Callose deposition, phloem-protein conformational changes and cell wall thickening are calcium-me... more Callose deposition, phloem-protein conformational changes and cell wall thickening are calcium-mediated occlusions occurring in the plant sieve elements in response to different biotic and abiotic stresses. However, the significance of these structures in plant-phytoplasma interactions requires in-depth investigations. We adopted a novel integrated approach, based on the combined use of microscopic and molecular analyses, to investigate the structural modifications induced in tomato leaf tissues in presence of phytoplasmas, focusing on vascular bundles and on the occlusion structures. Phloem hyperplasia and string-like arrangement of xylem vessels were found in infected vascular tissue. The diverse occlusion structures were differentially modulated in the phloem in response to phytoplasma infection. Callose amount was higher in midribs from infected plants than in healthy ones. Callose was observed at sieve plates but not at pore-plasmodesma units. A putative callose synthase gene e...
When expressed in transgenic tobacco plants, transgene mRNA that includes the 3' untranslated... more When expressed in transgenic tobacco plants, transgene mRNA that includes the 3' untranslated region (3' UTR) of Lettuce mosaic virus served as template for synthesis of complementary (-)-strand RNA following an infection by Tobacco etch virus, Tobacco vein mottle virus or Pepper mottle virus, but not when infected with Cucumber mosaic virus. Deletion of the 3' UTR from the transgene abolished the synthesis of (-)-strand transcripts. Similar results were obtained in transgenic tobacco plants expressing mRNA that includes the RNA3 3' UTR of Cucumber mosaic virus when infected with Tomato aspermy virus. These results show that the viral RNA-dependent RNA polymerase of several potyviruses and Tomato aspermy virus have the ability to recognize heterologous 3' UTRs when included in transgene mRNAs, and to use them as transcription promoters.
The plant cytoskeleton plays an important role in many biological processes, including cell divis... more The plant cytoskeleton plays an important role in many biological processes, including cell division and expansion, organogenesis, tip growth and intracellular signaling 1,2. The plant cytoskeleton is composed primarily of a network of microtubules and microfilaments (polymers of tubulin and actin, respectively) and diverse associated proteins. Whereas direct cell-to-cell communication is provided in animals and fungi by gap junctions and septal pores, respectively, within plants, cell-to-cell cytoplasmic trafficking takes place through plasmodesmata, wall spanning co-axial membranous organelles that bridge the cytoplasm of contiguous cells 3,4. Plasmodesmata are considered to enable both physiological and developmental coordination of the plant 5. Major insights in plasmodesmata functions have arisen from both viral movement studies and microinjection experiments 4 , and by the use of transiently expressed green fluorescent protein (GFP) fusion proteins 6. Functional studies have underlined that plasmodesmata are dynamic structures that Plant cell-to-cell communication is achieved by membranous conduits called plasmodesmata, which bridge the cytoplasm of neighboring cells. A growing body of immunolocalization data shows an association of the cytoskeleton machinery with plasmodesmata. The role of the cytoskeleton in the plasmodesmata-mediated transport has been well documented for virus movement. Because viruses are known to exploit existing host pathways and because the cytoskeleton is involved in intracellular trafficking, the cytoskeleton is thought to drive and target macromolecules to plasmodesmata. It is this link between plasmodesmata and the cytoskeleton that will be described here.
