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PLoS Pathogens Dec 2022Tobacco mosaic virus movement protein (TMV MP) is essential for virus spread between cells. To accomplish its task, TMV MP binds viral RNA, interacts with components of...
Tobacco mosaic virus movement protein (TMV MP) is essential for virus spread between cells. To accomplish its task, TMV MP binds viral RNA, interacts with components of the cytoskeleton, and increases the size exclusion limit (SEL) of plasmodesmata. Plasmodesmata are gated intercellular channels that allow passage of small molecules and macromolecules, including RNA and protein, between plant cells. Moreover, plasmodesmata are diverse and those connecting different cell types appear to have unique mechanisms to regulate macromolecular trafficking, which likely contributes to the establishment of distinct cell boundaries. Consequently, TMV MP might be competent to mediate RNA transport through some but not all plasmodesmal gates. Due to a lack of viral mutants defective for movement between specific cell types, the ability of TMV MP in this regard is incompletely understood. In contrast, a number of trafficking impaired Potato spindle tuber viroid (PSTVd) mutants have been identified. PSTVd is a systemically infectious non-coding RNA that nevertheless can perform all functions required for replication as well as cell-to-cell and systemic spread. Previous studies have shown that PSTVd employs different structure and sequence elements to move between diverse cell types in host plants, and mutants defective for transport between specific cell types have been identified. Therefore, PSTVd may serve as a tool to analyze the functions of MPs of viral and cellular origin. To probe the RNA transport activity of TMV MP, transgenic plants expressing the protein were inoculated with PSTVd mutants. Remarkably, TMV MP complemented a PSTVd mutant defective for mesophyll entry but could not support two mutants impaired for phloem entry, suggesting it fails to productively interface with plasmodesmata at the phloem boundary and that additional viral and host factors may be required. Consistent with this idea, TMV co-infection, but not the combination of MP and coat protein (CP) expression, was able to complement one of the phloem entry mutants. These observations suggest that phloem loading is a critical impediment to establishing systemic infection that could involve the entire ensemble of TMV proteins. They also demonstrate a novel strategy for analysis of MPs.
Topics: Tobacco Mosaic Virus; Viroids; Solanum tuberosum; Phloem; RNA, Viral; Plant Viral Movement Proteins; Nicotiana
PubMed: 36574436
DOI: 10.1371/journal.ppat.1011062 -
The New Phytologist Mar 2023Fruit malformation is a major constrain in fruit production worldwide resulting in substantial economic losses. The farmers for decades noticed that the chilling...
Fruit malformation is a major constrain in fruit production worldwide resulting in substantial economic losses. The farmers for decades noticed that the chilling temperature before blooming often caused malformed fruits. However, the molecular mechanism underlying this phenomenon is unclear. Here we examined the fruit development in response to cold stress in tomato, and demonstrated that short-term cold stress increased the callose accumulation in both shoot apical and floral meristems, resulting in the symplastic isolation and altered intercellular movement of WUS. In contrast to the rapidly restored SlWUS transcription during the recovery from cold stress, the callose removal was delayed due to obstructed plasmodesmata. The delayed reinstatement of cell-to-cell transport of SlWUS prevented the activation of SlCLV3 and TAG1, causing the interrupted feedback inhibition of SlWUS expression, leading to the expanded stem cell population and malformed fruits. We further showed that the callose dynamics in response to short-term cold stress presumably exploits the mechanism of bud dormancy during the seasonal growth, involving two antagonistic hormones, abscisic acid and gibberellin. Our results provide a novel insight into the cold stress regulated malformation of fruit.
Topics: Cold-Shock Response; Fruit; Gene Expression Regulation, Plant; Meristem; Plant Proteins; Solanum lycopersicum; Stem Cells; Feedback, Physiological
PubMed: 36564973
DOI: 10.1111/nph.18699 -
Viruses Dec 2022Movement proteins (MPs) of plant viruses enable the translocation of viral genomes from infected to healthy cells through plasmodesmata (PD). The MPs functions involve...
