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Mots-clésarabidopsis - Erwinia amylovora - Dickeya dadantii - Botrytis cinerea - azote - nutrition - stress multiple - stress biotique - stress abiotique

Ecole(s) doctorale(s) de rattachement : ED 145 Sciences du végétal, Université Paris-Sud 11, Orsay

Contacts :

Institut Jean-Pierre Bourgin, UMR1318 INRA-AgroParisTech
Bâtiment
INRA Centre de Versailles-Grignon
Route de St-Cyr (RD10)
78026 Versailles Cedex France

tél : +33 (0)1 30 83 30 00 - fax : +33 (0)1 30 83 33 19



 

Mathilde Fagard                   
Responsable
Directeur de recherche

 

Alia Dellagi
Maître de conférences AgroParisTech

Marie-Christine Soulié
Maître de conférences Paris VI

Martine Rigault
Assistant ingénieur AgroParisTech

 

Mahsa Farjad
Doctorant
Du 1/10/13 au 30/9/16

 

 

Camille Verly
Doctorant
Du 1/11/15 au 31/10/18

 

Dominique Expert
Directeur de recherche émérite-CNRS

 

 


Anciens membres
de l'équipe

 




 

Résumé :
 
Nitrate is the main form of nitrogen absorbed by plants in our temperate countries. Numerous agronomic data and some recent functional data show that relationships between infection process and nitrogen metabolism are complex and common, involving poorly described mechanisms. Our project is to analyze the relationship between nitrogen metabolism and necrotrophic infection processes and to identify the regulation pathways involved. Our expertise on pathogens and their pathogenicity concerns three necrotrophic microorganisms: i.e., the bacteria Erwinia amylovora and Dickeya dadantii and the fungus Botrytis cinerea including Arabidopsis thaliana responses to those pathogens. Our first objective is to analyze nitrogen nutrition impact on the infection process: pathogenic factors and defense response expression. Relationships between nitrogen metabolism and pathogenicity will be studied taking advantages of the mutants studied within the APE department « Adaptation of Plants to the Environment ». Our ultimate objective is, through the study of natural variation and/or the establishment of genetic screenings, to identify plant factors allowing combined biotic and abiotic stress signals.

Figure 1: Arabidopsis leaf infected (top half) or not (bottom half) with E. amylovora and stained with DCFH-DA which reveals intracellular ROS accumulation in response to bacterial infection.

   

Résultats marquants :


The NPI team was created in December 2012. The founding members of the NPI team were formerly part of three separate teams of the “Plant-Pathogens Interactions laboratory” (LIPP) in Paris. The objectives of the LIPP were to study different molecular aspects of plant-pathogen interactions involving necrotrophic pathogens:


Role of the type III effector DspA/E in the interaction between Arabidopsis and Erwinia amylovora (Ea)

jpg  
Ea is a necrotrophic bacterium responsible for fire blight in plants of the rosaceace family such as apple and pear trees. The pathogenicity of Ea is depends on its type three secretion system (T3SS) which allow the bacteria to inject effector proteins inside the plant cells to alter plant defense production and or plant physiology to favor bacterial growth in the apoplast. We studied the role of the Ea type III effector DspA/E during the interaction of Ea with Arabidopsis. We found that although Arabidopsis displays non-host resistance towards Ea, the bacteria are able to transiently multiply and that DspA/E is required for this growth as in host plants. To determine the role of the toxicity of DspA/E in this process we constructed Arabidopsis plants expressing DspA/E, which we then mutagenized using the chemical agent EMS. The screening for suppressor mutants led to the identification of four mutants corresponding to two complementation groups that are currently under study. We also analyzed the role of reactive oxygen species such as hydrogen peroxide (H2O2) in the toxicity of DspA/E and during the interaction with Arabidopsis.
More recently, we demonstrated that a member of the nitrate transporter family, NRT2.6, which is strongly induced following Arabidopsis inoculation with Ea, contributes to non-host resistance of Arabidopsis to Ea.

Figure 2 : Wild-type Arabidopsis (Col-0), the transgenic line bearing the dspA/E gene under the control of an estradiol-inducible promoter (13-1-2) and the suppressor mutant 18 were grown on medium with (+) or without (-) estradiol.

   


Iron metabolism during the infection process of Arabidopsis by Dickeya dadantii (Dd).

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Iron is essential for most forms of life. However, it is poorly available under aerobic conditions and it can catalyze the formation of toxic free radicals via the Fenton reaction. Dd is an enterobacterium causing important losses on crops such as potato, chicory and ornamentals such as Saintpaulia. We have studied the role of iron homeostasis in the interaction between Arabidopsis and the necrotrophic bacterium Dickeya dadantii (Dd). We showed that iron nutrition modulates defense responses and resistance to several pathogens (Dd and the fungus Botrytis cinerea) in Arabidopsis. We showed that several genes involved in iron homeostasis in Arabidopsis (iron transport and storage) are involved in plant resistance to the pathogen Dd and that siderophores, ubiquitous microbial molecules with high iron scavenging capacity, are activators of plant defense. They can be used in plant protection or as tools to study the connections between iron homeostasis and defense. Their elicitor property relies on their strong iron scavenging capacity.
More recently, we have identified a novel MPK gene potentially involved in the perception of the signal of iron deficiency in the leaves and immunity of A. thaliana. The role of this gene in the Dd-Arabidopsis interaction is currently under investigation.