Commercial tobacco cultivars BB16 (burley) and PBD6 (dark air cured) were transformed with the co... more Commercial tobacco cultivars BB16 (burley) and PBD6 (dark air cured) were transformed with the coat protein gene of lettuce mosaic potyvirus (LMV). Transgenic BB16 plants showed resistance to potato virus Y (PVY) infection, against the necrotic strain PVY-N Versailles, as well as the resistance breaking necrotic strain PVY-N 107. Transgenic PBD6, which carries the recessive ‘va’ gene conferring resistance to
The complex process of phloem sugar transport involves symplasmic and apoplasmic events. We chara... more The complex process of phloem sugar transport involves symplasmic and apoplasmic events. We characterized Arabidopsis thaliana lines ectopically expressing a phloem-specific gene encoding NDR1/HIN1-like26 (NHL26), a putative membrane protein. NHL26 overexpressor plants grew more slowly than wild-type plants, accumulated high levels of carbohydrates in mature leaves, and had a higher shoot biomass, contrasting with slower root growth and a lower seed yield. Similar effects were observed when NHL26 was overexpressed in companion cells, under the control of a companion cell-specific promoter. The soluble sugar content of the phloem sap and sink organs was lower than that in the wild type, providing evidence of a sugar export defect. This was confirmed in a phloem-export assay with the symplastic tracer carboxyfluorescein diacetate. Leaf sugar accumulation was accompanied by higher organic acid, amino acid, and protein contents, whereas analysis of the metabolite profile of phloem sap exudate revealed no change in amino acid or organic acid content, indicating a specific effect on sugar export. NHL26 was found to be located in the phloem plasmodesmata and the endoplasmic reticulum. These findings reveal that NHL26 accumulation affects either the permeability of plasmodesmata or sugar signaling in companion cells, with a specific effect on sugar export.
Phloem protein 2 (PP2) is one of the most abundant and enigmatic proteins in the phloem sap. Alth... more Phloem protein 2 (PP2) is one of the most abundant and enigmatic proteins in the phloem sap. Although thought to be associated with structural P-protein, PP2 is translocated in the assimilate stream where its lectin activity or RNA-binding properties can exert effects over long distances. Analyzing the diversity of these proteins in vascular plants led to the identification ofPP2-like genes in species from 17 angiosperm and gymnosperm genera. This wide distribution of PP2 genes in the plant kingdom indicates that they are ancient and common in vascular plants. Their presence in cereals and gymnosperms, both of which lack structural P-protein, also supports a wider role for these proteins. Within this superfamily, PP2 proteins have considerable size polymorphism. This is attributable to variability in the length of the amino terminus that extends from a highly conserved domain. The conserved PP2 domain was identified in the proteins encoded by six genes from several cucurbits, celery...
Poleroviruses are phytoviruses strictly transmitted by phloem-feeding aphids in a circulative and... more Poleroviruses are phytoviruses strictly transmitted by phloem-feeding aphids in a circulative and nonpropagative mode. During ingestion, aphids sample virions in sieve tubes along with sap. Therefore, any sap protein bound to virions will be acquired by the insects and could potentially be involved in the transmission process. By developing in vitro virus-overlay assays on sap proteins collected from cucumber, we observed that approximately 20 proteins were able to bind to purified particles of Cucurbit aphid borne yellows virus (CABYV). Among them, eight proteins were identified by mass spectrometry. The role of two candidates belonging to the PP2-like family (predominant lectins found in cucurbit sap) in aphid transmission was further pursued by using purified orthologous PP2 proteins from Arabidopsis. Addition of these proteins to the virus suspension in the aphid artificial diet greatly increased virus transmission rate. This shift was correlated with an increase in the number o...
Phloem Protein2 (PP2) is a component of the phloem protein bodies found in sieve elements. We des... more Phloem Protein2 (PP2) is a component of the phloem protein bodies found in sieve elements. We describe here the lectin properties of the Arabidopsis (Arabidopsis thaliana) PP2-A1. Using a recombinant protein produced in Escherichia coli, we demonstrated binding to N-acetylglucosamine oligomers. Glycan array screening showed that PP2-A1 also bound to high-mannose N-glycans and 9-acyl-N-acetylneuraminic sialic acid. Fluorescence spectroscopy-based titration experiments revealed that PP2-A1 had two classes of binding site for N,N',N''-triacetylchitotriose, a low-affinity site and a high-affinity site, promoting the formation of protein dimers. A search for structural similarities revealed that PP2-A1 aligned with the Cbm4 and Cbm22-2 carbohydrate-binding modules, leading to the prediction of a beta-strand structure for its conserved domain. We investigated whether PP2-A1 interacted with phloem sap glycoproteins by first characterizing abundant Arabidopsis phloem sap prot...
