Bacterial rhamnolipids and their 3-hydroxyalkanoate precursors activate Arabidopsis innate immunity through two independent mechanisms.

Plant innate immunity is activated upon perception of invasion pattern molecules by plant cell-surface immune receptors. Several bacteria of the genera Pseudomonas and Burkholderia produce rhamnolipids (RLs) from l-rhamnose and (R)-3-hydroxyalkanoate precursors (HAAs). RL and HAA secretion is required to modulate bacterial surface motility, biofilm development, and thus successful colonization of hosts. Here, we show that the lipidic secretome from the opportunistic pathogen Pseudomonas aeruginosa, mainly comprising RLs and HAAs, stimulates Arabidopsis immunity. We demonstrate that HAAs are sensed by the bulb-type lectin receptor kinase LIPOOLIGOSACCHARIDE-SPECIFIC REDUCED ELICITATION/S-DOMAIN-1-29 (LORE/SD1-29), which also mediates medium-chain 3-hydroxy fatty acid (mc-3-OH-FA) perception, in the plant Arabidopsis thaliana HAA sensing induces canonical immune signaling and local resistance to plant pathogenic Pseudomonas infection. By contrast, RLs trigger an atypical immune response and resistance to Pseudomonas infection independent of LORE. Thus, the glycosyl moieties of RLs, although abolishing sensing by LORE, do not impair their ability to trigger plant defense. Moreover, our results show that the immune response triggered by RLs is affected by the sphingolipid composition of the plasma membrane. In conclusion, RLs and their precursors released by bacteria can both be perceived by plants but through distinct mechanisms.

Comparative analysis of defence responses induced by the endophytic plant growth-promoting rhizobacterium Burkholderia phytofirmans strain PsJN and the non-host bacterium Pseudomonas syringae pv. pisi in grapevine cell suspensions.

Plant growth-promoting rhizobacteria (PGPR) are beneficial microorganisms that colonize the rhizosphere of many plant species and confer beneficial effects, such as an increase in plant growth. PGPR are also well known as inducers of systemic resistance to pathogens in plants. However, the molecular mechanisms involved locally after direct perception of these bacteria by plant cells still remain largely unknown. Burkholderia phytofirmans strain PsJN is an endophytic PGPR that colonizes grapevine and protects the plant against the grey mould disease caused by Botrytis cinerea. This report focuses on local defence events induced by B. phytofirmans PsJN after perception by the grapevine cells. It is demonstrated that, after addition to cell suspension cultures, the bacteria were tightly attaching to plant cells in a way similar to the grapevine non-host bacteria Pseudomonas syringae pv. pisi. B. phytofirmans PsJN perception led to a transient and monophasic extracellular alkalinization but no accumulation of reactive oxygen species or cell death were detected. By contrast, challenge with P. syringae pv. pisi induced a sustained and biphasic extracellular alkalinization, a two phases oxidative burst, and a HR-like response. Perception of the PGPR also led to the production of salicylic acid (SA) and the expression of a battery of defence genes that was, however, weaker in intensity compared with defence gene expression triggered by the non-host bacteria. Some defence genes up-regulated after B. phytofirmans PsJN challenge are specifically induced by exogenous treatment with SA or jasmonic acid, suggesting that both signalling pathways are activated by the PGPR in grapevine.

Characterization of a F-box gene up-regulated by phytohormones and upon biotic and abiotic stresses in grapevine.