Movement proteins (MPs) of plant viruses enable the translocation of viral genomes from infected to healthy cells through plasmodesmata (PD). The MPs functions involve the increase of the PD permeability and routing of viral genome both to the PD entrance and through the modified PD. encodes two MPs, termed BMB1 and BMB2, which act in concert to accomplish virus cell-to-cell transport. BMB1, representing an NTPase/helicase domain-containing RNA-binding protein, localizes to the cytoplasm and the nucleoplasm. BMB2 is a small hydrophobic protein that interacts with the endoplasmic reticulum (ER) membranes and induces local constrictions of the ER tubules. In plant cells, BMB2 localizes to PD-associated membrane bodies (PAMBs) consisting of modified ER tubules and directs BMB1 to PAMBs. Here, we demonstrate that BMB1 and BMB2 interact in vitro and in vivo, and that their specific interaction is essential for BMB2-directed targeting of BMB1 to PAMBs. Using mutagenesis, we show that the interaction involves the C-terminal BMB1 region and the N-terminal region of BMB2.
Topics: Hibiscus; Plant Viruses; Endoplasmic Reticulum; RNA Viruses; Plant Viral Movement Proteins; Nicotiana; Plasmodesmata
PubMed: 36560746
DOI: 10.3390/v14122742 -
Plant Biotechnology Journal Mar 2023Molecular farming technology using transiently transformed Nicotiana plants offers an economical approach to the pharmaceutical industry to produce an array of protein...
Molecular farming technology using transiently transformed Nicotiana plants offers an economical approach to the pharmaceutical industry to produce an array of protein targets including vaccine antigens and therapeutics. It can serve as a desirable alternative approach for those proteins that are challenging or too costly to produce in large quantities using other heterologous protein expression systems. However, since cost metrics are such a critical factor in selecting a production host, any system-wide modifications that can increase recombinant protein yields are key to further improving the platform and making it applicable for a wider range of target molecules. Here, we report on the development of a new approach to improve target accumulation in an established plant-based expression system that utilizes viral-based vectors to mediate transient expression in Nicotiana benthamiana. We show that by engineering the host plant to support viral vectors to spread more effectively between host cells through plasmodesmata, protein target accumulation can be increased by up to approximately 60%.
Topics: Recombinant Proteins; Plants, Genetically Modified; Tobacco Mosaic Virus; Nicotiana; Protein Transport; Genetic Vectors
PubMed: 36511837
DOI: 10.1111/pbi.13977 -
The Plant Journal : For Cell and... Feb 2023Arabinogalactan proteins (AGPs) are a plant-specific family of extracellular proteoglycans characterized by large and complex galactose-rich polysaccharide chains....
Arabinogalactan proteins (AGPs) are a plant-specific family of extracellular proteoglycans characterized by large and complex galactose-rich polysaccharide chains. Functional elucidation of AGPs, however, has been hindered by the high degree of redundancy of AGP genes. To uncover as yet unexplored roles of AGPs in Arabidopsis, a mutant of Hyp O-galactosyltransferase (HPGT), a critical enzyme that catalyzes the common initial step of Hyp-linked arabinogalactan chain biosynthesis, was used. Here we show, using the hpgt1,2,3 triple mutant, that a reduction in functional AGPs leads to a stomatal patterning defect in which two or more stomata are clustered together. This defect is attributed to increased and dysregulated symplastic transport following changes in plasmodesmata structure, such that highly permeable complex branched plasmodesmata with cavities in branching parts increased in the mutant. We also found that the hpgt1,2,3 mutation causes a reduction of cellulose in the cell wall and accumulation of pectin, which controls cell wall porosity. Our results highlight the importance of AGPs in the correct biogenesis of plasmodesmata, possibly acting through the regulation of cell wall properties surrounding the plasmodesmata.