The positive-strand RNA bromoviruses encode two nonstructural proteins, la and 2a, involved in RN... more The positive-strand RNA bromoviruses encode two nonstructural proteins, la and 2a, involved in RNA-dependent RNA replication. These proteins have extensive sequence similarities with methyltransferase, helicase, and polymerase proteins of other plant and animal viruses. la and 2a can also form a complex in vitro. To explore whether la-2a interaction is required for RNA replication in vivo, we reassorted the la and 2a genes from two different bromoviruses, brome mosaic virus (BMV) and cowpea chlorotic mottle virus (CCMV). la and 2a were expressed independently of viral replication by using RNAor DNA-based transient expression, and their in vivo RNA replication activities were tested in protoplasts with BMV and CCMV RNA3 templates. RNA-based transient expression confirmed prior indications that bromovirus RNA replication is more sensitive to reductions in la expression than to reductions in 2a expression. DNA-based expression of the homologous combinations of la and 2a supported high levels of RNA synthesis, but both la-2a heterologous combinations exhibited RNA synthesis defects. The combination of CCMV la and BMV 2a did not support detectable synthesis of negative-strand, positive-strand, or subgenomic RNA. The converse combination of BMV la and CCMV 2a was preferentially defective in positive-strand and subgenomic RNA accumulation, showing that la-2a interaction is involved in these processes in ways distinct from negative-strand RNA synthesis, which was only slightly affected. These results indicate that at least some functions of la and 2a operate in a mutually dependent manner in vivo and that the mechanisms of positiveand negative-strand RNA synthesis are differentiated in part by features of such interactions.
(J. Kehr). importantes pour l'analyse de la composition de la séve et la physiologie des tubes cr... more (J. Kehr). importantes pour l'analyse de la composition de la séve et la physiologie des tubes criblés. Un tableau assez complet des metabolites, des ions, des hormones, des protéines et des ARN presents dans la séve du phloéme est désormais disponible. Ces avancées ont largement étayé le role du phloéme comme réseau de signalisation, en plus de son role central dans la repartition des photo-assimilats. Ces méthodes d'echantillonnage de la séve élaborée ont aussi des applications potentielles remarquables pour l'analyse de la nutrition, la physiologie et les reactions de defense des plantes. Plusieurs études ayant suggéré que la qualité de la séve phloémienne pouvait affecter le comportement alimentaire des pucerons, des tentatives de manipulation de la composition de la séve phloémienne ont été engagées, basées sur la deregulation des genes controlant la teneur en acides amines ou en sucres de la séve, dans les plantes transgéniques. De telles approches ouvrent des strategies nouvelles pour limiter le développement des pucerons sur leur plante hote et ainsi leur nuisibilité.
Phloem sieve elements are highly differentiated cells involved in the long-distance transport of ... more Phloem sieve elements are highly differentiated cells involved in the long-distance transport of photoassimilates. These cells contain both aggregated phloem-proteins (P-proteins) and soluble proteins, which are also translocated by mass flow. We used liquid chromatography-tandem mass spectrometry (LC-MS/MS) to carry out a proteomic survey of the phloem exudate of Arabidopsis thaliana, collected by the ethylenediaminetetraacetic acid (EDTA)-facilitated method. We identified 287 proteins, a large proportion of which were enzymes involved in the metabolic precursor generation and amino acid synthesis, suggesting that sieve tubes display high levels of metabolic activity. RNA-binding proteins, defence proteins and lectins were also found. No putative P-proteins were detected in the EDTA-exudate fraction, indicating a lack of long-distance translocation of such proteins in Arabidopsis. In parallel, we investigated the organization of P-proteins, by high-resolution transmission electron ...
