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• In an attempt to advance the understanding of ectomycorrhizal symbiosis, which improves mineral nutrition and resistance to abiotic and biotic stresses of most woody plants from temperate and boreal regions, we produced a cDNA library of the ectomycorrhizal fungusHebeloma cylindrosporumin a yeast expression vector. The library was used for functional complementation cloning and for sequencing of 4036 expressed sequence tags (ESTs).• Expression of the library in a yeastade2andtrp1mutant strain allowed cloning of twoHebelomagenes,HcADE2andHcTRP1by functional complementation. This experiment highlighted both the efficiency of the complementation strategy and the quality of the library.• All sequence data available onHebeloma(4596) were analysed and contiged. The whole EST resource is accessibleviaa web interface, which allows searching and navigation through clusters and tentative contigs (TCs).• Analysis of the database revealed a low level of counterparts already identified in other ectomycorrhizal fungi. For example, it allowed identification of a large set of membrane transporters, such as phosphate, potassium, sulphate or micronutrient transporters. Thus, this EST resource provides a valuable tool for research on ectomycorrhiza and symbiotic interactions.New Phytologist(2004)doi: 10.1111/j.1469-8137.2004.01185.x© New Phytologist(2004) [ABSTRACT FROM AUTHOR]

Ectomycorrhizal symbiosis between fungi and woody plants strongly improves plant mineral nutrition and constitutes a major biological process in natural ecosystems. Molecular identification and functional characterization of fungal transport systems involved in nutrient uptake are crucial steps toward understanding the improvement of plant nutrition and the symbiotic relationship itself. In the present report a transporter belonging to the Trk family is identified in the model ectomycorrhizal fungus Hebeloma cylindrosporum and named HcTrk1. The Trk family is still poorly characterized, although it plays crucial roles in K+ transport in yeasts and filamentous fungi. In Saccharomyces cerevisiae K+ uptake is mainly dependent on the activity of Trk transporters thought to mediate H+:K+ symport. The ectomycorrhizal HcTrk1 transporter was functional when expressed in Xenopus oocytes, enabling the first electrophysiological characterization of a transporter from the Trk family. HcTrk1 mediates instantaneously activating inwardly rectifying currents, is permeable to both K+ and Na+, and displays channel-like functional properties. The whole set of data and particularly a phenomenon reminiscent of the anomalous mole fraction effect suggest that the transport does not occur according to the classical alternating access model. Permeation appears to occur through a single-file pore, where interactions between Na+ and K+ might result in Na+:K+ co-transport activity. HcTrk1 is expressed in external hyphae that explore the soil when the fungus grows in symbiotic condition. Thus, it could play a major role in both the K+ and Na+ nutrition of the fungus (and of the plant) in nutrient-poor soils. [ABSTRACT FROM AUTHOR]