Topics: Plasmodesmata; Plant Proteins; Arabidopsis; Mucoproteins; Cell Wall
PubMed: 36511822
DOI: 10.1111/tpj.16061 -
Plant, Cell & Environment Feb 2023Cell walls are essential for plant growth and development, providing support and protection from external environments. Callose is a glucan that accumulates in... (Review)
Review
Cell walls are essential for plant growth and development, providing support and protection from external environments. Callose is a glucan that accumulates in specialized cell wall microdomains including around intercellular pores called plasmodesmata. Despite representing a small percentage of the cell wall (~0.3% in the model plant Arabidopsis thaliana), callose accumulation regulates important biological processes such as phloem and pollen development, cell division, organ formation, responses to pathogenic invasion and to changes in nutrients and toxic metals in the soil. Callose accumulation modifies cell wall properties and restricts plasmodesmata aperture, affecting the transport of signaling proteins and RNA molecules that regulate plant developmental and environmental responses. Although the importance of callose, at and outside plasmodesmata cell walls, is widely recognized, the underlying mechanisms controlling changes in its synthesis and degradation are still unresolved. In this review, we explore the most recent literature addressing callose metabolism with a focus on the molecular factors affecting callose accumulation in response to mutualistic symbionts and pathogenic elicitors. We discuss commonalities in the signaling pathways, identify research gaps and highlight opportunities to target callose in the improvement of plant responses to beneficial versus pathogenic microbes.
Topics: Plasmodesmata; Plants; Arabidopsis; Glucans; Cell Wall
PubMed: 36478232
DOI: 10.1111/pce.14510 -
Biophysical Journal Jan 2023Intercellular communication and transport is the essential prerequisite for the function of multicellular organisms. Simple diffusion as a transport mechanism is often...
Intercellular communication and transport is the essential prerequisite for the function of multicellular organisms. Simple diffusion as a transport mechanism is often inefficient in sustaining the effective exchange of metabolites, and other active transport mechanisms become involved. In this paper, we use the giant cells of characean algae as a model system to explore the role of advection and diffusion in intercellular transport. Using fluorescent dye as a tracer, we study the kinetics of the permeation of the fluorophore through the plasmodesmata complex in the node of tandem cells and its further distribution across the cell. To explore the role of cytoplasmic streaming and the nodal cell complex in the transport mechanism, we modulate the cytoplasmic streaming using action potential to separate the diffusive permeation from the advective contribution. The results imply that the plasmodesmal transport of fluorescent probe through the central and peripheral cells of the nodal complex is differentially regulated by a physiological signal, the action potential. The passage of the probe through the central cells of the nodal complex ceases transiently after elicitation of the action potential in the internodal cell, whereas the passage through the peripheral cells of the node was retained. A diffusion-advection model is developed to describe the transport kinetics and extract the permeability of the node-internode cell wall from experimental data.
Topics: Characeae; Fluorescent Dyes; Chara; Biological Transport; Cytoplasmic Streaming
PubMed: 36463405
DOI: 10.1016/j.bpj.2022.11.2948 -
Plant Physiology Feb 2023Intracellular movement is an important step for the initial spread of virus in plants during infection. This process requires virus-encoded movement proteins (MPs) and...
Intracellular movement is an important step for the initial spread of virus in plants during infection. This process requires virus-encoded movement proteins (MPs) and their interaction with host factors. Despite the large number of known host factors involved in the movement of different viruses, little is known about host proteins that interact with one of the MPs encoded by potexviruses, the triple-gene-block protein 3 (TGBp3). The main obstacle lies in the relatively low expression level of potexviral TGBp3 in hosts and the weak or transient nature of interactions. Here, we used TurboID-based proximity labeling to identify the network of proteins directly or indirectly interacting with the TGBp3 of a potexvirus, Bamboo mosaic virus (BaMV). Endoplasmic reticulum (ER) luminal-binding protein 4 and calreticulin 3 of Nicotiana benthamiana (NbBiP4 and NbCRT3, respectively) associated with the functional TGBp3-containing BaMV movement complexes, but not the movement-defective mutant, TGBp3M. Fluorescent microscopy revealed that TGBp3 colocalizes with NbBiP4 or NbCRT3 and the complexes move together along ER networks to cell periphery in N. benthamiana. Loss- and gain-of-function experiments revealed that NbBiP4 or NbCRT3 is required for the efficient spread and accumulation of BaMV in infected leaves. In addition, overexpression of NbBiP4 or NbCRT3 enhanced the targeting of BaMV TGBp1 to plasmodesmata (PD), indicating that NbBiP4 and NbCRT3 interact with TGBp3 to promote the intracellular transport of virion cargo to PD that facilitates virus cell-to-cell movement. Our findings revealed additional roles for NbBiP4 and NbCRT3 in BaMV intracellular movement through ER networks or ER-derived vesicles to PD, which enhances the spread of BaMV in N. benthamiana.
