Recherche et caractérisation chez le haricot commun de gènes dont l'expression est modulée dans les racines lors d'un stress hydrique

La sécheresse constitue une des contraintes environnementales majeures à la production de haricot commun (Phaseolus vulgaris L.). La réponse des plantes au déficit hydrique et la mise en place de mécanismes de tolérance passent par la régulation d'un grand nombre de gènes, mais seule une douzaine de ces gènes sont connus chez le haricot. L'objectif de cette thèse a été d'isoler des gènes, dont l'expression est modulée en réponse à un déficit hydrique dans les racines de haricot. Un système expérimental a été mis au point, basé sur la déshydratation de plantes cultivées en aéroponie. Parmi les 1200 ADNc observés par differential display RT-PCR, 8,7 % sont régulés lors du stress dans les racines. Quarante-sept ADNc partiels ont ainsi été caractérisés qui correspondent tous, sauf un, à des gènes nouveaux chez le haricot. Vingt d'entre eux, nommés PvDR pour Phaseolus vulgaris Dehydration-Responsive, présentent une modulation d'expression précoce, c'est à dire antérieure aux modifications du statut hydrique et à la biosynthèse d'acide abscissique (ABA). D'après les comparaisons de séquence, ces ADNc ont été classés dans des mécanismes comme la division cellulaire, le contrôle de l'expression génique ou encore le turn-over des lipides et des protéines. La caractérisation des familles multigéniques correspondant à deux ADNc PvDR, codant pour un facteur d'initiation de la traduction EIF4A et une HSP/HSC70 (protéine de choc chaud de 70 kDa) a été réalisée sur des expériences de déshydratation, de chocs thermiques et d'un traitement à l'ABA. Enfin, un nouveau type de transporteur chez les plantes, similaire aux transporteurs de cations organiques des animaux, a été isolé. Les transcrits correspondants, quasi- indétectables dans les feuilles, sont localisés au niveau du phloème dans les racines et les tiges. Son profil d'expression lors de la déshydratation puis de la réhydratation des racines suggère une implication réelle de ce transporteur dans la réponse au déficit hydrique.Water deficit is one of the major constraints to the productivity of common bean (Phaseolus vulgaris L.). To overcome this limitation and improve production efficiency, more drought-tolerant plants must be developed. Plants respond to drought through transcriptional modulations of a certain number of genes. Few of them have been described for bean. The aim of this work was to characterize genes differentially expressed in response to water stress in P. vulgaris roots. Plants growing in an aeroponic system were submitted to dehydration by the arrest of spraying. Among 1200 cDNAs observed in DDRT-PCR experiments, 8,7% showed specific dehydration-responsive patterns in roots. Forty-seven partial cDNAs were further analyzed. All except one correspond to new genes in bean. Twenty cDNAs, named PvDR for Phaseolus vulgaris Dehydration Responsive, are regulated by dehydration before the significant change of root water status and the ABA synthesis. Based on sequence homologies, the corresponding genes were classified as involved in cellular division, control of gene expression as well as turn-over of lipids or proteins. Two multigenic families corresponding to a translation initiation factor EIF4A and a heat shock protein HSP70 have been analyzed in detail. Expression analyses of the two PveIF4A and three genes coding for HSP/HSC70 cytosolic isoforms showed variable responses to dehydration, heat shock and ABA. A new type of plant transporter was isolated corresponding to organic cation transporters in animals. Specific transcripts were detected in phloem cells of roots and stems. Its expression profile during a dehydration / rehydration cycle, strongly suggests an implication of this transporter in the response to water stress.ORSAY-PARIS 11-BU Sciences (914712101) / SudocSudocFranceF

Noncoding RNAs, emerging regulators in root endosymbioses

Endosymbiosis interactions allow plants to grow in nutrient deficient soil environments. The arbuscular mycorrhizal (AM) symbiosis is an ancestral interaction between land plants and fungi, whereas nitrogen-fixing symbioses are highly specific for certain plants, notably major crop legumes. The signaling pathways triggered by specific lipochitooligosaccharide molecules involved in these interactions have common components that also overlap with plant root development. These pathways include receptor-like kinases, transcription factors (TFs), and various intermediate signaling effectors, including noncoding (nc)RNAs. These latter molecules have emerged as major regulators of gene expression and small ncRNAs, composed of micro (mi)RNAs and small interfering (si)RNAs, are known to control gene expression at transcriptional (chromatin) or posttranscriptional levels. In this review, we describe exciting recent data connecting variants of conserved si/miRNAs with the regulation of TFs, such as NSP2, NFY-A1, auxin-response factors, and AP2-like proteins, known to be involved in symbiosis. The link between hormonal regulations and these si- and miRNA-TF nodes is proposed in a model in which different feedback loops or regulations controlling endosymbiosis signaling are integrated. The diversity and emerging regulatory networks of young legume miRNAs are also highlighted

