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Francis Martin - One of the best experts on this subject based on the ideXlab platform.
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Laser microdissection and microarray analysis of Tuber melanosporum ectomycorrhizas reveal functional heterogeneity between mantle and Hartig Net compartments
Environmental Microbiology, 2013Co-Authors: Stéphane Hacquard, Francis Martin, Annick Brun, Emilie Tisserant, Valérie Legué, Annegret KohlerAbstract:The ectomycorrhizal (ECM) symbiosis, a mutualistic plant-fungus association, plays a fundamental role in forest ecosystems by enhancing plant growth and by providing host protection from root diseases. The cellular complexity of the symbiotic organ, characterized by the differentiation of structurally specialized tissues (i.e. the fungal mantle and the Hartig Net), is the major limitation to study fungal gene expression in such specific compartments. We investigated the transcriptional landscape of the ECM fungus Tuber melanosporum during the major stages of its life cycle and we particularly focused on the complex symbiotic stage by combining the use of laser capture microdissection and microarray gene expression analysis. We isolated the fungal/soil (i.e. the mantle) and the fungal/plant (i.e. the Hartig Net) interfaces from transverse sections of T.melanosporum/Corylus avellana ectomycorrhizas and identified the distinct geNetic programmes associated with each compartment. Particularly, nitrogen and water acquisition from soil, synthesis of secondary metabolites and detoxification mechanisms appear to be important processes in the fungal mantle. In contrast, transport activity is enhanced in the Hartig Net and we identified carbohydrate and nitrogen-derived transporters that might play a key role in the reciprocal resources' transfer between the host and the symbiont.
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immunolocalization of hydrophobin hydpt 1 from the ectomycorrhizal basidiomycete pisolithus tinctorius during colonization of eucalyptus globulus roots
New Phytologist, 2001Co-Authors: Denis Tagu, O M H De Vries, Giovanni Piccoli, Ray Bellis, Paola Bonfante, Raffaella Balestrini, Vilberto Stocchi, Francis MartinAbstract:• The immunolocalization of one of the hydrophobins of Pisolithustinctorius (HYDPt-1) is reported. Hydrophobin proteins play key roles in adhesion and aggregation of fungal hyphae, and it is already known that formation of ectomycorrhizas on eucalypt roots enhances the accumulation of hydrophobin mRNAs in the mycelium of Pisolithus tinctorius. • Purification of SDS-insoluble proteins from the mycelium of P. tinctorius showed the presence of a 13 kDa polypeptide with properties of class I hydrophobin. • Polyconal antibodies were raised against a recombinant HYDPt-1 polypeptide, and these were used for immunofluorescence-coupled transmission electron microscopy. • HYDPt-1 is a cell wall protein located at the surface of the hyphae with no preferential accumulation in the fungal cells of the different tissues of the ectomycorrhiza (i.e. extraradical hyphae, mantle or Hartig Net).
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cell wall proteins of the ectomycorrhizal basidiomycete pisolithus tinctorius identification function and expression in symbiosis
Fungal Genetics and Biology, 1999Co-Authors: Francis Martin, Catherine Voiblet, D. Carvalho, Paola Bonfante, Raffaella Balestrini, P. Laurent, Denis TaguAbstract:Abstract Specific cell–cell and cell–substrate interactions direct the growth of ectomycorrhizal fungi to their host root targets. These elaborate mechanisms lead to the differentiation of distinct multihyphal structures, the mantle, and the Hartig Net. In the ectomycorrhizal basidiomycete Pisolithus tinctorius, the use of two-dimensional gel electrophoresis, immunocytochemical microscopy, and RNA blot analysis has demonstrated the differential expression of cell wall proteins (CWPs), such as hydrophobins, adhesins, and mannoproteins, during symbiotic interaction. In other fungi, these CWPs have been suggested to play a role in hyphae aggregation, intracellular signaling cascades, and cytoskeletal changes. The recent cloning of the genes for several of these CWPs in P. tinctorius allows us to address their function in symbiosis. This review summarizes our knowledge of CWPs in P. tinctorius and considers parallels with other biotrophic fungi as a possible framework for future work.
Ivano Brunner - One of the best experts on this subject based on the ideXlab platform.
