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1.
BMC Plant Biol ; 21(1): 318, 2021 Jul 03.
Artigo em Inglês | MEDLINE | ID: mdl-34217217

RESUMO

BACKGROUND: Cassava (Manihot esculenta Crantz) efficiently accumulates starch in its storage roots. However, how photosynthates are transported from the leaves to the phloem (especially how they are unloaded into parenchymal cells of storage roots) remains unclear. RESULTS: Here, we investigated the sucrose unloading pattern and its impact on cassava storage root development using microstructural and physiological analyses, namely, carboxyfluorescein (CF) and C14 isotope tracing. The expression profiling of genes involved in symplastic and apoplastic transport was performed, which included enzyme activity, protein gel blot analysis, and transcriptome sequencing analyses. These finding showed that carbohydrates are transported mainly in the form of sucrose, and more than 54.6% was present in the stem phloem. Sucrose was predominantly unloaded symplastically from the phloem into storage roots; in addition, there was a shift from apoplastic to symplastic unloading accompanied by the onset of root swelling. Statistical data on the microstructures indicated an enrichment of plasmodesmata within sieve, companion, and parenchyma cells in the developing storage roots of a cultivar but not in a wild ancestor. Tracing tests with CF verified the existence of a symplastic channel, and [14C] Suc demonstrated that sucrose could rapidly diffuse into root parenchyma cells from phloem cells. The relatively high expression of genes encoding sucrose synthase and associated proteins appeared in the middle and late stages of storage roots but not in primary fibrous roots, or secondary fibrous roots. The inverse expression pattern of sucrose transporters, cell wall acid invertase, and soluble acid invertase in these corresponding organs supported the presence of a symplastic sucrose unloading pathway. The transcription profile of genes involved in symplastic unloading and their significantly positive correlation with the starch yield at the population level confirmed that symplastic sucrose transport is vitally important in the development of cassava storage roots. CONCLUSIONS: In this study, we revealed that the cassava storage root phloem sucrose unloading pattern was predominantly a symplastic unloading pattern. This pattern is essential for efficient starch accumulation in high-yielding varieties compared with low-yielding wild ancestors.


Assuntos
Manihot/metabolismo , Floema/fisiologia , Fotossíntese/fisiologia , Raízes de Plantas/metabolismo , Amido/metabolismo , Transporte Biológico , Biomassa , Parede Celular/metabolismo , Difusão , Fluoresceínas/metabolismo , Regulação da Expressão Gênica de Plantas , Manihot/genética , Modelos Biológicos , Floema/citologia , Floema/ultraestrutura , Plasmodesmos/metabolismo , Frações Subcelulares/metabolismo , Sacarose/metabolismo , Açúcares/metabolismo
2.
Plant J ; 107(1): 7-20, 2021 07.
Artigo em Inglês | MEDLINE | ID: mdl-34058040

RESUMO

The sensing of abiotic stress, mechanical injury or pathogen attack by a single plant tissue results in the activation of systemic signals that travel from the affected tissue to the entire plant. This process is essential for plant survival during stress and is termed systemic signaling. Among the different signals triggered during this process are calcium, electric, reactive oxygen species and hydraulic signals. These are thought to propagate at rapid rates through the plant vascular bundles and to regulate many of the systemic processes essential for plant survival. Although the different signals activated during systemic signaling are thought to be interlinked, their coordination and hierarchy still need to be determined. Here, using a combination of advanced whole-plant imaging and hydraulic pressure measurements, we studied the activation of all four systemic signals in wild-type and different Arabidopsis thaliana mutants subjected to a local treatment of high-light (HL) stress or wounding. Our findings reveal that activation of systemic membrane potential, calcium, reactive oxygen species and hydraulic pressure signals, in response to wounding, is dependent on glutamate receptor-like proteins 3.3 and 3.6. In contrast, in response to HL stress, systemic changes in calcium and membrane potential depended on glutamate receptor-like 3.3 and 3.6, while systemic hydraulic signals did not. We further show that plasmodesmata functions are required for systemic changes in membrane potential and calcium during responses to HL stress or wounding. Our findings shed new light on the different mechanisms that integrate different systemic signals in plants during stress.


