Les fluctuations du taux de calcium (Ca2+) intracellulaire sont impliquées dans les événements de signalisation et de régulation de différents processus cellulaires. Alors que le role du Ca2+ dans la réponse des plantes lors des interactions mycorhiziennes à arbuscules (MA) interactions est bien documentée, il n’existe aucune information concernant la régulation ou le rôle de ce messager secondaire chez le symbiote fongique. La base moléculaire de l'homéostasie calcique fongique dans la symbiose MA a été analysée en étudiant l'expression de gènes fongiques liés au Ca2+. Dans un premier temps, des gènes de G. mosseae codant putativement pour une protéine kinase-like MAP3k (Gm2) et une P-type ATPase (Gm152) ont été étudiés. L’expression des deux gènes est stimulée par les exudats racinaires d’A. sinicum, suggérant un rôle dans les interactions précoces avant l'établissement de la symbiose. L’obtention de la séquence d'ADNc pleine longueur de Gm152 a confirmé son identité. Une étude plus approfondie du rôle de Ca2+ dans les processus fongiques impliqués dans la symbiose MA a été réalisée chez G. intraradices. L'expression de sept gènes fongiques encodant six protéines de transport membranaire calcique et une protéine kinase nucléaire, sélectionnés du séquençage transcriptomique du G. intraradices, était stimulée lors de la colonisation des racines de M. truncatula type sauvage (lignée J5) mais pas chez le mutant non-mycorhizienne dmi3/Mtsym13. La cartographie par microdissection laser des transcrits des gènes fongiques a indiqué une activation différentielle dans les arbuscules et/ou dans hyphes intercellulaires. Les variations tempo-spatiales de l'expression des gènes fongiques suggèrent des roles différents dans le développement ou le fonctionnement de la symbiose MA. L’ADNc pleine longueur a été obtenue de trois gènes de G. intraradices encodant un PMR1-like réticulum endoplasmique ATPase, un VCX1-like transporteur ionique vacuolaire et un CCaMK nucléaire pour des analyses fonctionnelles chez la levure afin de mieux comprendre leur rôle dans la symbiose MA. Les mécanismes par lesquels les protéines liées au Ca2+ pourraient jouer un rôle chez G. intraradices dans la mobilisation et la perception du messager secondaire au cours des interactions MA sont discutés / Fluctuations in intracellular (Ca2+) calcium levels generate signaling events and regulate different cellular processes. Whilst the implication of Ca2+ in plant cell responses during arbuscular mycorrhiza (AM) interactions is well documented, nothing is known about the regulation or role of this secondary meesenger in the fungal symbiont. The molecular basis of fungal calcium homeostasis in the AM symbiosis was analyzed by investigating the expression of Ca2+-related fungal genes. In a first study, G. mosseae genes putatively encoding a MAP3k-like protein kinase (Gm2) and a P-type ATPase (Gm152) were investigated. Both Ca2+-related genes were up-regulated by A. sinicum root exudates, suggesting a role in early interactions prior to symbiosis establishment. The full-length cDNA sequence of Gm152 obtained from germinating spores of G. mosseae confirmed its identity. The role of Ca2+ in fungal processes leading to establishment of an AM symbiosis was investigated in more detail in G. intraradices-M. truncatula interactions. Enhanced expression of genes encoding six membrane transport proteins and one nuclear protein kinase, selected from the G. intraradices transcriptome database, was related to colonization of wild-type M. truncatula (line J5) roots and not observed with the mycorrhiza-resistant mutant dmi3/Mtsym13. Laser microdissection mapping of transcripts indicated that the Ca2+-related G. intraradices genes were differentially up-regulated in arbuscules and/or in intercellular hyphae. The tempo-spatial variations in fungal gene expression suggest different roles in the development or functioning of the AM symbiosis. Full-length cDNA of three G. intraradices genes putatively encoding a PMR-like endoplasmic reticulum P-type ATPase, a VCX1-like vacuolar Ca2+ ion transporter and a nuclear CCaMK were obtained for functional analyses in yeast mutants to gain insight into their role in the mycorrhizal symbiosis. Possible mechanisms are discussed in which Ca2+-related proteins of G. intraradices may play a role in the mobilization and perception of the intracellular messenger by the AM fungus during symbiotic interactions with host roots