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Freeze fracturing for low-temperature scanning electron microscopy of Hartig Net in synthesized Picea abies - Hebeloma crustuliniforme and - Tricholoma vaccinum ectomycorrhizas
New Phytologist, 2006Co-Authors: Christoph Scheidegger, Ivano BrunnerAbstract:SUMMARY Rapidly frozen ectomycorrhizal roots were freeze-fractured under high vacuum in a dedicated cryo-preparation unit and investigated in the frozen, partially freeze-dried, or fully hydrated state. Tangential, radial and occasional transverse surface views of Hartig Net were obtained after freeze-fracturing. Protoplasmic and exoplasmic fracture faces of the fungal plasmalemma und occasional fractures between the fungal cell wall and the extracellular matrix were seen. Primary pit-fields with plasmodesmata occurred in non-ectomycorrhizal parts of the root as well as in small gaps of Hartig Net. Septa with dolipores were observed in the finger-like complexes of the Hartig Net.
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extracellular complexation of cd in the Hartig Net and cytosolic zn sequestration in the fungal mantle of picea abies hebeloma crustuliniforme ectomycorrhizas
Plant Cell and Environment, 2000Co-Authors: Beat Frey, Karl Zierold, Ivano BrunnerAbstract:Compartmentation of heavy metals on or within mycorrhizal fungi may serve as a protective function for the roots of forest trees growing in soils containing elevated concentrations of metals such as Cd and Zn. In this paper we present the first quantitative measurements by X-ray microanalysis of heavy metals in high-pressure frozen and cryosectioned ectomycorrhizal fungal hyphae. We used this technique to analyse the main sites of Cd and Zn in fungal cells of mantle and Hartig Net hyphae and in cortical root cells of symbiotic Picea abies - Hebeloma crustuliniforme associations to gain new insights into the mechanisms of detoxification of these two metals in Norway spruce seedlings. The mycorrhizal seedlings were exposed in growth pouches to either 1 mM Cd or 2 mM Zn for 5 weeks. The microanalytical data revealed that two distinct Cd- and Zn-binding mechanisms are involved in cellular compartmentation of Cd and Zn in the mycobiont. Whereas extracellular complexation of Cd occurred predominantly in the Hartig Net hyphae, both extracellular complexation and cytosolic sequestration of Zn occurred in the fungal tissue. The vacuoles were presumed not to be a significant pool for Cd and Zn storage. Cadmium was almost exclusively localized in the cell walls of the Hartig Net (up to 161 mmol kg -1 DW) compared with significantly lower concentrations in the cell walls of mantle hyphae (22 mmol kg -1 DW) and in the cell walls of cortical cells (15 mmol kg -1 DW). This suggests that the apoplast of the Hartig Net is a primary accumulation site for Cd. Zinc accumulated mainly in the cell walls of the mantle hyphae (111 mmol kg -1 DW), the Hartig Net hyphae (130 mmol kg -1 DW) and the cortical cells (152 mmol kg -1 DW). In addition, Zn occurred in high concentrations in the cytoplasm of the fungal mantle hyphae (up to 164 mmol kg -1 DW) suggesting that both the cell walls and the cytoplasm of fungal tissue are the main accumulation sites for Zn in P. abies resulting in decreased Zn transfer from the fungus to the root.
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production of ectomycorrhizal ppicea abies hebeloma crustuliniforme seedlings for ecological studies effects of synthesis techniques on the morphology of the symbiosis
Water Air and Soil Pollution, 1993Co-Authors: Ivano BrunnerAbstract:Various in vitro synthesis techniques were applied to produce numerous ectomycorrhizal Picea abies — Hebeloma crustuliniforme seedlings which could be later used for experiments in the field of environmental research. Included synthesis methods are the Erlenmeyer flask technique, the cuvette technique, and the growth pouch technique. A mantle and a Hartig Net were present in all synthesized and naturally grown symbioses. Stable features were the white color of the mantle, its two-layered structure, and the structure of the Hartig Net. A few characters such as the thickness of the mantle and the development of the Hartig Net formation were variable due to the synthesis method. The techniques suitable for producing ectomycorrhizal seedlings are the Erlenmeyer flask technique using vermiculite and peat moss and the growth pouch technique: the first technique because of sterility and good ectomycorrhizal formation within approximately two months, the second technique because of direct observation of ectomycorrhizal formation within one to two months and complete removing of ectomycorrhizae including extramatrical mycelia.
