Functional characterization of the rice transcription factors OsMADS25 and OsMADS26 in regard to development and biotic and abiotic stresses resistance

Le riz (Oryza sativa L) est la source principale d'alimentation pour plus de la moitié de la population mondiale (Khush, 2005). La production de riz devrait augmenter de plus de 40% en 2030 pour satisfaire la demande de croissance de la population. Chaque année, environ 25% de la production est perdue à cause des insectes ravageurs, des maladies et des mauvaises herbes (Khus, 2005). Des pertes semblables sont dues aux stress abiotiques comme la sécheresse. L'objectif de mon travail de thèse a consisté à étudier la fonction de deux facteurs de transcription (FT) à boîte MADS OsMADS25 et OsMADS26 dans la réponse aux stress ou dans le développement. Pour cela, j'ai généré des lignées de riz surexprimant les ADNc codant ces FT et aussi des lignées interférées pour le gène OsMADS26 en utilisant deux GST différentes pour induire de l'ARN interférence destiné à détruire les ARNm OsMADS26. Dans le cas du gène OsMADS25 qui appartient à un groupe de cinq gènes phylogénétiquement proches, j'ai généré des plantes exprimant la protéine OsMADS25 fusionnée avec le motif répresseur dominant de la transcription EAR. Les lignées T2 exprimant le FT OsMADS25 fusionné au motif EAR présentent un phénotype semblable à celui d'une lignée d'insertion de TDNA dans ce gène. Ces plantes sont caractérisées par une forte réduction du nombre de leur talle et par une hauteur plus importante de la talle principale. Les plantes qui surexpriment OsMADS25 natif ne présentent pas de phénotype particulier. Ceci suggère que le gène OsMADS25 pourrait être impliqué dans la régulation du nombre de talles chez le riz bien qu'il soit exprimé au niveau de la racine. Le mode d'action du gène OsMADS25 sur le contrôle du développement des méristèmes axillaire du riz reste à préciser. Les lignées interférées OsMADS26 présentent une meilleure résistance à Magnaporthe oryzae (Mo) et à Xanthomonas oryzae pv. Oryzae (Xoo), deux principaux pathogènes du riz, et aussi une meilleure capacité de restauration après l'application d'un stress hydrique par rapport aux lignées témoin tandis que les lignées surexprimant OsMADS26 sont plus sensibles à ces stress. Les analyses de QPCR et du transcriptome que nous avons effectuées ont mis en évidence l'expression constitutive plus élevée dans les lignées interférées de plusieurs gènes de réponse aux stress biotique et abiotique. Ces résultats suggèrent que OsMADS26 pourrait être un inhibiteur général des mécanismes de défense de la plante et que les plantes interférée OsMADS26 sont dans un statut physiologique de type primed-like qui leur permettent d'être plus résistantes aux stress. Les lignées interférées pour OsMADS26 sont très peu affectées dans leur développement. Le gène OsMADS26 est donc un gène très intéressant pour les programmes d'amélioration du rizMADS-box transcription factors (TF) have been mostly characterized for their involvement of plant development such as floral organogenesis and flowering time. Some of them are involved in stress related developmental processes such as abscission, fruit ripening and senescence. Overexpression of the rice OsMADS26 TF suggested a function in stress response. Here we report that OsMADS26 interfered lines presented a better resistance against two major pathogens of rice, Xanthomonas oryzae (Xoo) and Magnaportae oryzae (Mo) and a better recovery capacity after a water stress period. Transcriptome analysis revealed that several biotic and abiotic stresses related genes were up regulated in OsMADS26 interfered lines. In addition QPCR analysis showed that the expression of a set of biotic and abiotic genes was induced when OsMADS26 interfered lines were infected by Xoo or submitted to a water stress. This indicated that OsMADS26 is a negative regulator of biotic and abiotic stress response in rice. Taking in account the data previously published that showed that inducible overexpression of OsMADS26 resulted in the activation of expression of genes involved in jasmonic acid or reactive oxygen species biosynthesis, we postulate that OsMADS26 may be a hub regulator of stress response in rice and that it may be posttranscriptional regulated to modulate negatively or positively rice response to various stresses.In addition we have shown in this thesis that an insertion mutant line disrupting the OsMADS25 gene is characterized by a reduced number of tiller. This phenotype was also obtained in transgenic lines expressing the OsMADS25 transcription factor fused with a dominant motif inhibitor of transcription. Thissuggested that OsMADS25 is involved in the control of tiller development in rice.Key words: Rice, stress, blast, tillering, MADS-box, transcription factor, OsMADS26, OsMADS25, transcriptom