To access phloem sap, aphids have developed a furtive strategy, their stylets progressing towards... more To access phloem sap, aphids have developed a furtive strategy, their stylets progressing towards sieve tubes mainly through the apoplasmic compartment. Aphid feeding requires that they overcome a number of plant responses, ranging from sieve tube occlusion and activation of phytohormone-signalling pathways to expression of anti-insect molecules. In addition to bypassing plant defences, aphids have been shown to affect plant primary metabolism, which could be a strategy to improve phloem sap composition in nutrients required for their growth. During compatible interactions, leading to successful feeding and reproduction, aphids cause alterations in their host plant, including morphological changes, modified resource allocation and various local as well as systemic symptoms. Repeated salivary secretions injected from the first probe in the epidermal tissue up to ingestion of sieve-tube sap may play a crucial role in the compatibility between the aphid and the plant.
Phloem sieve elements are highly differentiated cells involved in the long-distance transport of ... more Phloem sieve elements are highly differentiated cells involved in the long-distance transport of photoassimilates. These cells contain both aggregated phloem-proteins (P-proteins) and soluble proteins, which are also translo-cated by mass flow. We used liquid chromatography– tandem mass spectrometry (LC-MS/MS) to carry out a proteomic survey of the phloem exudate of Arabidopsis thaliana, collected by the ethylenediaminetetraacetic acid (EDTA)-facilitated method. We identified 287 proteins, a large proportion of which were enzymes involved in the metabolic precursor generation and amino acid synthesis, suggesting that sieve tubes display high levels of metabolic activity. RNA-binding proteins, defence proteins and lectins were also found. No putative P-proteins were detected in the EDTA-exudate fraction, indicating a lack of long-distance translocation of such proteins in Arabidopsis. In parallel, we investigated the organization of P-proteins, by high-resolution transmission electron microscopy, and the localization of the phloem lectin PP2, a putative P-protein component, by immunolocalization with antibodies against PP2-A1. Transmission electron microscopy observations of P-proteins revealed bundles of filaments resembling strings of beads. PP2-A1 was found weakly associated with these structures in the sieve elements and bound to plastids. These observations suggest that PP2-A1 is anchored to P-proteins and organelles rather than being a structural component of P-proteins.
Callose deposition, phloem-protein conformational changes and cell wall thickening are calcium-me... more Callose deposition, phloem-protein conformational changes and cell wall thickening are calcium-mediated occlusions occurring in the plant sieve elements in response to different biotic and abiotic stresses. However, the significance of these structures in plant-phytoplasma interactions requires in-depth investigations. We adopted a novel integrated approach, based on the combined use of microscopic and molecular analyses, to investigate the structural modifications induced in tomato leaf tissues in presence of phytoplasmas, focusing on vascular bundles and on the occlusion structures. Phloem hyperplasia and string-like arrangement of xylem vessels were found in infected vascular tissue. The diverse occlusion structures were differentially modulated in the phloem in response to phytoplasma infection. Callose amount was higher in midribs from infected plants than in healthy ones. Callose was observed at sieve plates but not at pore-plasmodesma units. A putative callose synthase gene e...
When expressed in transgenic tobacco plants, transgene mRNA that includes the 3' untranslated... more When expressed in transgenic tobacco plants, transgene mRNA that includes the 3' untranslated region (3' UTR) of Lettuce mosaic virus served as template for synthesis of complementary (-)-strand RNA following an infection by Tobacco etch virus, Tobacco vein mottle virus or Pepper mottle virus, but not when infected with Cucumber mosaic virus. Deletion of the 3' UTR from the transgene abolished the synthesis of (-)-strand transcripts. Similar results were obtained in transgenic tobacco plants expressing mRNA that includes the RNA3 3' UTR of Cucumber mosaic virus when infected with Tomato aspermy virus. These results show that the viral RNA-dependent RNA polymerase of several potyviruses and Tomato aspermy virus have the ability to recognize heterologous 3' UTRs when included in transgene mRNAs, and to use them as transcription promoters.