In plants, long distance transport of sugars from photosynthetic source leaves to sink organs comprises different crucial steps depending on the species and organ types. Sucrose, the main carbohydrate for long distance transport is synthesized in the mesophyll and then loaded into the phloem. After long distance transport through the phloem vessels, sucrose is finally unloaded towards sink organs. Alternatively, sugar can also be transferred to non‐plant sinks and plant colonization by heterotrophic organisms increases the sink strength and creates an additional sugar demand for the host plant. These sugar fluxes are coordinated by transport systems. Main sugar transporters in plants comprise sucrose (SUTs) and monosaccharide (MSTs) transporters which constitute key components for carbon partitioning at the whole plant level and in interactions with fungi. Although complete SUTs and MSTs gene families have been identified from the reference Dicot Arabidopsis thaliana and Monocot rice (Oriza sativa), sugar transporter families of the leguminous plant Medicago truncatula, which represents a widely used model for studying plant-fungal interactions in arbuscular mycorrhiza (AM), have not yet been investigated.With the recent completion of the M. truncatula genome sequencing as well as the release of transcriptomic databases, monosaccharide and sucrose transporter families of M. truncatula were identified and now comprise 62 MtMSTs and 6 MtSUTs. I focused on the study of the newly identified MtSUTs at a full family scale; phylogenetic analyses showed that the 6 members of the MtSUT family distributed in all three Dicotyledonous SUT clades; they were named upon phylogenetic grouping into particular clades: MtSUT1-1, MtSUT1-2, MtSUT1-3, MtSUT2, MtSUT4-1 and MtSUT4-2. Functional analyses by yeast complementation and expression profiles obtained by quantitative RT-PCR revealed that MtSUT1-1 and MtSUT4-1 are H+/sucrose symporters and represent key members of the MtSUT family. Conservation of transport capacity between orthologous leguminous proteins, expression profiles and subcellular localization compared to previously characterized plant SUTs indicate that MtSUT1-1 is the main protein involved in phloem loading in source leaves whilst MtSUT4-1 mediates vacuolar sucrose export for remobilization of intracellular reserve.The AM symbiosis between plants and fungi from the phylum Glomeromycota is characterized by trophic exchanges between the two partners. The fungus supplies the autotrophic host with nutrients and thereby promotes plant growth. In return, the host plant provides photosynthate (sugars) to the heterotrophic symbiont. Here, sugar fluxes from plant source leaves towards colonized sink roots in the association between the model leguminous plant M. truncatula and the arbuscular mycorrhizal fungus (AMF) Glomus intraradices were investigated. Sugar transporter candidates from both the plant and fungal partners presenting differential expression profiles using available transcriptomic tools were pinpointed. Gene expression profiles of MtSUTs and sugar quantification analyses upon high and low phosphorus nutrient supply and inoculation by the AMF suggest a mycorrhiza-driven stronger sink in AM roots with a fine-tuning regulation of MtSUT gene expression. Conserved regulation patterns were observed for orthologous SUTs in response to colonization by glomeromycotan fungi.In parallel, a non-targeted strategy consisting in the development of a M. truncatula - G. intraradices expression library suitable for yeast functional complementation and screening of symbiotic marker genes, similar to the approach that led to the identification of the first glomeromycotan hexose transporter (GpMST1), has been developed in this study. [...]

Fluctuations in intracellular (Ca2+) calcium levels generate signaling events and regulate different cellular processes. Whilst the implication of Ca2+ in plant cell responses during arbuscular mycorrhiza (AM) interactions is well documented, nothing is known about the regulation or role of this secondary meesenger in the fungal symbiont. The molecular basis of fungal calcium homeostasis in the AM symbiosis was analyzed by investigating the expression of Ca2+-related fungal genes. In a first study, G. mosseae genes putatively encoding a MAP3k-like protein kinase (Gm2) and a P-type ATPase (Gm152) were investigated. Both Ca2+-related genes were up-regulated by A. sinicum root exudates, suggesting a role in early interactions prior to symbiosis establishment. The full-length cDNA sequence of Gm152 obtained from germinating spores of G. mosseae confirmed its identity. The role of Ca2+ in fungal processes leading to establishment of an AM symbiosis was investigated in more detail in G. intraradices-M. truncatula interactions. Enhanced expression of genes encoding six membrane transport proteins and one nuclear protein kinase, selected from the G. intraradices transcriptome database, was related to colonization of wild-type M. truncatula (line J5) roots and not observed with the mycorrhiza-resistant mutant dmi3/Mtsym13. Laser microdissection mapping of transcripts indicated that the Ca2+-related G. intraradices genes were differentially up-regulated in arbuscules and/or in intercellular hyphae. The tempo-spatial variations in fungal gene expression suggest different roles in the development or functioning of the AM symbiosis. Full-length cDNA of three G. intraradices genes putatively encoding a PMR-like endoplasmic reticulum P-type ATPase, a VCX1-like vacuolar Ca2+ ion transporter and a nuclear CCaMK were obtained for functional analyses in yeast mutants to gain insight into their role in the mycorrhizal symbiosis. Possible mechanisms are discussed in which Ca2+-related proteins of G. intraradices may play a role in the mobilization and perception of the intracellular messenger by the AM fungus during symbiotic interactions with host roots