F-box proteins are key components of the ubiquitin (Ub)/26S proteasome pathway that mediates selective degradation of regulatory proteins involved in a wide variety of cellular processes affecting eukaryotic cells. In plants, F-box genes form one of the largest multigene superfamilies and control many important biological functions. Among the F-box genes characterized to date only few have been involved in the regulation of plant defense responses. Moreover, no F-box genes have been studied and characterized in grapevine. Using a differential display approach we isolated a F-box gene (BIG-24.1), which is up-regulated during Botrytis cinerea infection of grapevine leaves. BIG-24.1 encodes a polypeptide of 386 amino acids with a conserved F-box domain in the N-terminus region and a kelch domain. By investigating expression profiles of BIG-24.1, we show that the gene expression is strongly stimulated in B. cinerea infected berries and in grapevine cells challenged by MAMP rhamnolipids, a non-host bacterium and an endophytic rhizobacterium. The gene is also strongly induced by abiotic stresses including UV-C and wounding or by salicylic acid, methyl-jasmonate, ethylene and abscisic acid that are known to be involved in defense signalling pathways. In addition, sequence analysis of the BIG-24.1 promoter revealed the presence of several regulatory elements involved in the activation of plant defense responses.

Biosurfactants in Plant Protection Against Diseases: Rhamnolipids and Lipopeptides Case Study.

Biosurfactants are amphiphilic surface-active molecules that are produced by a variety of microorganisms including fungi and bacteria. Pseudomonas, Burkholderia, and Bacillus species are known to secrete rhamnolipids and lipopeptides that are used in a wide range of industrial applications. Recently, these compounds have been studied in a context of plant-microbe interactions. This mini-review describes the direct antimicrobial activities of these compounds against plant pathogens. We also provide the current knowledge on how rhamnolipids and lipopeptides stimulate the plant immune system leading to plant resistance to phytopathogens. Given their low toxicity, high biodegradability and ecological acceptance, we discuss the possible role of these biosurfactants as alternative strategies to reduce or even replace pesticide use in agriculture.

Microspore embryogenesis and programmed cell death in barley: effects of copper on albinism in recalcitrant cultivars.

Albinism remains a major problem in cereal improvement programs that rely on doubled haploid (DH) technology, and the factors controlling the phenomenon are not well understood. Here we report on the positive influence of copper on the production of DH plants obtained through microspore embryogenesis (ME) in recalcitrant cultivars of barley (Hordeum vulgare L.). The presence of copper sulphate in the anther pre-treatment medium improved green DH plant regeneration from cultivars known to produce exclusively albino plants using classical procedures. In plastids, the effect of copper was characterized by a decrease in starch and a parallel increase in internal membranes. The addition of copper sulphate in the ME pre-treatment medium should enable breeders to exploit the genetic diversity of recalcitrant cultivars through DH technology. We examined programmed cell death (PCD) during microspore development to determine whether PCD may interfere with the induction of ME and/or the occurrence of albinism. By examining the fate of nuclei in various anther cell layers, we demonstrated that the kinetics of PCD in anthers differed between the barley cultivars Igri and Cork that show a low and a high rate of albinism, respectively. However, no direct correlation between PCD in the anther cell layers and the rate of albinism was observed and copper had no influence on the PCD kinetic in these cultivars. It was concluded that albinism following ME was not due to PCD in anthers, but rather to another unknown phenomenon that appears to specifically affect plastids during microspore/pollen development.

The herbicide flumioxazin stimulates pathogenesis-related gene expression and enzyme activities in Vitis vinifera.

In this work, the capacity of the soil-applied herbicide flumioxazin (fmx) to trigger defence mechanisms was assessed using 6-week-old in vitro grown Vitis vinifera L. plantlets. Time-course studies demonstrated that the herbicide induced the expression of basic beta-1,3-glucanase (Vvglu), basic chitinase (Vvchit1b) and PR10 (VvPR10.3) genes encoding three pathogenesis-related (PR) proteins involved in grapevine defence against pathogens. Thus, all transcripts accumulated in grapevine tissues to reach maximum values after 24-72 h of herbicide exposure, except for VvPR10.3 gene expression, which was induced in roots and stems but not in leaves. Induction of PR genes was observed to a greater extent in roots and leaves, and its intensity diminished in the stems although still remained noteworthy. The activities of beta-1,3-glucanase and chitinase enzymes significantly increased in the whole plant after herbicide exposure and were still stimulated 21 days after the beginning of treatments. Similarly, the most remarkable effect occurred in roots. However, all enzyme activities tested were stimulated in the upper aerial tissues as well, indicating that fmx or a derived product acts systemically, likely via root uptake.