The plant cytoskeleton plays an important role in many biological processes, including cell divis... more The plant cytoskeleton plays an important role in many biological processes, including cell division and expansion, organogenesis, tip growth and intracellular signaling 1,2. The plant cytoskeleton is composed primarily of a network of microtubules and microfilaments (polymers of tubulin and actin, respectively) and diverse associated proteins. Whereas direct cell-to-cell communication is provided in animals and fungi by gap junctions and septal pores, respectively, within plants, cell-to-cell cytoplasmic trafficking takes place through plasmodesmata, wall spanning co-axial membranous organelles that bridge the cytoplasm of contiguous cells 3,4. Plasmodesmata are considered to enable both physiological and developmental coordination of the plant 5. Major insights in plasmodesmata functions have arisen from both viral movement studies and microinjection experiments 4 , and by the use of transiently expressed green fluorescent protein (GFP) fusion proteins 6. Functional studies have underlined that plasmodesmata are dynamic structures that Plant cell-to-cell communication is achieved by membranous conduits called plasmodesmata, which bridge the cytoplasm of neighboring cells. A growing body of immunolocalization data shows an association of the cytoskeleton machinery with plasmodesmata. The role of the cytoskeleton in the plasmodesmata-mediated transport has been well documented for virus movement. Because viruses are known to exploit existing host pathways and because the cytoskeleton is involved in intracellular trafficking, the cytoskeleton is thought to drive and target macromolecules to plasmodesmata. It is this link between plasmodesmata and the cytoskeleton that will be described here.
Commercial tobacco cultivars BB16 (burley) and PBD6 (dark air cured) were transformed with the co... more Commercial tobacco cultivars BB16 (burley) and PBD6 (dark air cured) were transformed with the coat protein gene of lettuce mosaic potyvirus (LMV). Transgenic BB16 plants showed resistance to potato virus Y (PVY) infection, against the necrotic strain PVY-N Versailles, as well as the resistance breaking necrotic strain PVY-N 107. Transgenic PBD6, which carries the recessive ‘va’ gene conferring resistance to
The complex process of phloem sugar transport involves symplasmic and apoplasmic events. We chara... more The complex process of phloem sugar transport involves symplasmic and apoplasmic events. We characterized Arabidopsis thaliana lines ectopically expressing a phloem-specific gene encoding NDR1/HIN1-like26 (NHL26), a putative membrane protein. NHL26 overexpressor plants grew more slowly than wild-type plants, accumulated high levels of carbohydrates in mature leaves, and had a higher shoot biomass, contrasting with slower root growth and a lower seed yield. Similar effects were observed when NHL26 was overexpressed in companion cells, under the control of a companion cell-specific promoter. The soluble sugar content of the phloem sap and sink organs was lower than that in the wild type, providing evidence of a sugar export defect. This was confirmed in a phloem-export assay with the symplastic tracer carboxyfluorescein diacetate. Leaf sugar accumulation was accompanied by higher organic acid, amino acid, and protein contents, whereas analysis of the metabolite profile of phloem sap exudate revealed no change in amino acid or organic acid content, indicating a specific effect on sugar export. NHL26 was found to be located in the phloem plasmodesmata and the endoplasmic reticulum. These findings reveal that NHL26 accumulation affects either the permeability of plasmodesmata or sugar signaling in companion cells, with a specific effect on sugar export.
Phloem protein 2 (PP2) is one of the most abundant and enigmatic proteins in the phloem sap. Alth... more Phloem protein 2 (PP2) is one of the most abundant and enigmatic proteins in the phloem sap. Although thought to be associated with structural P-protein, PP2 is translocated in the assimilate stream where its lectin activity or RNA-binding properties can exert effects over long distances. Analyzing the diversity of these proteins in vascular plants led to the identification ofPP2-like genes in species from 17 angiosperm and gymnosperm genera. This wide distribution of PP2 genes in the plant kingdom indicates that they are ancient and common in vascular plants. Their presence in cereals and gymnosperms, both of which lack structural P-protein, also supports a wider role for these proteins. Within this superfamily, PP2 proteins have considerable size polymorphism. This is attributable to variability in the length of the amino terminus that extends from a highly conserved domain. The conserved PP2 domain was identified in the proteins encoded by six genes from several cucurbits, celery...
