diseases seriously threaten rice production. Plasmodesmata (PD)-associated proteins are deemed to play a key role in viral infection in host plants. However, few...
Viral diseases seriously threaten rice production. Plasmodesmata (PD)-associated proteins are deemed to play a key role in viral infection in host plants. However, few PD-associated proteins have been discovered in rice to afford viral infection. Here, inspired by the infection mechanism in insect vectors, we identified a member of the Flotillin family taking part in the cell-to-cell transport of rice stripe virus (RSV) in rice. Flotillin1 interacted with RSV nucleocapsid protein (NP) and was localized on PD. In flotillin1 knockout mutant rice, which displayed normal growth, RSV intercellular movement was retarded, leading to significantly decreased disease incidence. The PD pore sizes of the mutant rice were smaller than those of the wild type due to more callose deposits, which was closely related to the upregulation of two callose synthase genes. RSV infection stimulated flotillin1 expression and enlarged the PD aperture via RSV NP. In addition, flotillin1 knockout decreased disease incidences of southern rice black-streaked dwarf virus (SRBSDV) and rice dwarf virus (RDV) in rice. Overall, our study reveals a new PD-associated protein facilitating virus cell-to-cell trafficking and presents the potential of flotillin1 as a target to produce broad-spectrum antiviral rice resources in the future.
Topics: Animals; Plasmodesmata; Viral Proteins; Virus Diseases; Oryza; Plant Diseases; Hemiptera; Membrane Proteins
PubMed: 38130080
DOI: 10.1111/pbi.14274 -
Molecular Plant Jun 2024Circular single-stranded DNA (ssDNA) viruses have been rarely found in fungi, and the evolutionary and ecological relationships among ssDNA viruses infecting fungi and...
Circular single-stranded DNA (ssDNA) viruses have been rarely found in fungi, and the evolutionary and ecological relationships among ssDNA viruses infecting fungi and other organisms remain unclear. In this study, a novel circular ssDNA virus, tentatively named Diaporthe sojae circular DNA virus 1 (DsCDV1), was identified in the phytopathogenic fungus Diaporthe sojae isolated from pear trees. DsCDV1 has a monopartite genome (3185 nt in size) encapsidated in isometric virions (21-26 nm in diameter). The genome comprises seven putative open reading frames encoding a discrete replicase (Rep) split by an intergenic region, a putative capsid protein (CP), several proteins of unknown function (P1-P4), and a long intergenic region. Notably, the two split parts of DsCDV1 Rep share high identities with the Reps of Geminiviridae and Genomoviridae, respectively, indicating an evolutionary linkage with both families. Phylogenetic analysis based on Rep or CP sequences placed DsCDV1 in a unique cluster, supporting the establishment of a new family, tentatively named Gegemycoviridae, intermediate to both families. DsCDV1 significantly attenuates fungal growth and nearly erases fungal virulence when transfected into the host fungus. Remarkably, DsCDV1 can systematically infect tobacco and pear seedlings, providing broad-spectrum resistance to fungal diseases. Subcellular localization analysis revealed that DsCDV1 P3 is systematically localized in the plasmodesmata, while its expression in trans-complementation experiments could restore systematic infection of a movement-deficient plant virus, suggesting that P3 is a movement protein. DsCDV1 exhibits unique molecular and biological traits not observed in other ssDNA viruses, serving as a link between fungal and plant ssDNA viruses and presenting an evolutionary connection between ssDNA viruses and fungi. These findings contribute to expanding our understanding of ssDNA virus diversity and evolution, offering potential biocontrol applications for managing crucial plant diseases.
Topics: Fungal Viruses; Plant Diseases; Phylogeny; DNA, Single-Stranded; Ascomycota; DNA Viruses; Disease Resistance; Genome, Viral; Pyrus; Nicotiana
PubMed: 38745413
DOI: 10.1016/j.molp.2024.05.003 -
Science Bulletin Apr 2024The microdomains of plasmodesmata, specialized cell-wall channels responsible for communications between neighboring cells, are composed of various plasmodesmata-located...