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Ontogeny of synthesized Picea abies (L.) Karst.–Hebeloma crustuliniforme (Bull. ex St Amans) Quél. ectomycorrhizas
New Phytologist, 1992Co-Authors: Ivano Brunner, Christoph ScheideggerAbstract:summary Ectomycorrhizas were synthesized between Picea abies (L.) Karst. and Hebeloma crustuliniforme (Bull. ex St Amans) Quel. in growth pouches containing activated charcoal filter paper. Immediately emanating hyphae from inoculated plugs contacted root hairs and root surfaces, hyphal morphology was altered. They became short-celled with few clamp connections, highly branched, and lacked cell wall ornamentation. Then, hyphae on root surfaces formed an inner synenchymatous mantle and loose hyphae formed an outer prosenchymatous mantle. Simultaneously, root cap cells became invaded by fungal hyphae and their remnants were incorporated into the inner mantle. Further root elongation took place but root hair production was suppressed. Thirdly, a Hartig Net began to form in basal portions of developing ectomycorrhizas and the fungal mantle became denser and thicker. Fourthly, growth of the root was reduced and the hyphae of the Hartig Net peNetrated between cortical cells moving toward the apex. Fifthly, metacutis occasionally surrounded the meristem. At this stage the formation of the ectomycorrhiza can be considered to be complete and the rootlet itself is temporarily in a dormant state.
Denis Tagu - One of the best experts on this subject based on the ideXlab platform.
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immunolocalization of hydrophobin hydpt 1 from the ectomycorrhizal basidiomycete pisolithus tinctorius during colonization of eucalyptus globulus roots
New Phytologist, 2001Co-Authors: Denis Tagu, O M H De Vries, Giovanni Piccoli, Ray Bellis, Paola Bonfante, Raffaella Balestrini, Vilberto Stocchi, Francis MartinAbstract:• The immunolocalization of one of the hydrophobins of Pisolithustinctorius (HYDPt-1) is reported. Hydrophobin proteins play key roles in adhesion and aggregation of fungal hyphae, and it is already known that formation of ectomycorrhizas on eucalypt roots enhances the accumulation of hydrophobin mRNAs in the mycelium of Pisolithus tinctorius. • Purification of SDS-insoluble proteins from the mycelium of P. tinctorius showed the presence of a 13 kDa polypeptide with properties of class I hydrophobin. • Polyconal antibodies were raised against a recombinant HYDPt-1 polypeptide, and these were used for immunofluorescence-coupled transmission electron microscopy. • HYDPt-1 is a cell wall protein located at the surface of the hyphae with no preferential accumulation in the fungal cells of the different tissues of the ectomycorrhiza (i.e. extraradical hyphae, mantle or Hartig Net).
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cell wall proteins of the ectomycorrhizal basidiomycete pisolithus tinctorius identification function and expression in symbiosis
Fungal Genetics and Biology, 1999Co-Authors: Francis Martin, Catherine Voiblet, D. Carvalho, Paola Bonfante, Raffaella Balestrini, P. Laurent, Denis TaguAbstract:Abstract Specific cell–cell and cell–substrate interactions direct the growth of ectomycorrhizal fungi to their host root targets. These elaborate mechanisms lead to the differentiation of distinct multihyphal structures, the mantle, and the Hartig Net. In the ectomycorrhizal basidiomycete Pisolithus tinctorius, the use of two-dimensional gel electrophoresis, immunocytochemical microscopy, and RNA blot analysis has demonstrated the differential expression of cell wall proteins (CWPs), such as hydrophobins, adhesins, and mannoproteins, during symbiotic interaction. In other fungi, these CWPs have been suggested to play a role in hyphae aggregation, intracellular signaling cascades, and cytoskeletal changes. The recent cloning of the genes for several of these CWPs in P. tinctorius allows us to address their function in symbiosis. This review summarizes our knowledge of CWPs in P. tinctorius and considers parallels with other biotrophic fungi as a possible framework for future work.