Poleroviruses are phytoviruses strictly transmitted by phloem-feeding aphids in a circulative and... more Poleroviruses are phytoviruses strictly transmitted by phloem-feeding aphids in a circulative and nonpropagative mode. During ingestion, aphids sample virions in sieve tubes along with sap. Therefore, any sap protein bound to virions will be acquired by the insects and could potentially be involved in the transmission process. By developing in vitro virus-overlay assays on sap proteins collected from cucumber, we observed that approximately 20 proteins were able to bind to purified particles of Cucurbit aphid borne yellows virus (CABYV). Among them, eight proteins were identified by mass spectrometry. The role of two candidates belonging to the PP2-like family (predominant lectins found in cucurbit sap) in aphid transmission was further pursued by using purified orthologous PP2 proteins from Arabidopsis. Addition of these proteins to the virus suspension in the aphid artificial diet greatly increased virus transmission rate. This shift was correlated with an increase in the number o...
Phloem Protein2 (PP2) is a component of the phloem protein bodies found in sieve elements. We des... more Phloem Protein2 (PP2) is a component of the phloem protein bodies found in sieve elements. We describe here the lectin properties of the Arabidopsis (Arabidopsis thaliana) PP2-A1. Using a recombinant protein produced in Escherichia coli, we demonstrated binding to N-acetylglucosamine oligomers. Glycan array screening showed that PP2-A1 also bound to high-mannose N-glycans and 9-acyl-N-acetylneuraminic sialic acid. Fluorescence spectroscopy-based titration experiments revealed that PP2-A1 had two classes of binding site for N,N',N''-triacetylchitotriose, a low-affinity site and a high-affinity site, promoting the formation of protein dimers. A search for structural similarities revealed that PP2-A1 aligned with the Cbm4 and Cbm22-2 carbohydrate-binding modules, leading to the prediction of a beta-strand structure for its conserved domain. We investigated whether PP2-A1 interacted with phloem sap glycoproteins by first characterizing abundant Arabidopsis phloem sap prot...
The positive-strand RNA bromoviruses encode two nonstructural proteins, la and 2a, involved in RN... more The positive-strand RNA bromoviruses encode two nonstructural proteins, la and 2a, involved in RNA-dependent RNA replication. These proteins have extensive sequence similarities with methyltransferase, helicase, and polymerase proteins of other plant and animal viruses. la and 2a can also form a complex in vitro. To explore whether la-2a interaction is required for RNA replication in vivo, we reassorted the la and 2a genes from two different bromoviruses, brome mosaic virus (BMV) and cowpea chlorotic mottle virus (CCMV). la and 2a were expressed independently of viral replication by using RNAor DNA-based transient expression, and their in vivo RNA replication activities were tested in protoplasts with BMV and CCMV RNA3 templates. RNA-based transient expression confirmed prior indications that bromovirus RNA replication is more sensitive to reductions in la expression than to reductions in 2a expression. DNA-based expression of the homologous combinations of la and 2a supported high levels of RNA synthesis, but both la-2a heterologous combinations exhibited RNA synthesis defects. The combination of CCMV la and BMV 2a did not support detectable synthesis of negative-strand, positive-strand, or subgenomic RNA. The converse combination of BMV la and CCMV 2a was preferentially defective in positive-strand and subgenomic RNA accumulation, showing that la-2a interaction is involved in these processes in ways distinct from negative-strand RNA synthesis, which was only slightly affected. These results indicate that at least some functions of la and 2a operate in a mutually dependent manner in vivo and that the mechanisms of positiveand negative-strand RNA synthesis are differentiated in part by features of such interactions.