Figure 3  : Methylated pectin staining of Arabidopsis leaves infected with Dickeya dadantii.

   


Cell wall of Botrytis cinerea (Bc) and its role during the infection

jpg
 
Bc is a filamentous fungus responsible for the grey mould disease. Considered as a model organism for necrotrophic plant pathogens, two Botrytis strains B0510 and T4 have been annotated and are publicly available. In order to limit fungus development, it is essential to understand the mechanisms involved in fungal growth and colonization. We have characterized chitin synthases (CHS) of the fungus Bc), which are potential specific targets for antifungal drugs. Our work showed that  has eight CHS genes. Several mutants constructed by reverse genetics have been characterized: the Bcchs1 mutant displays cell wall weakening and reduced virulence, the Bcchs3a mutant has a drastic reduction in virulence. The BcCHS6 seems to be essential for Bc and therefore could represent a valuable antifungal target. Phenotypic features of the Bcchs3a mutant being particularly interesting, we have followed its characterization in more detail and found that mutant Bcchs3a was virulent on pad2 and pad3 mutant leaves defective in the phytoalexin camalexin (an antimicrobial compound). The Bcchs3 mutant induced camalexin accumulation at the infection site on Col-0 plants, correlated with overexpression of the PAD3 gene. Altogether, these results led us to assign a critical role to the BcCHS3a chitin synthase isoform, both in fungal virulence and plant defense response.
Figure 4: Left: Maceration symptoms provoked by wild-type Botrytis cinerea Arabidopsis leaves. Right: the Bcchs3 mutant, affected in a gene encoding a chitin
   


Links between nitrogen supply and metabolism and plant-pathogen interactions

Currently, the NPI team is investigating the links between nitrogen supply and metabolism and plant-pathogen interactions. Nitrate is the main form of nitrogen absorbed by plants in our temperate countries. Numerous agronomic data and some recent functional data show that relationships between infection process and nitrogen metabolism are complex and common, involving poorly described mechanisms. Our project is to analyze the relationship between nitrogen metabolism and necrotrophic infection processes and to identify the regulation pathways involved. Our expertise on pathogens and their pathogenicity concerns three necrotrophic microorganisms: i.e., the bacteria Erwinia amylovora and Dickeya dadantii and the fungus Botrytis cinerea including Arabidopsis thaliana responses to those pathogens. Our first objective is to analyze nitrogen nutrition impact on the infection process: pathogenic factors and defense response expression. Relationships between nitrogen metabolism and pathogenicity will be studied taking advantages of the mutants studied within the APE department « Adaptation of Plants to the Environment ». Our ultimate objective is, through the study of natural variation and/or the establishment of genetic screenings, to identify plant factors allowing combined biotic and abiotic stress signals.

 


Publications représentatives :

Fagard M, Launay A, Clément G, Courtial J, Dellagi A, Farjad M, Krapp A, Soulié MC and Masclaux-Daubresse C (2014). Nitrogen metabolism meets phytopathology. JExp Bot (doi: 10.1093/jxb/eru323) (abstract)

Degrave A, Moreau M, Launay A, Barny MA, Brisset MN, Patrit O, Taconnat L, Vedel R and Fagard M (2013). The bacterial effector DspA/E is toxic in A. thaliana and is required for multiplication and survival of fire blight pathogen. Mol Plant Pathol 14: 506-517.

Magellan H, Boccara M, Drujon T, Soulié M-C, Guillou C, Dubois J and Becker H (2013). Discovery of two new inhibitors of Botrytis cinerea chitin synthase by a chemical library screening. Bioorg. Med. Chem. 21, 4997-5003.

Morcx S, Kunz C, Choquer M, Assie S, Blondet E, Simond-Cote E, Gajek K, Chapeland-Leclerc F, Expert D, Soulié M-C (2013). Disruption of Bcchs4, Bcchs6 or Bcchs7 chitin synthase genes in Botrytis cinerea and the essential role of class VI chitin synthase (Bcchs6). Fungal genetics and biology. FG & B 52: 1-8.

Dechorgnat J, Patrit O, Krapp A, Fagard M and Daniel-Vedele F (2012). Characterization of the Nrt2.6 gene in Arabidopsis thaliana: a link with plant response to biotic and abiotic stress. PLoS One 7: e42491. (pdf)

Kieu NP, Aznar A, Segond D, Rigault M, Simond-Côte E, Kunz C, Soulie MC, Expert D and Dellagi A (2012). Iron deficiency affects plant defence responses and confers resistance to Dickeya dadantii and Botrytis cinerea. Mol Plant Pathol 13: 816-827.