Transcriptional and post-transcriptional regulation of a NAC1 transcription factor in Medicago truncatula roots

Legume roots develop two types of lateral organs, lateral roots and nodules. Nodules develop as a result of a symbiotic interaction with rhizobia and provide a niche for the bacteria to fix atmospheric nitrogen for the plant. The Arabidopsis NAC1 transcription factor is involved in lateral root formation, and is regulated post-transcriptionally by miRNA164 and by SINAT5-dependent ubiquitination. We analyzed in Medicago truncatula the role of the closest NAC1 homolog in lateral root formation and in nodulation. MtNAC1 shows a different expression pattern in response to auxin than its Arabidopsis homolog and no changes in lateral root number or nodulation were observed in plants affected in MtNAC1 expression. In addition, no interaction was found with SINA E3 ligases, suggesting that post-translational regulation of MtNAC1 does not occur in M. truncatula. Similar to what was found in Arabidopsis, a conserved miR164 target site was retrieved in MtNAC1, which reduced protein accumulation of a GFP-miR164 sensor. Furthermore, miR164 and MtNAC1 show an overlapping expression pattern in symbiotic nodules, and overexpression of this miRNA led to a reduction in nodule number. This work suggests that regulatory pathways controlling a conserved transcription factor are complex and divergent between M. truncatula and Arabidopsis

miR396 affects mycorrhization and root meristem activity in the legume Medicago truncatula

The root system is crucial for acquisition of resources from the soil. In legumes, the efficiency of mineral and water uptake by the roots may be reinforced due to establishment of symbiotic relationships with mycorrhizal fungi and interactions with soil rhizobia. Here, we investigated the role of miR396 in regulating the architecture of the root system and in symbiotic interactions in the model legume Medicago truncatula. Analyses with promoter–GUS fusions suggested that the mtr-miR396a and miR396b genes are highly expressed in root tips, preferentially in the transition zone, and display distinct expression profiles during lateral root and nodule development. Transgenic roots of composite plants that over-express the miR396b precursor showed lower expression of six growth-regulating factor genes (MtGRF) and two bHLH79-like target genes, as well as reduced growth and mycorrhizal associations. miR396 inactivation by mimicry caused contrasting tendencies, with increased target expression, higher root biomass and more efficient colonization by arbuscular mycorrhizal fungi. In contrast to MtbHLH79, repression of three GRF targets by RNA interference severely impaired root growth. Early activation of mtr-miR396b, concomitant with post-transcriptional repression of MtGRF5 expression, was also observed in response to exogenous brassinosteroids. Growth limitation in miR396 over-expressing roots correlated with a reduction in cell-cycle gene expression and the number of dividing cells in the root apical meristem. These results link the miR396 network to the regulation of root growth and mycorrhizal associations in plants.Fil: Bazin, Jeremie. Centre National de la Recherche Scientifique. Institut des Sciences du Veg etal; Francia. Universite Paris Diderot - Paris 7; FranciaFil: Khan. Ghazanfar Abbas. Centre National de la Recherche Scientifique. Institut des Sciences du Veg etal; FranciaFil: Combier, Jean Philippe. Laboratoire de Recherche en Sciences Veg etales; Francia. Centre National de la Recherche Scientifique; FranciaFil: Bustos Sanmamed, Maria del Pilar. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Rosario. Instituto de Biología Molecular y Celular de Rosario; Argentina. Centre National de la Recherche Scientifique. Institut des Sciences du Veg etal; FranciaFil: Debernardi, Juan Manuel. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Rosario. Instituto de Biología Molecular y Celular de Rosario; ArgentinaFil: Rodriguez, Ramiro. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Rosario. Instituto de Biología Molecular y Celular de Rosario; ArgentinaFil: Sorin, Celine. Centre National de la Recherche Scientifique. Institut des Sciences du Veg etal; Francia. Universite Paris Diderot - Paris 7; FranciaFil: Palatnik, Javier Fernando. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Rosario. Instituto de Biología Molecular y Celular de Rosario; ArgentinaFil: Hartmann, Caroline. Centre National de la Recherche Scientifique. Institut des Sciences du Veg etal; Francia. Universite Paris Diderot - Paris 7; FranciaFil: Crespi, Martin. Centre National de la Recherche Scientifique. Institut des Sciences du Veg etal; FranciaFil: Lelandais Briere, Christine. Centre National de la Recherche Scientifique. Institut des Sciences du Veg etal; Francia. Universite Paris Diderot - Paris 7; Franci

Long ncRNAs and epigenetic regulations are attractive candidate elements for the orchestration of symbiotic gene expression in M. truncatula.

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