Potential of the coffee endophytic Bacillus cereus sensu lato strain CCBLR15 to control the plant-parasitic nematode Radopholus duriophilus

International audienceBacillus cereus sensu lato strain CCBLR15 is highly efficient in inducing in vitro the mortality of coffee-parasitic nematodes, including Pratylenchus coffeae and Radopholus duriophilus. In the present study, this strain was further characterised to investigate (1) its taxonomic position, (2) the secreted/non-secreted nature of its nematicidal compounds, and (3) its in planta biocontrol capacity towards R. duriophilus. Firstly, analysis of 16S rRNA gene sequences and motility test indicated that CCBLR15 was closely related to B. cereus sensu stricto. Secondly, nematodes separate expositions in vitro to CCBLR15 culture filtrate and cell lysate revealed that nematicidal activity was attributed to compounds released after bacterial cell lysis. Thirdly, CCBLR15 nematode biocontrol capacity was studied using three Coffea arabica cultivars, either seed- or cutting-propagated. Plants were therefore characterised by different developmental stages, referred to as 'early' and 'advanced', with respectively 2-3 and 6-8 leaf pairs. Additionally, plants were grown in unsterilised soil, with a one-month delay between inoculations of bacteria and nematodes to assess the persistence of the nematicidal effects. The presence of R. duriophilus in roots decreased plant growth only in one cultivar whose plants were inoculated at the 'early' stage. Nonetheless, CCBLR15 pre-inoculation significantly reduced the number of nematodes in roots of all cultivars and inhibited the negative effects of nematodes in 'early'-inoculated plants. These results demonstrate the capacity of CCBLR15 to control R. duriophilus and constitute an important step before studying the persistence of its biocontrol effects on a longer period or after plants establishment in the field

Defence mechanisms associated with mycorrhiza-induced resistance in wheat against powdery mildew

International audienceTo develop a more sustainable agriculture using alternative control strategies, mechanisms involved in the biocontrol ability of the arbuscular mycorrhizal fungus Funneliformis mosseae to protect wheat against the foliar biotrophic pathogen Blumeria graminis f. sp. tritici were investigated under controlled conditions. B. graminis infection on wheat leaves was reduced by 78% in mycorrhizal plants compared with non-mycorrhizal ones (control). Wheat roots inoculated with F. mosseae revealed a systemic resistance in leaves to B. graminis, after a 6-week co-culture. Accordingly, this resistance was associated with a significant reduction of B. graminis haustorium formation in epidermal leaf cells of mycorrhizal wheat and an accumulation of phenolic compounds and H2O2 at B. graminis penetration sites. Moreover, gene expression analysis demonstrated upregulation of genes encoding for several defence markers, such as peroxidase, phenylalanine ammonia lyase, chitinase 1 and nonexpressor of pathogenesis-related proteins 1 in mycorrhizal wheat only in the absence of the pathogen. This study showed that protection of wheat obtained against B. graminis in response to mycorrhizal inoculation by F. mosseae could be interpreted as a mycorrhiza-induced resistance (MIR). Our findings also suggest that MIR-associated mechanisms impaired the B. graminis development process and corresponded to a systemic elicitation of plant defences rather than a primed state in wheat leaves