W. G. Allaway - One of the best experts on this subject based on the ideXlab platform.
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apoplasmic barriers and their significance in the exodermis and sheath of eucalyptus pilularis pisolithus tinctorius ectomycorrhizas
New Phytologist, 2000Co-Authors: Peter A Vesk, Annelaure Markovina, Anne E. Ashford, W. G. AllawayAbstract:The apoplasmic permeability of ectomycorrhizal roots of intact Eucalyptus pilularis seedlings infected with Pisolithus tinctorius on aseptic agar plates was examined using the nonbinding fluorochrome 8-hydroxypyrene-1,3,6-trisulphonate and lanthanum ions in conjunction with anhydrous freeze substitution and dry sectioning. Most mycorrhizas formed in the air above the agar surface, and in these the sheath rapidly became nonwettable and impermeable to the fluorochrome but was nevertheless permeable to lanthanum ions. In a few mycorrhizas which developed in contact with the agar the sheath remained permeable to both tracers when fully developed. This increased hydrophobicity of the sheath in mycorrhizas in the air above the agar surface might be explained by deposition of hydrophobins, but nevertheless it still allows an apoplasmic pathway for radial movement of ions. Regardless of their sheath permeation both apoplasmic tracers were always found throughout the Hartig Net and were arrested at the Casparian bands and suberin lamellae of the exodermis. It is concluded that the fluorochrome must have moved longitudinally along the Hartig Net which is a region of higher permeability than the sheath. Casparian bands in the exodermis of ectomycorrhizal roots have similar properties to those in nonmycorrhizal roots in excluding solutes and their exclusion of lanthanum ions indicates that they are not permeable to ions. The data do not support the concept of a totally sealed apoplasmic exchange compartment, but the differential permeability suggests that the sheath might allow radial transfer of ions but block loss of sugars and organic molecules of similar size.
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dispersed polyphosphate in fungal vacuoles in eucalyptus pilularis pisolithus tinctorius ectomycorrhizas
Fungal Genetics and Biology, 1999Co-Authors: A E Ashford, Peter A Vesk, Annelaure Markovina, David A Orlovich, W. G. AllawayAbstract:Ectomycorrhizas produced between Pisolithus tinctorius and Eucalyptus pilularis under axenic conditions were rapidly frozen, freeze-substituted in tetrahydrofuran and embedded anhydrously, and dry-sectioned for X-ray microanalysis. The vacuoles of the sheath and Hartig Net hyphae were rich in phosphorus and potassium. They also contained sulfur and variable amounts of chlorine. In anhydrously processed freeze-substituted mycorrhizas, dispersed electron-opaque material filled the fungal vacuoles. X-ray maps indicated that P was distributed evenly throughout the entire vacuole profile and was not concentrated in spherical bodies or subregions of the vacuole. There were no electron-opaque granules surrounded by electron-lucent areas, such as are commonly seen in chemically fixed material. The fungal vacuoles were also rich in K, which similarly gave a signal from the entire vacuolar profile. Such P-rich vacuoles occurred in both the mycorrhizal sheath and Hartig Net hyphae. Stained sections of ether-acrolein freeze-substituted mycorrhizas also showed only dispersed material in the fungal vacuoles as, in most cases, did acetone-osmium freeze-substituted material. Precipitation of metachromatic granules by ethanol suggested that large amounts of polyphosphate are stored in these regions under the conditions of our experiments, as well as in the tips of actively growing hyphae of the same fungus. The higher plant vacuoles of ectomycorrhizas gave a much lower signal for K, and P was barely detectable. Much more K was located in the vacuoles of the root exodermal cells than in epidermal cells. The analysis of element distribution between the vacuole and cytoplasm in root cells agrees well with that found for other plant species using other techniques. We conclude that polyphosphate is indeed present in the vacuoles of the fungal cells of these ectomycorrhizas, but that in vivo it is in a dispersed form, not in granules.
Erika Kothe - One of the best experts on this subject based on the ideXlab platform.
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dehydrogenase genes in the ectomycorrhizal fungus tricholoma vaccinum a role for ald1 in mycorrhizal symbiosis
Journal of Basic Microbiology, 2016Co-Authors: Catarina Henke, Erika Kothe, Elkemartina Jung, Annekatrin Voit, Katrin KrauseAbstract:: Ectomycorrhizal symbiosis is important for forest ecosystem functioning with tree-fungal cooperation increasing performance and countering stress conditions. Aldehyde dehydrogenases (ALDHs) are key enzymes for detoxification and thus may play a role in stress response of the symbiotic association. With this focus, eight dehydrogenases, Ald1 through Ald7 and TyrA, of the ectomycorrhizal basidiomycete Tricholoma vaccinum were characterized and phylogeNetically investigated. Functional analysis was performed through differential expression analysis by feeding different, environmentally important substances. A strong effect of indole-3-acetic acid (IAA) was identified, linking mycorrhiza formation and auxin signaling between the symbiosis partners. We investigated ald1 overexpressing strains for performance in mycorrhiza with the host tree spruce (Picea abies) and observed an increased width of the apoplast, accommodating the Hartig' Net hyphae of the T. vaccinum over-expressing transformants. The results support a role for Ald1 in ectomycorrhiza formation and underline functional differentiation within fungal aldehyde dehydrogenases in the family 1 of ALDHs.
