Analysis of the role of nitric oxide (NO) in the cross‐regulation between immunity, growth and iron homeostasis in plants

Studies performed in our Agroecology Department show that the immune response of plants is linked to their iron nutrition and is modulated by pyoverdine, a siderophore produced by the plant beneficial rhizobacteria Pseudomonas fluorescens C7R12. Accordingly, Arabidopsis thaliana plantlets exposed to iron deficiency and treated with pyoverdine in its iron non‐chelated structure (apo‐pyo) show an enhanced growth but a decreased immune response capacity. We hypothesize that nitric oxide (NO), a universal signaling molecule, is a key component of the regulation of the immune response in plants exposed to apo‐pyo and to the C7R12 strain. We checked by fluorescence microscopy that NO is actually …

Study of the stress-related signalling of endoplasmic reticulum in grapevine immunity associated to LysM receptor kinases (VvLYKs)

In the actual situation of decreasing the use of chemicals in agriculture, enhancing our understanding of plant immunity is a critical task to develop more sustainable plant health protection methods. An interesting strategy is to study how plants, and in particular grapevine, perceives and responds to different microorganisms. Microorganisms are notably recognizedby LysM Receptor-like Kinase (LYKs) and previous works have identified 16 LYKs encoded by the grapevine genome (VvLYKs) (Roudaire et al. 2023). Among them, VvLYK1-1, VvLYK1-2 and VvLYK5-1 are involved in chitin perception and thus play a role in the plant immunity. Interestingly, immune responses are also involved during symbiotic…

Antimicrobial Activity of Resveratrol Analogues

Stilbenes, especially resveratrol and its derivatives, have become famous for their positive effects on a wide range of medical disorders, as indicated by a huge number of published studies. A less investigated area of research is their antimicrobial properties. A series of 13 trans-resveratrol analogues was synthesized via Wittig or Heck reactions, and their antimicrobial activity assessed on two different grapevine pathogens responsible for severe diseases in the vineyard. The entire series, together with resveratrol, was first evaluated on the zoospore mobility and sporulation level of Plasmopara viticola (the oomycete responsible for downy mildew). Stilbenes displayed a spectrum of acti…

The Pseudomonas fluorescens Siderophore Pyoverdine Weakens Arabidopsis thaliana Defense in Favor of Growth in Iron-Deficient Conditions

SPE EA BIOME IPM UB INRA; International audience; Pyoverdines are siderophores synthesized by fluorescent Pseudomonas spp. Under iron-limiting conditions, these high-affinity ferric iron chelators are excreted by bacteria in the soil to acquire iron. Pyoverdines produced by beneficial Pseudomonas spp. ameliorate plant growth. Here, we investigate the physiological incidence and mode of action of pyoverdine from Pseudomonas fluorescens C7R12 on Arabidopsis (Arabidopsis thaliana) plants grown under iron-sufficient or iron-deficient conditions. Pyoverdine was provided to the medium in its iron-free structure (apo-pyoverdine), thus mimicking a situation in which it is produced by bacteria. Rema…

NO signaling in cryptogein-induced immune responses in tobacco

Nitric oxide production in tobacco leaf cells: a generalized stress response?

The function of nitric oxide (NO), a gaseous free radical emitted by many plants, is incompletely understood. In the present study the hypothesis that NO generation, like that of the reactive oxygen species, occurs as a general response to different environmental cues was tested. Leaf peels and mesophyll cell suspensions of Nicotiana tabacum cv. Xanthi were loaded with the NO-specific fluorophore, diaminofluorescein, and subjected to an abiotic stressor. Light stress and mechanical injury had no apparent effect on NO production. In contrast, high temperatures, hyperosmotic stress, salinity and epi-illumination in a microscope all led to rapid surges in NO-induced fluorescence. The fluoresce…

Evolutionary diversification of type-2 HDAC structure, function and regulation in Nicotiana tabacum

Ministère de l'Education Nationale et de la Recherche ; Conseil Régional de Bourgogne (PARI AGRALE8) ; Association pour la Recherche sur les Nicotianacées ; Conseil Régional de Bourgogne; International audience; Type-2 HDACs (HD2s) are plant-specific histone deacetylases that play diverse roles during development and in responses to biotic and abiotic stresses. In this study we characterized the six tobacco genes encoding HD2s that mainly differ by the presence or the absence of a typical zinc finger in their C-terminal part. Of particular interest, these HD2 genes exhibit a highly conserved intron/exon structure. We then further investigated the phylogenetic relationships among the HD2 gen…

La pyoverdine, un sidérophore produit par la souche bénéfique du sol Pseudomonas fluorescens C7R12, impacte l’homéostasie du fer et promeut la croissance aux dépens des réactions de défense d’Arabidopsis thaliana cultivée en condition de carence en fer

Identification and functional characterization of S-nitrosated proteins from Klebsormidium nitens