Topics: Viral Proteins; Carrier Proteins; Calreticulin; Plants; Nicotiana; Endoplasmic Reticulum; Potexvirus
PubMed: 36459587
DOI: 10.1093/plphys/kiac547 -
The Plant Genome Dec 2023As conserved regulatory agents, noncoding RNAs (ncRNAs) have an important impact on many aspects of plant life, including growth, development, and environmental... (Review)
Review
As conserved regulatory agents, noncoding RNAs (ncRNAs) have an important impact on many aspects of plant life, including growth, development, and environmental response. Noncoding RNAs can travel through not only plasmodesma and phloem but also intercellular barriers to regulate distinct processes. Increasing evidence shows that the intercellular trans-kingdom transmission of ncRNAs is able to modulate many important interactions between plants and other organisms, such as plant response to pathogen attack, the symbiosis between legume plants and rhizobia and the interactions with parasitic plants. In these interactions, plant ncRNAs are believed to be sorted into extracellular vesicles (EVs) or other nonvesicular vehicles to pass through cell barriers and trigger trans-kingdom RNA interference (RNAi) in recipient cells from different species. There is evidence that the features of extracellular RNAs and associated RNA-binding proteins (RBPs) play a role in defining the RNAs to retain in cell or secrete outside cells. Despite the few reports about RNA secretion pathway in plants, the export of extracellular ncRNAs is orchestrated by a series of pathways in plants. The identification and functional analysis of mobile small RNAs (sRNAs) are attracting increasing attention in recent years. In this review, we discuss recent advances in our understanding of the function, sorting, transport, and regulation of plant extracellular ncRNAs.
Topics: Gene-Environment Interaction; RNA Interference; RNA, Untranslated; RNA, Plant; Plants
PubMed: 36444889
DOI: 10.1002/tpg2.20289 -
Frontiers in Plant Science 2022Plant viruses use a variety of strategies to infect their host. During infection, viruses cause symptoms of varying severity, which are often associated with altered...
Plant viruses use a variety of strategies to infect their host. During infection, viruses cause symptoms of varying severity, which are often associated with altered leaf pigmentation due to structural and functional damage to chloroplasts that are affected by viral proteins. Here we demonstrate that Kunitz peptidase inhibitor-like protein (KPILP) gene is induced in response to potato virus X (PVX) infection. Using reverse genetic approach, we have demonstrated that downregulates expression of and genes of antenna light-harvesting complex proteins, gene encoding glutamyl-tRNA reductase, which participates in tetrapyrrole biosynthesis, and gene encoding RuBisCO small subunit isoform involved in the antiviral immune response. Thus, is a regulator of chloroplast retrograde signaling system during developing PVX infection. Moreover, was demonstrated to affect carbon partitioning: reduced glucose levels during PVX infection were associated with upregulation. Another KPILP function is associated with plasmodesmata permeability control. Its ability to stimulate intercellular transport of reporter 2xGFP molecules indicates that KPILP is a positive plasmodesmata regulator. Moreover, natural KPILP glycosylation is indispensable for manifestation of this function. During PVX infection increased expression leads to the reduction of plasmodesmata callose deposition. These results could indicate that KPILP affects plasmodesmata permeability callose-dependent mechanism. Thus, virus entering a cell and starting reproduction triggers expression, which leads to downregulation of nuclear-encoded chloroplast genes associated with retrograde signaling, reduction in photoassimilates accumulation and increase in intercellular transport, creating favorable conditions for reproduction and spread of viral infection.
PubMed: 36438111
DOI: 10.3389/fpls.2022.1041867