Assuntos
Arabidopsis/metabolismo , Sinalização do Cálcio/fisiologia , Espécies Reativas de Oxigênio/metabolismo , Arabidopsis/genética , Arabidopsis/fisiologia , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Cálcio/metabolismo , Potenciais da Membrana , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Mutação , NADPH Oxidases/genética , NADPH Oxidases/metabolismo , Plasmodesmos/metabolismo , Receptores de Glutamato/genética , Receptores de Glutamato/metabolismo , Transdução de Sinais , Estresse Fisiológico
3.
J Exp Bot ; 72(15): 5534-5552, 2021 07 28.
Artigo em Inglês | MEDLINE | ID: mdl-33974689

RESUMO

In mature leaves, cell-to-cell transport via plasmodesmata between mesophyll cells links the production of assimilates by photosynthesis with their export to sink organs. This study addresses the question of how signals derived from chloroplasts and photosynthesis influence plasmodesmata permeability. Cell-to-cell transport was analyzed in leaves of the Arabidopsis chlorophyll b-less ch1-3 mutant, the same mutant complemented with a cyanobacterial CAO gene (PhCAO) overaccumulating chlorophyll b, the trxm3 mutant lacking plastidial thioredoxin m3, and the ntrc mutant lacking functional NADPH:thioredoxin reductase C. The regulation of plasmodesmata permeability in these lines could not be traced back to the reduction state of the thioredoxin system or the types and levels of reactive oxygen species produced in chloroplasts; however, it could be related to chloroplast ATP and NADPH production. The results suggest that light enables plasmodesmata closure via an increase in the ATP and NADPH levels produced in photosynthesis, providing a control mechanism for assimilate export based on the rate of photosynthate production in the Calvin-Benson cycle. The level of chlorophyll b influences plasmodesmata permeability via as-yet-unidentified signals. The data also suggest a role of thioredoxin m3 in the regulation of cyclic electron flow around photosystem I.


Assuntos
Proteínas de Arabidopsis , Arabidopsis , Trifosfato de Adenosina/metabolismo , Arabidopsis/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Clorofila/metabolismo , Cloroplastos/metabolismo , NADP/metabolismo , Oxirredução , Fotossíntese , Folhas de Planta/metabolismo , Plasmodesmos/metabolismo
4.
Cells ; 10(4)2021 03 26.
Artigo em Inglês | MEDLINE | ID: mdl-33810252

RESUMO

Plasmodesmata form intercellular channels which ensure the transport of various molecules during embryogenesis and postembryonic growth. However, high permeability of plasmodesmata may interfere with the establishment of auxin maxima, which are required for cellular patterning and the development of distinct tissues. Therefore, diffusion through plasmodesmata is not always desirable and the symplastic continuum must be broken up to induce or accomplish some developmental processes. Many data show the role of auxin maxima in the regulation of auxin-responsive genes and the establishment of various cellular patterns. However, still little is known whether and how these maxima are formed in the embryo proper before 16-cell stage, that is, when there is still a nonpolar distribution of auxin efflux carriers. In this work, we focused on auxin-dependent regulation of plasmodesmata function, which may provide rapid and transient changes of their permeability, and thus take part in the regulation of gene expression.


Assuntos
Genes de Plantas , Ácidos Indolacéticos/metabolismo , Plantas/embriologia , Plantas/genética , Plasmodesmos/metabolismo , Modelos Biológicos , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo
5.
Mol Plant Pathol ; 22(5): 539-550, 2021 05.
Artigo em Inglês | MEDLINE | ID: mdl-33723908

RESUMO

On infection, plant-parasitic nematodes establish feeding sites in roots from which they take up carbohydrates among other nutrients. Knowledge on how carbohydrates are supplied to the nematodes' feeding sites is limited. Here, gene expression analyses showed that RNA levels of OsSWEET11 to OsSWEET15 were extremely low in both Meloidogyne graminicola (Mg)-caused galls and noninoculated roots. All the rice sucrose transporter genes, OsSUT1 to OsSUT5, were either down-regulated in Mg-caused galls compared with noninoculated rice roots or had very low transcript abundance. OsSUT1 was the only gene up-regulated in galls, at 14 days postinoculation (dpi), after being highly down-regulated at 3 and 7 dpi. OsSUT4 was down-regulated at 3 dpi. No noticeable OsSUTs promoter activities were detected in Mg-caused galls of pOsSUT1 to -5::GUS rice lines. Loading experiments with carboxyfluorescein diacetate (CFDA) demonstrated that symplastic connections exist between phloem and Mg-caused giant cells (GCs). According to data from OsGNS5- and OsGSL2-overexpressing rice plants that had decreased and increased callose deposition, respectively, callose negatively affected Mg parasitism and sucrose supply to Mg-caused GCs. Our results suggest that plasmodesmata-mediated sucrose transport plays a pivotal role in sucrose supply from rice root phloem to Mg-caused GCs, and OsSWEET11 to -15 and OsSUTs are not major players in it, although further functional analysis is needed for OsSUT1 and OsSUT4.