The small gaseous molecule nitric oxide (NO) is well established as a major ubiquitous component of cell signalling. A key signalling mechanism mediating NO effects is S-nitrosation, a post-translational modification by which NO can impact the target protein activities, subcellular localizations and capacities to form protein complexes. The identification of proteins targeted by NO is of major interest in order to elucidate NO functions. Interestingly, land plants lack NO synthase (NOS), which is the main enzyme for NO synthesis in metazoans, while a few algal species possess it, thus raising many interrogations. Therefore, we focused on the identification of S-nitrosated proteins during sa…

Identification and functional characterization of S-nitrosated proteins from Klebsormidium nitens, a model alga to study plant adaptation to land

The small gaseous molecule nitric oxide (NO) is well established as a major ubiquitous component of cell signalling. A key signalling mechanism mediating NO effects is S-nitrosation, a post-translational modification by which NO can impact the target protein activities, subcellular localizations, and capacities to form protein complexes. The identification of proteins targeted by NO is of major interest in order to elucidate NO functions. Interestingly, land plants lack NO synthase (NOS), which is the main enzyme for NO synthesis in metazoans, while a few algal species possess it, thus raising many interrogations. Therefore, we focused on the identification of S-nitrosated proteins during s…

Induced resistance as a strategy for vineyard protection

National audience; As most grown grapevine Vitis vinifera varieties are susceptible to diseases such as downy and powdery mildews, numerous treatments are required to ensure a satisfactory yield and harvest quality. However, the use of phytochemical fungicides has serious drawbacks: some of them are potentially harmful for the environment and human health and contribute to the selection of resistant pathogen strains. Nowadays, in an objective of sustainable viticulture, there is increasing societal request, political incitation and winegrower’s awareness to reduce the use of pesticides. For these reasons, alternative strategies of protection are under research. In our laboratory, we are stu…

S-Nitrosation of Arabidopsis thaliana Protein Tyrosine Phosphatase 1 Prevents Its Irreversible Oxidation by Hydrogen Peroxide

Tyrosine-specific protein tyrosine phosphatases (Tyr-specific PTPases) are key signaling enzymes catalyzing the removal of the phosphate group from phosphorylated tyrosine residues on target proteins. This post-translational modification notably allows the regulation of mitogen-activated protein kinase (MAPK) cascades during defense reactions. Arabidopsis thaliana protein tyrosine phosphatase 1 (AtPTP1), the only Tyr-specific PTPase present in this plant, acts as a repressor of H2O2 production and regulates the activity of MPK3/MPK6 MAPKs by direct dephosphorylation. Here, we report that recombinant histidine (His)-AtPTP1 protein activity is directly inhibited by H2O2 and nitric oxide (NO) …

Analysis of the cross‐regulation between immunity, growth and iron homeostasis in plants

The existence of a tightly regulated balance between growth and immunity in plants has recently emerged. In this study, we challenged this concept thanks to the biological model pyoverdine-Arabidopsis thaliana. Pyoverdine is a siderophore produced by the plant growth promoting rhizobacteria Pseudomonas fluorescens C7R12. Under iron deficiency, P. fluorescens excretes the iron free form of pyoverdine (apo‐pyo) in the soil. Once chelated with iron (ferri‐pyo), the complex is internalized by the bacteria. We demonstrated that Arabidopsis thaliana plants treated by apo‐pyo in a medium containing or not iron internalize pyoverdine. Interestingly, apo‐pyo-treated plants did not show a typical gro…

Role of nitric oxide synthase (NOS) of Klebsormidium nitens: Identification and characterization of partners

National audience; Nitric oxide (NO) is an important cellular signaling molecule across kingdoms. During bioticor abiotic stresses, NO burst is detected in both plants and mammals although no sequenceencoding the well described mammalian NO synthase (NOS) is highlighted in plants. Comparedto terrestrial plants, some algae present transcripts encoding the NOS-like enzyme. Amongthem, Klebsormidium nitens the model alga to study the early transition step from aquatic algaeto land plants is found. As mechanisms governing NO synthesis and signaling in green lineageremain unclear, the study of NOS from K. nitens (KnNOS) through (i) the identification ofregulator proteins, (ii) the identification …

Role of nitric oxide synthases from Klebsormidium nitens: structural characterization and identification of protein partners

Nitric oxide (NO) is an important cellular signalling molecule regulating various physiological processes, in both animals andplants. In animals, NO synthesis is mainly catalysed by NO synthase (NOS) enzymes. During biotic or abiotic stresses, NOSlike activities that are sensitive to mammalian NOS inhibitors have been detected in plants, although no sequences encodingthe well described mammalian NOS are highlighted in land plants. Interestingly, we identified NOS-like sequences in 20 algaespecies. Among them, NOSs are found in Klebsormidium nitens the model alga to study the early transition step from aquaticalgae to land plants.As mechanisms governing NO synthesis and signalling in green l…

