Réponses moléculaires du champignon ectomycorhizien Paxillus involutus aux métaux et effet de la symbiose sur la croissance et la survie de Betula pendula cultivé en sol pollué

Texte intégral accessible uniquement aux membres de l'Université de LorraineMolecular mechanisms underlying the response of the ectomycorrhizal fungi Paxillus involutus towards heavy metals have been studied. The availability of a P. involutus cDNA library has allowed the use of a cDNA macroarray hybridization, with 2040 cDNAs, to identify genes whose expression was modulated by cadmium (4 % of these clones were up or down regulated by Cd). Three cDNAs (from the cDNA library) coding for a superoxide dismutase (SOD), a metallothionein (MT) and a thioredoxin (TRX) have been characterized. The SOD and the MT functions have been demontrated respectively using a SOD-deleted bacteria and a MT-deleted yeast. The study of the SOD revealed a probable peroxysomal localization and mainly a regulation at the post-translational level. The expression study of MT and TRX has been performed using 3 metals : Cd, Cu et Zn and indicated an induction by only Cd and Cu. We have also measured the synthesis of non-protein thiols in P. involutus subjected to a Cd stress and demonstrated the overproduction of glutathion, the absence of phytochelatins and the presence of a unidentified component which could be a MT. We have also studied the impact of various P. involutus strains on different aspects of his host tree (birch) physiology grown on polluted soil. We have confirmed the filter function of P. involutus, but we also observed a weak survival rate of introduced fungal strains against indigenous strains.Les mécanismes moléculaires impliqués dans la réponse du champignon ectomycorhizien Paxillus involutus aux métaux ont été étudiés. La disponibilité d'une banque ADNc de ce champignon a permis l'étude de l'expression génique de 2040 clones et l'obtention d'une vision globale des gènes régulés par le Cd (environ 4 % sont régulés positivement ou négativement). Trois ADNc codant une superoxyde dismutase (SOD), une métallothionéine (MT) et une thioredoxine (TRX) ont été caractérisés. La fonction de la SOD et de la MT a été démontrée respectivement chez une bactérie SOD-déficiente et une levure MT-déficiente. Les études sur la SOD ont démontré une probable localisation peroxysomale et un niveau de régulation principalement post-traductionnel. L'étude de l'expression de la MT et de la TRX a été réalisée sur 3 métaux: Cd, Cu et Zn et a permis de montrer leur induction par le Cd et le Cu. La mesure de la synthèse des thiols non protéiques chez P. involutus soumis à un stress Cd indique la surproduction de glutathion, l'absence de synthèse de phytochélatines et la présence d'un composé pouvant être une MT. Enfin, le rôle de différentes souches de P. involutus sur les racines de bouleau cultivé sur sol pollué a été étudié. Ceci nous a permis de confirmer le rôle filtre de P. involutus vis à vis des métaux et de mettre en évidence la faible compétitivité des souches vis à vis de souches fongiques indigènes

Réponses moléculaires du champignon ectomycorhizien Paxillus involutus aux métaux et effet de la symbiose sur la croissance et la survie de Betula pendula cultivé en sol pollué