Assuntos
Proteínas de Membrana Transportadoras/metabolismo , Oryza/metabolismo , Doenças das Plantas/parasitologia , Proteínas de Plantas/metabolismo , Plasmodesmos/metabolismo , Sacarose/metabolismo , Tylenchoidea/fisiologia , Animais , Transporte Biológico , Expressão Gênica , Genes Reporter , Glucanos/metabolismo , Proteínas de Membrana Transportadoras/genética , Oryza/parasitologia , Floema/metabolismo , Floema/parasitologia , Proteínas de Plantas/genética , Raízes de Plantas/metabolismo , Raízes de Plantas/parasitologia , Tumores de Planta/parasitologia
6.
Plant Sci ; 304: 110800, 2021 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-33568299

RESUMO

Cell-to-cell communication is crucial in coordinating diverse biological processes in multicellular organisms. In plants, communication between adjacent cells occurs via nanotubular passages called plasmodesmata (PD). The PD passage is composed of an appressed endoplasmic reticulum (ER) internally, and plasma membrane (PM) externally, that traverses the cell wall, and associates with the actin-cytoskeleton. The coordination of the ER, PM and cytoskeleton plays a potential role in maintaining the architecture and conductivity of PD. Many data suggest that PD-associated proteins can serve as tethers that connect these structures in a functional PD, to regulate cell-to-cell communication. In this review, we summarize the organization and regulation of PD activity via tethering proteins, and discuss the importance of PD-mediated cell-to-cell communication in plant development and defense against environmental stress.


Assuntos
Membrana Celular/fisiologia , Plasmodesmos/fisiologia , Actinas/metabolismo , Parede Celular/metabolismo , Retículo Endoplasmático/metabolismo , Proteínas de Membrana/metabolismo , Proteínas de Plantas/metabolismo , Plantas/metabolismo , Plasmodesmos/metabolismo
7.
J Plant Physiol ; 257: 153341, 2021 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-33388666

RESUMO

Plant tissues exhibit a symplasmic organization; the individual protoplasts are connected to their neighbors via cytoplasmic bridges that extend through pores in the cell walls. These bridges may have diameters of a micrometer or more, as in the sieve pores of the phloem, but in most cell types they are smaller. Historically, botanists referred to cytoplasmic bridges of all sizes as plasmodesmata. The meaning of the term began to shift when the transmission electron microscope (TEM) became the preferred tool for studying these structures. Today, a plasmodesma is widely understood to be a 'nano-scale' pore. Unfortunately, our understanding of these nanoscopic channels suffers from methodological limitations. This is exemplified by the fact that state-of-the-art EM techniques appear to reveal plasmodesmal pore structures that are much smaller than the tracer molecules known to diffuse through these pores. In general, transport processes in pores that have dimensions in the size range of the transported molecules are governed by different physical parameters than transport process in the macroscopic realm. This can lead to unexpected effects, as experience in nanofluidic technologies demonstrates. Our discussion of problems of size in plasmodesma research leads us to conclude that the field will benefit from technomimetic reasoning - the utilization of concepts developed in applied nanofluidics for the interpretation of biological systems.