Les stimulateurs de défense des plantes (SDP) en viticulture - Un tour d’horizon sur les principes, les applications et les perspectives

IPMSPEINRAUBCNRS; Les stimulateurs de défense des plantes (SDP) en viticulture - Un tour d’horizon sur les principes, les applications et les perspectives

Dual mode of action of grape cane extracts against Botrytis cinerea

International audience; Crude extracts of Vitis vinifera canes represent a natural source of stilbene compounds with well characterized antifungals properties. In our trials, exogenous application of a stilbene extract (SE) obtained from grape canes on grapevine leaves reduces the necrotic lesions caused by Botrytis cinerea. The SE showed to possess a direct antifungal activity by inhibiting the mycelium growth. The activation of some grapevine defense mechanism was also investigated. H2O2 production and activation of mitogen-activated protein kinase (MAPK) phosphorylation cascades as well as accumulation of stilbenoid phytoalexins were explored on grapevine cell suspension. Moreover, the t…

Role of nitric oxide synthases from klebsormidium nitens: first structural characterization and partners identification

Objectives: Nitric oxide (NO) is an important cellular signaling molecule regulating various physiologicalprocesses, in both animals and plants. In animals, NO synthesis is mainly catalyzed by NO synthase(NOS) enzymes. In plants, NOS-like activities sensitive to mammalian NOS inhibitors have beenmeasured, although no sequences encoding mammalian NOSs have been found in land plants.Interestingly, we identified NOS-like sequences in 20 algae species. These latter include thefilamentous charophyte green algae Klebsormidium nitens, a biological model to study the earlytransition step from aquatic algae to land plants. In order to understand the mechanisms governingNO synthesis and signaling in …

Identification and functional characterization of S-nitrosated proteins from Klebsormidium nitens in response to salt stress

The small gaseous molecule nitric oxide (NO) is well established as a major ubiquitouscomponent of cell signalling. A key signalling mechanism mediating NO effects is Snitrosation, a post-translational modification by which NO can impact the target proteinactivities, subcellular localizations, and capacities to form protein complexes. The identificationof proteins targeted by NO is of major interest in order to elucidate NO functions. Interestingly,land plants lack NO synthase (NOS), which is the main enzyme for NO synthesis in metazoans,while a few algal species possess it, thus raising many interrogations. Therefore, we focused onthe identification of S-nitrosated proteins during salt str…

Development of a chitosan-based biocontrol product against crop diseases

Nitric oxide production and signalling in algae

International audience

Nitric oxide and glutathione impact the expression of iron uptake- and iron transport-related genes as well as the content of metals in A. thaliana plants grown under iron deficiency

International audience; Mounting evidence indicate that nitric oxide (NO) acts as a signaling molecule mediating iron deficiency responses through the upregulation of the expression of iron uptake-related genes. Accordingly, NO donors such as nitrosoglutathione (GSNO) were reported to improve the fitness of plants grown under iron deficiency. Here, we showed that glutathione, a by-product of GSNO, triggered the upregulation of the expression of iron uptake- and transport-related gene and an increase of iron concentration in Arabidopsis thaliana seedlings facing iron deficiency. Furthermore, we provided evidence that under iron deficiency, NO released by GSNO did not improve the root iron co…

The chaperone-like protein Cdc48 regulates ubiquitin-proteasome system in plants.

The degradation of misfolded proteins is mainly mediated by the ubiquitin-proteasome system (UPS). UPS can be assisted by the protein Cdc48 but the relationship between UPS and Cdc48 in plants has been poorly investigated. Here, we analyzed the regulation of UPS by Cdc48 in tobacco thanks to two independent cell lines overexpressing Cdc48 constitutively and plant leaves overexpressing Cdc48 transiently. In the cell lines, the accumulation of ubiquitinated proteins was affected both quantitatively and qualitatively and the amount of proteasomal subunits was modified, while proteolytic activities were unchanged. Similarly, the over-expression of Cdc48 in planta impacted the accumulation of ub…

β-Aminobutyric Acid (BABA)-Induced Resistance in Arabidopsis thaliana: Link with Iron Homeostasis

International audience; Bêta-Aminobutyric acid (BABA) is a nonprotein amino acid inducing resistance in many different plant species against a wide range of abiotic and biotic stresses. Nevertheless, how BABA primes plant natural defense reactions remains poorly understood. Based on its structure, we hypothesized and confirmed that BABA is able to chelate iron (Fe) in vitro. In vivo, we showed that it led to a transient Fe deficiency response in Arabidopsis thaliana plants exemplified by a reduction of ferritin accumulation and disturbances in the expression of genes related to Fe homeostasis. This response was not correlated to changes in Fe concentrations, suggesting that BABA affects the…