Expression and in situ localization of two major PR proteins of grapevine berries during development and after UV-C exposition.

In grapevine Vitis vinifera L. cv Pinot noir, the Pathogenesis-Related (PR) proteins CHI4D and TL3 are among the most abundant extractable PR proteins of ripe berries and accumulate during berry ripening from véraison until full maturation. Evidence was supplied in favor of the involvement of these two protein families in plant defense mechanisms and plant development. In order to better understand CHI4D and TL3 function in grapevine, we analyzed their temporal and spatial pattern of expression during maturation and after an abiotic stress (UV-C) by in situ hybridization (ISH) and immunohistolocalization. In ripening berries, CHI4D and TL3 genes were mainly expressed in the exocarp and around vascular bundles of the mesocarp. In UV-C exposed berries, CHI4D and TL3 gene expression was strongly induced before véraison. Corresponding proteins localized in the exocarp and, to a lesser extent, around vascular bundles of the mesocarp. The spatial and temporal accumulation of the two PR proteins during berry maturation and after an abiotic stress is discussed in relation to their putative roles in plant defense.

Microspore embryogenesis in barley: anther pre-treatment stimulates plant defence gene expression.

Microspore embryogenesis (ME) is a process in which the gametophytic pollen programme of the microspore is reoriented towards a new embryo sporophytic programme. This process requires a stress treatment, usually performed in the anther or isolated microspores for several days. Despite the universal use of stress to induce ME, very few studies have addressed the physiological processes that occur in the anther during this step. To further understand the processes triggered by stress treatment, we followed the response of anthers by measuring the expression of stress-related genes in two barley (Hordeum vulgare L.) cultivars differing in their ME response. Genes encoding enzymes involved in oxidative stress (glutathione-S-transferase, GST; oxalate oxidase, OxO), in the synthesis of jasmonic acid (13-lipoxygenase, Lox; allene oxide cyclase, AOC; allene oxide synthase, AOS) and in the phenylpropanoid pathway (phenylalanine ammonia lyase, PAL), as well as those encoding PR proteins (Barwin, chitinase 2b, Chit 2b; glucanase, Gluc; basic pathogenesis-related protein 1, PR1; pathogenesis-related protein 10, PR10) were up-regulated in whole anthers upon stress treatment, indicating that anther perceives stress and reacts by triggering general plant defence mechanisms. In particular, both OxO and Chit 2b genes are good markers of anther reactivity owing to their high level of induction during the stress treatment. The effect of copper sulphate appeared to limit the expression of defence-related genes, which may be correlated with its positive effect on the yield of microspore embryos.

Programmed cell death progressively models the development of anther sporophytic tissues from the tapetum and is triggered in pollen grains during maturation.

To characterize the spatial and temporal occurrence of programmed cell death (PCD) in Lilium anther tissues, we used both microscopical and molecular markers of apoptosis for developmental stages from meiosis to pollen release. The first hallmarks of PCD include cell condensation and shrinkage of the cytoplasm, separation of chromatin into delineated masses, and DNA fragmentation in the tapetum as early as the premeiosis stage. PCD then extended to other anther sporophytic tissues, leading to anther dehiscence. Although the PCD clearly affected the endothecium and the epidermis, these two cell layers remained alive until anther dehiscence. In pollen, no sign of PCD was found until pollen mitosis I, after what apoptotic features developed progressively in the vegetative cell. In addition, DNA ladders were detected in all sporophytic tissues and cell types throughout pollen development, whereas in the male gametophyte DNA ladders were only detected during pollen maturation. Our data suggest that PCD is a progressive and active process affecting all the anther tissues, first being triggered in the tapetum.