Assuntos
Plasmodesmos/metabolismo , Transporte Biológico , Floema/metabolismo , Terminologia como Assunto
8.
Cell Mol Life Sci ; 78(3): 799-816, 2021 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-32920696

RESUMO

Plasmodesmata are intercellular pores connecting together most plant cells. These structures consist of a central constricted form of the endoplasmic reticulum, encircled by some cytoplasmic space, in turn delimited by the plasma membrane, itself ultimately surrounded by the cell wall. The presence and structure of plasmodesmata create multiple routes for intercellular trafficking of a large spectrum of molecules (encompassing RNAs, proteins, hormones and metabolites) and also enable local signalling events. Movement across plasmodesmata is finely controlled in order to balance processes requiring communication with those necessitating symplastic isolation. Here, we describe the identities and roles of the molecular components (specific sets of lipids, proteins and wall polysaccharides) that shape and define plasmodesmata structural and functional domains. We highlight the extensive and dynamic interactions that exist between the plasma/endoplasmic reticulum membranes, cytoplasm and cell wall domains, binding them together to effectively define plasmodesmata shapes and purposes.


Assuntos
Transporte Biológico/fisiologia , Plantas/metabolismo , Plasmodesmos/metabolismo , Comunicação Celular , Parede Celular/química , Parede Celular/metabolismo , Estruturas Citoplasmáticas/química , Retículo Endoplasmático/metabolismo , Lipídeos de Membrana/química , Lipídeos de Membrana/metabolismo , Proteínas de Plantas/química , Proteínas de Plantas/metabolismo , Plasmodesmos/química , Polissacarídeos/química , Polissacarídeos/metabolismo
9.
Plant Physiol ; 184(1): 53-64, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32719057

RESUMO

Plasmodesmata are small channels that connect plant cells. While recent technological advances have facilitated analysis of the ultrastructure of these channels, there are limitations to efficiently addressing their presence over an entire cellular interface. Here, we highlight the value of serial block electron microscopy for this purpose. We developed a computational pipeline to study plasmodesmata distributions and detect the presence/absence of plasmodesmata clusters, or pit fields, at the phloem unloading interfaces of Arabidopsis (Arabidopsis thaliana) roots. Pit fields were visualized and quantified. As the wall environment of plasmodesmata is highly specialized, we also designed a tool to extract the thickness of the extracellular matrix at and outside of plasmodesmata positions. We detected and quantified clear wall thinning around plasmodesmata with differences between genotypes, including the recently published plm-2 sphingolipid mutant. Our tools open avenues for quantitative approaches in the analysis of symplastic trafficking.


Assuntos
Arabidopsis/ultraestrutura , Microscopia Eletrônica/métodos , Plasmodesmos/ultraestrutura , Arabidopsis/genética , Arabidopsis/metabolismo , Genótipo , Floema/genética , Floema/metabolismo , Floema/ultraestrutura , Raízes de Plantas/genética , Raízes de Plantas/metabolismo , Raízes de Plantas/ultraestrutura , Plasmodesmos/metabolismo
10.
Plant Physiol ; 184(1): 407-420, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32636343

RESUMO

Plasma membranes encapsulated in the symplasmic nanochannels of plasmodesmata (PD) contain abundant lipid rafts, which are enriched with sphingolipids (SLs) and sterols. Reduction of sterols has highlighted the role played by lipid raft integrity in the intercellular trafficking of glycosylphosphatidylinositol (GPI)-anchored PD proteins, particularly in affecting callose enhancement. The presence of callose at PD is strongly attributed to the regulation of callose accumulation and callose degradation by callose synthases and ß-1,3-glucanases (BGs), respectively. SLs are implicated in signaling and membrane protein trafficking; however, the underlying processes linking SL composition to the control of symplasmic apertures remain unknown. The wide variety of SLs in plants prompted us to investigate which SL molecules are important for regulating symplasmic apertures in Arabidopsis (Arabidopsis thaliana). We introduced several potential SL pathway inhibitors and genetically modified SL contents using two independent SL pathway mutants. We were able to modulate callose deposition to control symplasmic connectivity through perturbations of SL metabolism. Alteration in glucosylhydroxyceramides or related SL composition particularly disturbed the secretory machinery for the GPI-anchored PdBG2 protein, resulting in an overaccumulation of callose. Moreover, our results revealed that SL-enriched lipid rafts link symplasmic channeling to PD callose homeostasis by controlling the targeting of GPI-anchored PdBG2. This study elevates our understanding of the molecular linkage underlying intracellular trafficking and precise targeting of GPI-anchored PD proteins incorporating glucosyl SLs.


