6533b827fe1ef96bd12857d4

RESEARCH PRODUCT

Déterminisme du pouvoir protecteur de Fusarium oxysporum : recherche de gènes impliqués dans l'interaction protectrice avec la tomate

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Fusarium oxysporum is a common soil borne fungus, well represented in every type of soils, throughout the world. This species includes pathogenic strains inducing severe diseases in many crops and strains able to protect a plant against the infection by a pathogenic strain. The protective strains are not only non pathogenic strains isolated from suppressive soils but also pathogenic strains applied to a non host plant. The protective capacity of these strains is mainly based on mechanisms of competition and induced resistance of the plant The main objective of this work was to identify fungal genes involved in the protective capacity of these strains and associated to the elicitation of plant defence mechanisms. The approach consisted in looking for genes differentially expressed during the interaction between tomato cell cultures and either a protective or a non protective strain of F. oxysporum. An experimental model was developed; it is based on a strain of F. oxysporum f. sp. melonis (Fom24) pathogenic on muskmelon (Fom24) and on its transposition mutant (rev157). The wild type strain has the capacity to protect flax and tomato against fusarium wilt, although rev157 has lost the protective capacity. This mutant being neither affected in its capacity to grow saprophytically, nor to colonize the plant roots, the hypothesis that it was affected in its capacity to elicitate the plant defence reactions was proposed. Indeed, when interacting with cell cultures it induced physiological events (H2O2 accumulation, cell death) different from that induced by the wild type strain Fom24. Having no a priori hypothesis regarding the functions of the genes affected by the mutation, a Rapid Subtraction Hybridization (RaSH) technique was used to identify genes differentially expressed during the interaction between tomato cell cultures and either the protective or the non protective strain of F. oxysporum. This technique enabled to identify sequences from both fungal and plant origin. Since data concerning fungal genes are scarce in Genbank, the most informative data concerned plant genes. Regarding fungi, the expression of two genes, a chitinase and a polyphenol oxydase respectively involved in the degradation of cell walls and the oxidative catalysis of phenols was analyzed by Northern blot. Regarding plant genes, no newly expressed sequence was detected, demonstrating that the protective interaction results from a differential regulation of genes already expressed during the non protective interaction between plant cells and conidia of F. oxysporum. The expression profile of 5 genes was analyzed by Northern blot and real time PCR. Two of them are involved in the plant response to pathogen infection (RIN4, chitinase), and the 3 others are involved in important functions related to metabolism and transport (ATPase, ferredoxin-NADP-reductase and porin). The low activity of the ferredoxin-NADP-reductase in the cells interacting with the protective strain can be correlated with the great of amount H2O2 produced by these cells. The most interesting result concerns the demonstration that RIN4, a gene implicated in a guard system, is involved in the interaction between cells of tomato of a susceptible variety and different strains of F. oxysporum. Depending on the genetic environment of the cell (presence or absence of resistance genes) this gene either favors the growth of the pathogen or stimulates the plant defence reactions. The gene RIN4 being down-regulated during the protective interaction, the plant defence reactions would be fully expressed.