Moreau M, Degrave A, Vedel R, Bitton F, Patrit O, Renou J-P, Barny M-A, Fagard M (2012) EDS1 Contributes to Nonhost Resistance of Arabidopsis thaliana Against Erwinia amylovora. Molecular Plant-Microbe Interactions 25: 421-430

Expert D, Franza T and Dellagi A (2012). Iron in plant-pathogen interactions.  Molecular Aspects of Iron Metabolism in Pathogenic and Symbiotic Plant Microbe Associations. SpringerBriefs in Biometals. (ed. Expert, D. O'Brian, M.).

Boureau T, Siamer S, Perino C, Gaubert S, Patrit O, Degrave A, Fagard M, Chevreau E and Barny M-A (2011). The HrpN Effector of Erwinia amylovora, Which Is Involved in Type III Translocation, Contributes Directly or Indirectly to Callose Elicitation on Apple Leaves. Molecular Plant-Microbe Interactions 24: 577-584.

Liu W, Soulié M-C, Perrino C, Fillinger S (2011). The osmosensing signal transduction pathway from Botrytis cinerea regulates cell wall integrity and MAP kinase pathways control melanin biosynthesis with influence of light. Fungal Genetics and Biology 48: 377-387.

Siamer S, Patrit O, Fagard M, Belgareh-Touzé N, Barny M-A (2011). Expressing the Erwinia amylovora type III effector DspA/E in the yeast Saccharomyces cerevisiae strongly alters cellular trafficking. FEBS Open Bio 1: 23-26

Arbelet D, Malfatti P, Simond-Cote E, Fontaine T, Desquilbet L, Expert D, Kunz C and Soulié M (2010). Disruption of the Bcchs3a Chitin Synthase Gene in Botrytis cinerea Is Responsible for Altered Adhesion and Overstimulation of Host Plant Immunity. Molecular Plant-Microbe Interactions 23: 1324-1334.

Dellagi A, Segond D, Rigault M, Fagard M, Simon C, Saindrenan P and Expert D (2009). Microbial Siderophores Exert a Subtle Role in Arabidopsis during Infection by Manipulating the Immune Response and the Iron Status. Plant Physiology 150: 1687-1696.

Segond D, Dellagi A, Lanquar V, Rigault M, Patrit O, Thomine S, Expert D (2009). NRAMP genes function in Arabidopsis thaliana resistance to Erwinia chrysanthemi infection. Plant J 58: 195-207.

Barny MA, Boureau T, Degrave A, Fagard M, Bouteau F, Gaubert S, Reboutier D and Brisset MN (2008). Type III effectors of E. amylovora: Synergistic and antagonistic effects. In KB Johnson, VO Stockwell, eds, Proceedings of the Eleventh International Workshop on Fire Blight, Vol 793, pp 215-220.

Degrave A, Fagard M, Perino C, Brisset MN, Gaubert S, Laroche S, Patrit O and Barny MA (2008). Erwinia amylovora type three-secreted proteins trigger cell death and defense responses in Arabidopsis thaliana. Molecular Plant-Microbe Interactions 21: 1076-1086

Fagard M, Dellagi A, Roux C, Périno C, Rigault M, Boucher V, Shevchik V, Expert D (2007) Arabidopsis thaliana expresses multiple lines of defense to counter-attack Erwinia chrysanthemi. Molecular Plant Microbe Interactions 20: 794-805.

Boughammoura A, Franza T, Dellagi A, Roux C, Matzanke-Markstein B, and Expert D (2007) Ferritins, bacterial virulence and plant defence. BioMetals 20: 347-353.

2006 and before (selected publications):

Choquer M, Boccara M, Goncalves IR, Soulié MC, Vidal-Cros A (2004) Survey of the Botrytis cinerea chitin synthase multigenic family through the analysis of six euascomycetes genomes. European Journal of Biochemistry 271: 2153-2164.

Dellagi A, Rigault M, Segond D, Roux C, Kraepiel Y, Cellier F, Briat J, Gaymard F, Expert D (2005) Siderophore-mediated upregulation of Arabidopsis ferritin expression in response to Erwinia chrysanthemi infection. Plant J 43: 262-272.
 
Soulié MC, Perino C, Piffeteau A, Choquer M, Malfatti P, Cimerman A, Kunz C, Boccara M, Vidal-Cros A (2006) Botrytis cinerea virulence is drastically reduced after disruption of chitin synthase class III gene (Bcchs3a). Cellular Microbiology 8: 1310–1321.

Soulié MC, Piffeteau A, Choquer M, Boccara M, Vidal-Cros A (2003) Disruption of Botrytis cinerea class I chitin synthase gene Bcchs1 results in cell wall weakening and reduced virulence. Fungal Genetics and Biology 8: 1310-1321.

 
 
 

 


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