Assuntos
Arabidopsis/metabolismo , Glucanos/metabolismo , Glicosilfosfatidilinositóis/metabolismo , Plasmodesmos/metabolismo , Esfingolipídeos/metabolismo , Proteínas de Arabidopsis/metabolismo
11.
Methods Mol Biol ; 2166: 103-120, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32710405

RESUMO

RNA transport and localization are evolutionarily conserved processes that allow protein translation to occur at specific subcellular sites and thereby having fundamental roles in the determination of cell fates, embryonic patterning, asymmetric cell division, and cell polarity. In addition to localizing RNA molecules to specific subcellular sites, plants have the ability to exchange RNA molecules between cells through plasmodesmata (PD). Plant RNA viruses hijack the mechanisms of intracellular and intercellular RNA transport to establish localized replication centers within infected cells and then to disseminate their infectious genomes between cells and throughout the plant organism with the help of their movement proteins (MP). In this chapter, we describe the transient expression of the tobacco mosaic virus movement protein (TMV-MP) and the application of the MS2 system for the in vivo labeling of the MP-encoding mRNA. The MS2 method is based on the binding of the bacteriophage coat protein (CP) to its origin of assembly (OAS) in the phage RNA. Thus, to label a specific mRNA in vivo, a tandem repetition of a 19-nucleotide-long stem-loop (SL) sequence derived from the MS2 OAS sequence (MSL) is transcriptionally fused to the RNA under investigation. The RNA is detected by the co-expression of fluorescent protein-tagged MS2 CP (MCP), which binds to each of the MSL elements. In providing a detailed protocol for the in vivo visualization of TMV-MP mRNA tagged with the MS2 system in Nicotiana benthamiana epidermal cells, we describe (1) the specific DNA constructs, (2) Agrobacterium tumefaciens-mediated transfection for their transient expression in plants, and (3) imaging conditions required to obtain high-quality mRNA imaging data.


Assuntos
Agrobacterium tumefaciens/genética , Levivirus/metabolismo , Proteínas do Movimento Viral em Plantas/genética , Transporte de RNA/genética , RNA Mensageiro/metabolismo , RNA de Plantas/metabolismo , RNA Viral/genética , Vírus do Mosaico do Tabaco/metabolismo , Transporte Biológico , Proteínas do Capsídeo/genética , Proteínas do Capsídeo/metabolismo , Clonagem Molecular , Expressão Gênica , Vetores Genéticos , Levivirus/genética , Proteínas Luminescentes , Microscopia de Fluorescência , Proteínas do Movimento Viral em Plantas/metabolismo , Plantas Geneticamente Modificadas/genética , Plasmodesmos/metabolismo , RNA Mensageiro/genética , Tabaco/genética , Tabaco/metabolismo , Vírus do Mosaico do Tabaco/genética
12.
Methods Mol Biol ; 2166: 145-155, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32710407

RESUMO

Multicellular organisms rely on systemic signals to orchestrate diverse developmental and physiological programs. To transmit environmental stimuli that perceived in the leaves, plants recruit many mobile molecules including mobile mRNAs as systemic signals for interorgan communication. The mobile mRNAs provide an efficient and specific remote control system for plants to cope with environmental dynamics. Upon being transcribed in local tissues, mobile mRNAs are selectively targeted to plasmodesmata for cell-to-cell and long-distance translocation. The mRNA labeling system based on the RNA-binding protein MS2 provides a useful tool to investigate intracellular trafficking of mobile mRNAs in plants. Here we describe the detailed protocol to visualize intracellular trafficking of plant mobile mRNAs by using the MS2 live-cell imaging system.


Assuntos
Agrobacterium tumefaciens/genética , Transporte Biológico/genética , Clonagem Molecular/métodos , Levivirus/genética , Microscopia Confocal/métodos , Folhas de Planta/metabolismo , Plasmodesmos/metabolismo , RNA Mensageiro/genética , Proteínas do Capsídeo/genética , Expressão Gênica , Vetores Genéticos , Processamento de Imagem Assistida por Computador , Proteínas Luminescentes/genética , Proteínas Luminescentes/metabolismo , Sinais de Localização Nuclear/genética , Sinais de Localização Nuclear/metabolismo , Folhas de Planta/genética , Plantas/genética , Plantas/metabolismo , RNA Mensageiro/metabolismo
13.
Commun Biol ; 3(1): 291, 2020 06 05.
Artigo em Inglês | MEDLINE | ID: mdl-32504045

