ABSTRACT
* Metallothioneins are small polypeptides involved in metal tolerance of many eukaryotes. Here we characterized the Pimt1 gene, coding for a metallothionein from the ectomycorrhizal fungus Paxillus involutus. * Expression of Pimt1 in P. involutus under metal stress conditions was measured by northern blot and RT-PCR analyses. The full-length cDNA was used to perform functional complementation in yeast mutant strains and agrotransformation of Hebeloma cylindrosporum. * Heterologous expression in yeast showed that PiMT1 was able to complement the hypersensitivity of mutant strains to cadmium (Cd) and copper (Cu), but not to zinc (Zn). Transcripts were almost undetectable under control conditions, whereas Cu and Cd, but not Zn, strongly induced Pimt1 expression in P. involutus. Constitutive overexpression of Pimt1 in H. cylindrosporum conferred a higher copper tolerance. * The present study identified PiMT1 as a potential determinant in the response of mycorrhizal fungi to Cu and Cd stress. Additionally, we demonstrated the usefulness of mycorrhizal fungi transformation using Agrobacterium technology to approach gene function.
Subject(s)
Agaricales/genetics , Basidiomycota/genetics , Fungal Proteins/genetics , Metallothionein/genetics , Metals/metabolism , Agaricales/metabolism , Amino Acid Sequence , Basidiomycota/metabolism , Cloning, Molecular , DNA, Complementary , Fungal Proteins/biosynthesis , Fungal Proteins/chemistry , Gene Expression Regulation, Fungal , Genetic Complementation Test , Metallothionein/biosynthesis , Metallothionein/chemistry , Molecular Sequence Data , Mycorrhizae/genetics , Mycorrhizae/metabolism , Organisms, Genetically Modified , Rhizobium/genetics , Saccharomyces cerevisiae/genetics , Sequence Alignment , Sequence Analysis, Protein , Transcription, Genetic , Transformation, GeneticABSTRACT
This review focuses on recent evidence that identifies potential extracellular and cellular mechanisms that may be involved in the tolerance of ectomycorrhizal fungi to excess metals in their environment. It appears likely that mechanisms described in the nonmycorrhizal fungal species are used in the ectomycorrhizal fungi as well. These include mechanisms that reduce uptake of metals into the cytosol by extracellular chelation through extruded ligands and binding onto cell-wall components. Intracellular chelation of metals in the cytosol by a range of ligands (glutathione, metallothioneins), or increased efflux from the cytosol out of the cell or into sequestering compartments are also key mechanisms conferring tolerance. Free-radical scavenging capacities through the activity of superoxide dismutase or production of glutathione add another line of defence against the toxic effect of metals.