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biosynthesis and secretion of indole 3 acetic acid and its morphological effects on tricholoma vaccinum spruce ectomycorrhiza
Applied and Environmental Microbiology, 2015Co-Authors: Katrin Krause, Catarina Henke, Theodore Asiimwe, Andrea Ulbricht, Sandra Klemmer, Doreen Schachtschabel, Wilhelm Boland, Erika KotheAbstract:ABSTRACT Fungus-derived indole-3-acetic acid (IAA), which is involved in development of ectomycorrhiza, affects both partners, i.e., the tree and the fungus. The biosynthesis pathway, excretion from fungal hyphae, the induction of branching in fungal cultures, and enhanced Hartig Net formation in mycorrhiza were shown. Gene expression studies, incorporation of labeled compounds into IAA, heterologous expression of a transporter, and bioinformatics were applied to study the effect of IAA on fungal morphogenesis and on ectomycorrhiza. Tricholoma vaccinum produces IAA from tryptophan via indole-3-pyruvate, with the last step of this biosynthetic pathway being catalyzed by an aldehyde dehydrogenase. The gene ald1 was found to be highly expressed in ectomycorrhiza and induced by indole-3-acetaldehyde. The export of IAA from fungal cells is supported by the multidrug and toxic extrusion (MATE) transporter Mte1 found in T. vaccinum. The addition of IAA and its precursors induced elongated cells and hyphal ramification of mycorrhizal fungi; in contrast, in saprobic fungi such as Schizophyllum commune, IAA did not induce morphogeNetic changes. Mycorrhiza responded by increasing its Hartig Net formation. The IAA of fungal origin acts as a diffusible signal, influencing root colonization and increasing Hartig Net formation in ectomycorrhiza.
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Hartig Net formation of tricholoma vaccinum spruce ectomycorrhiza in hydroponic cultures
Environmental Science and Pollution Research, 2015Co-Authors: Catarina Henke, Elkemartina Jung, Erika KotheAbstract:For re-forestation of metal-contaminated land, ectomycorrhizal trees may provide a solution. Hence, the study of the interaction is necessary to allow for comprehensive understanding of the mutually symbiotic features. On a structural level, hyphal mantle and the Hartig’ Net formed in the root apoplast are essential for plant protection and mycorrhizal functioning. As a model, we used the basidiomycete Tricholoma vaccinum and its host spruce (Picea abies). Using an optimized hydroponic cultivation system, both features could be visualized and lower stress response of the tree was obtained in non-challenged cultivation. Larger spaces in the apoplasts could be shown with high statistical significance. The easy accessibility will allow to address metal stress or molecular responses in both partners. Additionally, the proposed cultivation system will enable for other experimental applications like addressing flooding, biological interactions with helper bacteria, chemical signaling, or other biotic or abiotic challenges relevant in the natural habitat.
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a hydrophobin accumulated in the Hartig Net of ectomycorrhiza formed between tricholoma terreum and its compatible host tree is missing in an incompatible association
Journal of applied botany, 2000Co-Authors: A Mankel, Katrin Krause, M Genenger, G Kost, Erika KotheAbstract:The host specificity of Tricholoma species in ectomycorrhiza formation was exploited to show specific accumulation of hydrophobin protein using polyclonal antiserum raised against the hydrophobin Sc3 from the homobasidiomycete Schizophyllum commune. This small, hydrophobic protein is found in cell walls of monokaryotic aerial mycelium of S. commune. In immunofluoresence studies of in vitro synthesized ectomycorrhiza of Tricholoma terreum with pine and fir a specific accumulation in the Hartig' Net could be shown in the compatible interaction of T. terreum with Pinus sylvestris, while the incompatible interaction of T terreum with Picea abies gave no evidence of hydrophobin accumulation. The hyphal mantle was stained in both cases. A role for the Tricholoma hydrophobin in Hartig' Net assembly and cell/cell contact of both hyphal/hyphal contact and contact of hyphae to the lignin containing plant cell wall is discussed.