RESUMO

Numerous cell surface receptors and receptor-like proteins (RLPs) undergo activation or deactivation via a transmembrane domain (TMD). A subset of plant RLPs distinctively localizes to the plasma membrane-lined pores called plasmodesmata. Those RLPs include the Arabidopsis thaliana Plasmodesmata-located protein (PDLP) 5, which is well known for its vital function regulating plasmodesmal gating and molecular movement between cells. In this study, we report that the TMD, although not a determining factor for the plasmodesmal targeting, serves essential roles for the PDLP5 function. In addition to its role for membrane anchoring, the TMD mediates PDLP5 self-interaction and carries an evolutionarily conserved motif that is essential for PDLP5 to regulate cell-to-cell movement. Computational modeling-based analyses suggest that PDLP TMDs have high propensities to dimerize. We discuss how a specific mode(s) of TMD dimerization might serve as a common mechanism for PDLP5 and other PDLP members to regulate cell-to-cell movement.


Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Movimento Celular , Evolução Molecular , Proteínas de Membrana/metabolismo , Plasmodesmos/metabolismo , Motivos de Aminoácidos , Arabidopsis/genética , Arabidopsis/crescimento & desenvolvimento , Proteínas de Arabidopsis/genética , Comunicação Celular , Proteínas de Membrana/genética , Domínios Proteicos
14.
Biochemistry (Mosc) ; 85(5): 531-544, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32571183

RESUMO

Plasmodesmata (PD) are intercellular channels in plant tissues providing continuity of the cytoplasm, the plasma membrane (PM) and the endoplasmic reticulum (ER) of neighboring cells. These channels allow the active transport of macromolecules such as proteins or RNAs. Thus, PD are believed to play a critical role in the functional unity of plant tissues and the transport of signals required for plant development and responses to external stimuli. Recent findings indicate that the PD channel contains a specialized type of ER-PM membrane contact sites (MCSs), structural links formed between ER and PM with tethering proteins. As shown for animal cells, MCSs are essential for lipid and protein trafficking between ER and PM membranes as well as for stress responses or the maintenance of ER structural integrity. On the other hand, our knowledge of the PD-specific MCSs is still scarce, and experimentally supported models of organization of their structural elements are only starting to emerge. Here, we review the structural and functional properties of proteins that can take part in establishing MCSs in PD. We also discuss the significance of cytoskeleton, lipid membrane microdomains and cell wall components for the maintenance and remodeling of PD-specific MCS in response to various biotic and abiotic stresses.


Assuntos
Membrana Celular/metabolismo , Parede Celular/metabolismo , Microdomínios da Membrana/metabolismo , Células Vegetais/metabolismo , Plasmodesmos/metabolismo , Citoesqueleto/metabolismo , Retículo Endoplasmático/metabolismo , Microtúbulos/metabolismo , Células Vegetais/ultraestrutura , Transporte Proteico
15.
Philos Trans R Soc Lond B Biol Sci ; 375(1801): 20190408, 2020 06 22.
Artigo em Inglês | MEDLINE | ID: mdl-32362251

RESUMO

The signalling pathways that regulate intercellular trafficking via plasmodesmata (PD) remain largely unknown. Analyses of mutants with defects in intercellular trafficking led to the hypothesis that chloroplasts are important for controlling PD, probably by retrograde signalling to the nucleus to regulate expression of genes that influence PD formation and function, an idea encapsulated in the organelle-nucleus-PD signalling (ONPS) hypothesis. ONPS is supported by findings that point to chloroplast redox state as also modulating PD. Here, we have attempted to further elucidate details of ONPS. Through reverse genetics, expression of select nucleus-encoded genes with known or predicted roles in chloroplast gene expression was knocked down, and the effects on intercellular trafficking were then assessed. Silencing most genes resulted in chlorosis, and the expression of several photosynthesis and tetrapyrrole biosynthesis associated nuclear genes was repressed in all silenced plants. PD-mediated intercellular trafficking was changed in the silenced plants, consistent with predictions of the ONPS hypothesis. One striking observation, best exemplified by silencing the PNPase homologues, was that the degree of chlorosis of silenced leaves was not correlated with the capacity for intercellular trafficking. Finally, we measured the distribution of PD in silenced leaves and found that intercellular trafficking was positively correlated with the numbers of PD. Together, these results not only provide further support for ONPS but also point to a genetic mechanism for PD formation, clarifying a longstanding question about PD and intercellular trafficking. This article is part of the theme issue 'Retrograde signalling from endosymbiotic organelles'.


Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/fisiologia , Núcleo Celular/fisiologia , Cloroplastos/fisiologia , Plasmodesmos/metabolismo , Transdução de Sinais , Transporte Proteico
16.
Development ; 147(6)2020 03 30.
Artigo em Inglês | MEDLINE | ID: mdl-32229613

RESUMO

Auxin is a key signal regulating plant growth and development. It is well established that auxin dynamics depend on the spatial distribution of efflux and influx carriers on the cell membranes. In this study, we employ a systems approach to characterise an alternative symplastic pathway for auxin mobilisation via plasmodesmata, which function as intercellular pores linking the cytoplasm of adjacent cells. To investigate the role of plasmodesmata in auxin patterning, we developed a multicellular model of the Arabidopsis root tip. We tested the model predictions using the DII-VENUS auxin response reporter, comparing the predicted and observed DII-VENUS distributions using genetic and chemical perturbations designed to affect both carrier-mediated and plasmodesmatal auxin fluxes. The model revealed that carrier-mediated transport alone cannot explain the experimentally determined auxin distribution in the root tip. In contrast, a composite model that incorporates both carrier-mediated and plasmodesmatal auxin fluxes re-capitulates the root-tip auxin distribution. We found that auxin fluxes through plasmodesmata enable auxin reflux and increase total root-tip auxin. We conclude that auxin fluxes through plasmodesmata modify the auxin distribution created by efflux and influx carriers.


Assuntos
Ácidos Indolacéticos/metabolismo , Meristema/metabolismo , Plasmodesmos/metabolismo , Arabidopsis/genética , Arabidopsis/crescimento & desenvolvimento , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Transporte Biológico/fisiologia , Regulação da Expressão Gênica no Desenvolvimento , Regulação da Expressão Gênica de Plantas , Desenvolvimento Vegetal/fisiologia , Raízes de Plantas/metabolismo , Plantas Geneticamente Modificadas , Distribuição Tecidual
17.
BMC Microbiol ; 20(1): 72, 2020 03 30.
Artigo em Inglês | MEDLINE | ID: mdl-32228456

RESUMO

BACKGROUND: Plant viruses move through plasmodesmata (PD) to infect new cells. To overcome the PD barrier, plant viruses have developed specific protein(s) to guide their genomic RNAs or DNAs to path through the PD. RESULTS: In the present study, we analyzed the function of Pepper vein yellows virus P4 protein. Our bioinformatic analysis using five commonly used algorithms showed that the P4 protein contains an transmembrane domain, encompassing the amino acid residue 117-138. The subcellular localization of P4 protein was found to target PD and form small punctates near walls. The P4 deletion mutant or the substitution mutant constructed by overlap PCR lost their function to produce punctates near the walls inside the fluorescent loci. The P4-YFP fusion was found to move from cell to cell in infiltrated leaves, and P4 could complement Cucumber mosaic virus movement protein deficiency mutant to move between cells. CONCLUSION: Taking together, we consider that the P4 protein is a movement protein of Pepper vein yellows virus.


Assuntos
Biologia Computacional/métodos , Vírus de Plantas/fisiologia , Tabaco/virologia , Proteínas Virais/metabolismo , Algoritmos , Cucumovirus/fisiologia , Mutação , Folhas de Planta/virologia , Plasmodesmos/metabolismo , Plasmodesmos/virologia , Domínios Proteicos , Tabaco/metabolismo , Proteínas Virais/química , Proteínas Virais/genética
18.
Plant Physiol Biochem ; 151: 284-291, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32248039

RESUMO

In view of recent findings, it is still a matter of debate whether the composition of the phloem sap of higher plants is specific and based on a plasmodesmal selectivity filter for macromolecular transport, or whether simply related to size, abundance and half-life of the macromolecules within the phloem sap. A range of reports indicates specific function of phloem-mobile signaling molecules such as the florigen making it indispensable to discriminate specific macromolecules entering the phloem from others which cannot cross this selectivity filter. Nevertheless, several findings have discussed for a non-selective transport via plasmodesmata, or contamination of the phloem sap by degradation products coming from immature still developing young sieve elements undergoing differentiation. Here, we discuss several possibilities, and raise the question how selectivity of the phloem sap composition could be achieved thereby focusing on mobility and dynamics of sucrose transporter mRNA and proteins.