Les mécanismes moléculaires impliqués dans la réponse du champignon ectomycorhizien Paxillus involutus aux métaux ont été étudiés. La disponibilité d'une banque ADNc de ce champignon a permis l'étude de l'expression génique de 2040 clones et l'obtention d'une vision globale des gènes régulés par le Cd (environ 4 % sont régulés positivement ou négativement). Trois ADNc codant une superoxyde dismutase (SOD), une métallothionéine (MT) et une thioredoxine (TRX) ont été caractérisés. La fonction de la SOD et de la MT a été démontrée respectivement chez une bactérie SOD-déficiente et une levure MT-déficiente. Les études sur la SOD ont démontré une probable localisation peroxysomale et un niveau de régulation principalement post-traductionnel. L'étude de l'expression de la MT et de la TRX a été réalisée sur 3 métaux: Cd, Cu et Zn et a permis de montrer leur induction par le Cd et le Cu. La mesure de la synthèse des thiols non protéiques chez P. involutus soumis à un stress Cd indique la surproduction de glutathion, l'absence de synthèse de phytochélatines et la présence d'un composé pouvant être une MT. Enfin, le rôle de différentes souches de P. involutus sur les racines de bouleau cultivé sur sol pollué a été étudié. Ceci nous a permis de confirmer le rôle filtre de P. involutus vis à vis des métaux et de mettre en évidence la faible compétitivité des souches vis à vis de souches fongiques indigènes.Molecular mechanisms underlying the response of the ectomycorrhizal fungi Paxillus involutus towards heavy metals have been studied. The availability of a P. involutus cDNA library has allowed the use of a cDNA macroarray hybridization, with 2040 cDNAs, to identify genes whose expression was modulated by cadmium (4 % of these clones were up or down regulated by Cd). Three cDNAs (from the cDNA library) coding for a superoxide dismutase (SOD), a metallothionein (MT) and a thioredoxin (TRX) have been characterized. The SOD and the MT functions have been demontrated respectively using a SOD-deleted bacteria and a MT-deleted yeast. The study of the SOD revealed a probable peroxysomal localization and mainly a regulation at the post-translational level. The expression study of MT and TRX has been performed using 3 metals: Cd, Cu et Zn and indicated an induction by only Cd and Cu. We have also measured the synthesis of non-protein thiols in P. involutus subjected to a Cd stress and demonstrated the overproduction of glutathion, the absence of phytochelatins and the presence of a unidentified component which could be a MT. We have also studied the impact of various P. involutus strains on different aspects of his host tree (birch) physiology grown on polluted soil. We have confirmed the filter function of P. involutus, but we also observed a weak survival rate of introduced fungal strains against indigenous strains.NANCY1-SCD Sciences & Techniques (545782101) / SudocSudocFranceF

Transcriptomic responses to cadmium in the ectomycorrhizal fungus Paxillus involutus

AbstractThe molecular mechanisms underlying the response of ectomycorrhizal fungi to heavy metals in general and cadmium in particular remain poorly understood. We screened 2040 arrayed cDNAs of the ectomycorrhizal fungus Paxillus involutus to identify cadmium-responsive genes by using differential hybridization. Forty nine (2.4%) of the 2040 cDNAs were differentially expressed, among which transcripts coding a laccase, an aconitase, and a metallothionein were upregulated by 3.9-, 3.7- and 2.8-fold, respectively, whereas genes coding hydrophobins and threonine dehydratase were strongly downregulated. Our results suggest that complexation of cadmium by phenolic compounds, or by complexing peptides such as metallothioneins, is probably key determinant of the cellular response to cadmium in P. involutus. In addition, the present study suggests that the synthesis of hydrophobins may be efficiently reduced, thus redirecting Cys to the manufacture of Cys-enriched compounds

Extracellular and cellular mechanisms sustaining metal tolerance in ectomycorrhizal fungi

International audienceThis 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

Metal induction of a Paxillus involutus metallothionein and its heterologous expression in Hebeloma cylindrosporum

International audience• 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

Identification of genes involved in Cd hypertolerance by using crosses between Arabidopsis halleri and Arabidopsis lyrata ssp. petraea

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Transcriptome analysis of roots from crosses between Arabidopsis halleri ,a Zn, Cd hyperaccumulator and Arabidopsis petraea, a non-tolerant, non-accumulator relative

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Comparative cDNA-AFLP analysis of roots from crosses between Arabidopsis halleri, a Zn, Cd hyperaccumulator and Arabidopsis petraea, a non-tolerant, non-accumulator relative

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Identification of genes involved in Cd hypertolerance by using crosses between A. halleri and A. lyrata ssp. petraea

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A major QTL for Cd tolerance in Arabidopsis halleri co-localizes with HMA4, a gene encoding a heavy metal ATPase

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