The microdomains of plasmodesmata, specialized cell-wall channels responsible for communications between neighboring cells, are composed of various plasmodesmata-located proteins (PDLPs) and lipids. Here, we found that, among all PDLP or homologous proteins in Arabidopsis thaliana genome, PDLP5 and PDLP7 possessed a C-terminal sphingolipid-binding motif, with the latter being the only member that was significantly upregulated upon turnip mosaic virus and cucumber mosaic virus infections. pdlp7 mutant plants exhibited significantly reduced callose deposition, larger plasmodesmata diameters, and faster viral transmission. These plants exhibited increased glucosidase activity but no change in callose synthase activity. PDLP7 interacted specifically with glucan endo-1,3-β-glucosidase 10 (BG10). Consistently, higher levels of callose deposition and slower virus transmission in bg10 mutants were observed. The interaction between PDLP7 and BG10 was found to depend on the presence of the Gnk2-homologous 1 (GnK2-1) domain at the N terminus of PDLP7 with Asp-35, Cys-42, Gln-44, and Leu-116 being essential. In vitro supplementation of callose was able to change the conformation of the GnK2-1 domain. Our data suggest that the GnK2-1 domain of PDLP7, in conjunction with callose and BG10, plays a key role in plasmodesmata opening and closure, which is necessary for intercellular movement of various molecules.
PubMed: 38735789
DOI: 10.1016/j.scib.2024.04.063 -
Molecular Plant-microbe Interactions :... Feb 2024In plants, plasmodesmata establish cytoplasmic continuity between cells to allow for communication and resource exchange across the cell wall. While plant pathogens use...
In plants, plasmodesmata establish cytoplasmic continuity between cells to allow for communication and resource exchange across the cell wall. While plant pathogens use plasmodesmata as a pathway for both molecular and physical invasion, the benefits of molecular invasion (cell-to-cell movement of pathogen effectors) are poorly understood. To establish a methodology for identification and characterization of the cell-to-cell mobility of effectors, we performed a quantitative live imaging-based screen of candidate effectors of the fungal pathogen . We predicted effectors by their expression profiles, the presence of a secretion signal, and their predicted and in planta localization when fused to green fluorescent protein. We assayed for cell-to-cell mobility of nucleocytosolic effectors and identified 14 that are cell-to-cell mobile. We identified that three of these effectors are "hypermobile," showing cell-to-cell mobility greater than expected for a protein of that size. To explore the mechanism of hypermobility, we chose two hypermobile effectors and measured their impact on plasmodesmata function and found that even though they show no direct association with plasmodesmata, each increases the transport capacity of plasmodesmata. Thus, our methods for quantitative analysis of cell-to-cell mobility of candidate microbe-derived effectors, or any suite of host proteins, can identify cell-to-cell hypermobility and offer greater understanding of how proteins affect plasmodesmal function and intercellular connectivity. [Formula: see text] Copyright © 2024 The Author(s). This is an open access article distributed under the CC BY 4.0 International license.
Topics: Plasmodesmata; Plants; Cytoplasm; Cytosol; Cell Wall
PubMed: 37942798
DOI: 10.1094/MPMI-05-23-0052-TA -
Molecular Plant Pathology Jun 2024Fusarium head blight disease on small-grain cereals is primarily caused by the ascomycete fungal pathogen Fusarium graminearum. Infection of floral spike tissues is...
Fusarium head blight disease on small-grain cereals is primarily caused by the ascomycete fungal pathogen Fusarium graminearum. Infection of floral spike tissues is characterized by the biosynthesis and secretion of potent trichothecene mycotoxins, of which deoxynivalenol (DON) is widely reported due to its negative impacts on grain quality and consumer safety. The TRI5 gene encodes an essential enzyme in the DON biosynthesis pathway and the single gene deletion mutant, ΔTri5, is widely reported to restrict disease progression to the inoculated spikelet. In this study, we present novel bioimaging evidence revealing that DON facilitates the traversal of the cell wall through plasmodesmata, a process essential for successful colonization of host tissue. Chemical complementation of ΔTri5 did not restore macro- or microscopic phenotypes, indicating that DON secretion is tightly regulated both spatially and temporally. A comparative qualitative and quantitative morphological cellular analysis revealed infections had no impact on plant cell wall thickness. Immunolabelling of callose at plasmodesmata during infection indicates that DON can increase deposits when applied exogenously but is reduced when F. graminearum hyphae are present. This study highlights the complexity of the interconnected roles of mycotoxin production, cell wall architecture and plasmodesmata in this highly specialized interaction.
Topics: Trichothecenes; Fusarium; Triticum; Plant Diseases; Cell Wall; Plasmodesmata; Mycotoxins
PubMed: 38877764
DOI: 10.1111/mpp.13485