Assuntos
Floema , Fenômenos Fisiológicos Vegetais , Plasmodesmos , Floema/química , Floema/metabolismo , Plasmodesmos/metabolismo , Transdução de Sinais
19.
Plant Cell Physiol ; 61(5): 942-956, 2020 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-32101300

RESUMO

Cell-to-cell communication is tightly regulated in response to environmental stimuli in plants. We previously used a photoconvertible fluorescent protein Dendra2 as a model reporter to study this process. This experiment revealed that macromolecular trafficking between protonemal cells in Physcomitrella patens is suppressed in response to abscisic acid (ABA). However, it remains unknown which ABA signaling components contribute to this suppression and how. Here, we show that ABA signaling components SUCROSE NON-FERMENTING 1-RELATED PROTEIN KINASE 2 (PpSnRK2) and ABA INSENSITIVE 3 (PpABI3) play roles as an essential and promotive factor, respectively, in regulating ABA-induced suppression of Dendra2 diffusion between cells (ASD). Our quantitative imaging analysis revealed that disruption of PpSnRK2 resulted in defective ASD onset itself, whereas disruption of PpABI3 caused an 81-min delay in the initiation of ASD. Live-cell imaging of callose deposition using aniline blue staining showed that, despite this onset delay, callose deposition on cross walls remained constant in the PpABI3 disruptant, suggesting that PpABI3 facilitates ASD in a callose-independent manner. Given that ABA is an important phytohormone to cope with abiotic stresses, we further explored cellular physiological responses. We found that the acquisition of salt stress tolerance is promoted by PpABI3 in a quantitative manner similar to ASD. Our results suggest that PpABI3-mediated ABA signaling may effectively coordinate cell-to-cell communication during the acquisition of salt stress tolerance. This study will accelerate the quantitative study for ABA signaling mechanism and function in response to various abiotic stresses.


Assuntos
Bryopsida/metabolismo , Proteínas de Plantas/metabolismo , Plasmodesmos/metabolismo , Ácido Abscísico/farmacologia , Bryopsida/citologia , Bryopsida/efeitos dos fármacos , Bryopsida/crescimento & desenvolvimento , Sobrevivência Celular/efeitos dos fármacos , Plasmodesmos/efeitos dos fármacos , Tolerância ao Sal/efeitos dos fármacos
20.
Proc Natl Acad Sci U S A ; 117(9): 5049-5058, 2020 03 03.
Artigo em Inglês | MEDLINE | ID: mdl-32051250

RESUMO

The coordinated redistribution of sugars from mature "source" leaves to developing "sink" leaves requires tight regulation of sugar transport between cells via plasmodesmata (PD). Although fundamental to plant physiology, the mechanisms that control PD transport and thereby support development of new leaves have remained elusive. From a forward genetic screen for altered PD transport, we discovered that the conserved eukaryotic glucose-TOR (TARGET OF RAPAMYCIN) metabolic signaling network restricts PD transport in leaves. Genetic approaches and chemical or physiological treatments to either promote or disrupt TOR activity demonstrate that glucose-activated TOR decreases PD transport in leaves. We further found that TOR is significantly more active in mature leaves photosynthesizing excess sugars than in young, growing leaves, and that this increase in TOR activity correlates with decreased rates of PD transport. We conclude that leaf cells regulate PD trafficking in response to changing carbohydrate availability monitored by the TOR pathway.


Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Células Vegetais/metabolismo , Folhas de Planta/metabolismo , Plasmodesmos/metabolismo , Arabidopsis/embriologia , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Transporte Biológico , Metabolismo dos Carboidratos , Perfilação da Expressão Gênica , Regulação da Expressão Gênica no Desenvolvimento , Regulação da Expressão Gênica de Plantas , Técnicas de Silenciamento de Genes , Inativação Gênica , Folhas de Planta/crescimento & desenvolvimento , Transporte Proteico , Transdução de Sinais , Tabaco/genética , Tabaco/metabolismo
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