(J. Kehr). importantes pour l'analyse de la composition de la séve et la physiologie des tubes cr... more (J. Kehr). importantes pour l'analyse de la composition de la séve et la physiologie des tubes criblés. Un tableau assez complet des metabolites, des ions, des hormones, des protéines et des ARN presents dans la séve du phloéme est désormais disponible. Ces avancées ont largement étayé le role du phloéme comme réseau de signalisation, en plus de son role central dans la repartition des photo-assimilats. Ces méthodes d'echantillonnage de la séve élaborée ont aussi des applications potentielles remarquables pour l'analyse de la nutrition, la physiologie et les reactions de defense des plantes. Plusieurs études ayant suggéré que la qualité de la séve phloémienne pouvait affecter le comportement alimentaire des pucerons, des tentatives de manipulation de la composition de la séve phloémienne ont été engagées, basées sur la deregulation des genes controlant la teneur en acides amines ou en sucres de la séve, dans les plantes transgéniques. De telles approches ouvrent des strategies nouvelles pour limiter le développement des pucerons sur leur plante hote et ainsi leur nuisibilité.
Phloem sieve elements are highly differentiated cells involved in the long-distance transport of ... more Phloem sieve elements are highly differentiated cells involved in the long-distance transport of photoassimilates. These cells contain both aggregated phloem-proteins (P-proteins) and soluble proteins, which are also translocated by mass flow. We used liquid chromatography-tandem mass spectrometry (LC-MS/MS) to carry out a proteomic survey of the phloem exudate of Arabidopsis thaliana, collected by the ethylenediaminetetraacetic acid (EDTA)-facilitated method. We identified 287 proteins, a large proportion of which were enzymes involved in the metabolic precursor generation and amino acid synthesis, suggesting that sieve tubes display high levels of metabolic activity. RNA-binding proteins, defence proteins and lectins were also found. No putative P-proteins were detected in the EDTA-exudate fraction, indicating a lack of long-distance translocation of such proteins in Arabidopsis. In parallel, we investigated the organization of P-proteins, by high-resolution transmission electron ...
To access phloem sap, aphids have developed a furtive strategy, their stylets progressing towards... more To access phloem sap, aphids have developed a furtive strategy, their stylets progressing towards sieve tubes mainly through the apoplasmic compartment. Aphid feeding requires that they overcome a number of plant responses, ranging from sieve tube occlusion and activation of phytohormone-signalling pathways to expression of anti-insect molecules. In addition to bypassing plant defences, aphids have been shown to affect plant primary metabolism, which could be a strategy to improve phloem sap composition in nutrients required for their growth. During compatible interactions, leading to successful feeding and reproduction, aphids cause alterations in their host plant, including morphological changes, modified resource allocation and various local as well as systemic symptoms. Repeated salivary secretions injected from the first probe in the epidermal tissue up to ingestion of sieve-tube sap may play a crucial role in the compatibility between the aphid and the plant.
Phloem sieve elements are highly differentiated cells involved in the long-distance transport of ... more Phloem sieve elements are highly differentiated cells involved in the long-distance transport of photoassimilates. These cells contain both aggregated phloem-proteins (P-proteins) and soluble proteins, which are also translo-cated by mass flow. We used liquid chromatography– tandem mass spectrometry (LC-MS/MS) to carry out a proteomic survey of the phloem exudate of Arabidopsis thaliana, collected by the ethylenediaminetetraacetic acid (EDTA)-facilitated method. We identified 287 proteins, a large proportion of which were enzymes involved in the metabolic precursor generation and amino acid synthesis, suggesting that sieve tubes display high levels of metabolic activity. RNA-binding proteins, defence proteins and lectins were also found. No putative P-proteins were detected in the EDTA-exudate fraction, indicating a lack of long-distance translocation of such proteins in Arabidopsis. In parallel, we investigated the organization of P-proteins, by high-resolution transmission electron microscopy, and the localization of the phloem lectin PP2, a putative P-protein component, by immunolocalization with antibodies against PP2-A1. Transmission electron microscopy observations of P-proteins revealed bundles of filaments resembling strings of beads. PP2-A1 was found weakly associated with these structures in the sieve elements and bound to plastids. These observations suggest that PP2-A1 is anchored to P-proteins and organelles rather than being a structural component of P-proteins.
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