Signal Molecules for Plant Defense Responses to Biotic Stress

Plants are capable of recognizing the penetrating pathogens and of responding to their attack by the activation of the defense systems. Signal transduction from the receptor to the cell genome is required for this activation. Recently, signal molecules have been found, which are involved in the signal transduction triggered in response to biotic stress. The data accumulated imply the presence of a complex and well-coordinated signal network in plant cells. This net controls plant defense responses to pathogen attacks.     

Terpene Down-Regulation Triggers Defense Responses in Transgenic Orange Leading to Resistance against Fungal Pathogens

Terpenoid volatiles are isoprene compounds that are emitted by plants to communicate with the environment. In addition to their function in repelling herbivores and attracting carnivorous predators in green tissues, the presumed primary function of terpenoid volatiles released from mature fruits is the attraction of seed-dispersing animals. Mature oranges (Citrus sinensis) primarily accumulate terpenes in peel oil glands, with d-limonene accounting for approximately 97% of the total volatile terpenes. In a previous report, we showed that down-regulation of a d-limonene synthase gene alters monoterpene levels in orange antisense (AS) fruits, leading to resistance against Penicillium digitatum infection. A global gene expression analysis of AS versus empty vector (EV) transgenic fruits revealed that the down-regulation of d-limonene up-regulated genes involved in the innate immune response. Basal levels of jasmonic acid were substantially higher in the EV compared with AS oranges. Upon fungal challenge, salicylic acid levels were triggered in EV samples, while jasmonic acid metabolism and signaling were drastically increased in AS orange peels. In nature, d-limonene levels increase in orange fruit once the seeds are fully viable. The inverse correlation between the increase in d-limonene content and the decrease in the defense response suggests that d-limonene promotes infection by microorganisms that are likely involved in facilitating access to the pulp for seed-dispersing frugivores.Plants are sessile organisms that produce and emit a vast array of volatile organic compounds (VOCs) to communicate between parts of the same plant and with other plants. It is generally accepted that the original role of these compounds in nature is related to defense functions (Degenhardt et al., 2003). Most VOCs are terpenoids, fatty acid degradation compounds, phenylpropanoids, and amino acid-derived products. Among these, terpenoids are likely to be the most abundant and expensive to produce (Gershenzon, 1994). Terpenoids are isoprenoid-derived compounds synthesized through the condensation of C5 isoprene units, a process that is catalyzed by a wide diversity of terpene synthases using geranyl diphosphate (GDP), farnesyl diphosphate (FDP), and geranylgeranyl diphosphate (GGDP) as substrates. These reactions give rise to the C5 hemiterpenes, the C10 monoterpenes, the C15 sesquiterpenes, and the C20 diterpenes (Dudareva et al., 2006).In green tissues, volatile terpenoid synthesis is either induced upon wounding or occurs constitutively; terpenes can be then stored in specific organs or tissues where they would be most effective in defense responses, such as leaf trichomes, resin ducts and lacticifers, pockets near the epidermis, or secretory cavities in Citrus spp. (Langenheim, 1994; Turner et al., 2000; Trapp and Croteau, 2001; Voo et al., 2012). Genetic engineering experiments have demonstrated that specific terpenoid compounds emitted by leaves can intoxicate, repel, or deter herbivores (Aharoni et al., 2003; Wu et al., 2006), or they may attract the natural predators and parasitoids of damaging herbivores to protect plants from further damage (Kappers et al., 2005; Schnee et al., 2006). These terpenoids are naturally found in complex mixtures, and it has been proposed that they can act synergistically, as in conifer resin, for simultaneous protection against pests and pathogens (Phillips and Croteau, 1999). Although fatty acid degradation products (such as jasmonates) and phenylpropanoids (such as salicylates) as well as their volatile and nonvolatile precursors are clearly involved in many induced defense responses against pests and pathogens (Glazebrook, 2005), much less is known regarding the participation of terpenoid volatiles in the defense against microorganisms in plants and about the possible interactions of these terpenoids with phytohormones.In contrast to their function in leaves, when released from flowers and mature fruits, the main function of terpenoid volatiles is in the attraction of pollinators (Pichersky and Gershenzon, 2002; Kessler et al., 2008; Junker and Blüthgen, 2010; Schiestl, 2010) and seed-dispersing animals (Lomáscolo et al., 2010; Rodríguez et al., 2011b), respectively. Fruit maturation and ripening are usually associated with large increases in the synthesis and accumulation of specific flavored volatiles, which are proposed to function as signals for seed dispersal (Auldridge et al., 2006; Goff and Klee, 2006; Rodríguez et al., 2013).Upon wounding, plant responses to biotic stresses are orchestrated locally and systemically by signaling molecules. Among these molecules, the jasmonates regulate defenses against arthropod herbivores and necrotroph fungal pathogens as well as biotrophic pathogens, such as some mildews (Ellis and Turner, 2001; Stintzi et al., 2001; Kessler et al., 2004; Li et al., 2005; Wasternack, 2007; Browse and Howe, 2008). In addition to jasmonates, molecules such as salicylic acid (SA) and ethylene appear to regulate distinct defense pathways and are major synergistic (Mur et al., 2006) or antagonistic (De Vos et al., 2005) regulators of plant innate immunity. Plants produce a specific blend of these alarm signals after pathogen or pest attacks, and the production of these molecules varies greatly in quantity, composition, and timing. These signals activate differential sets of defense-related genes that eventually determine the nature of the defense response against the attacker (Reymond and Farmer, 1998; Rojo et al., 2003; De Vos et al., 2005). All genes that encode enzymes involved in the biosynthesis of jasmonates are jasmonic acid (JA) inducible (Wasternack, 2006), indicating that JA biosynthesis is regulated by positive feedback. The precursor for the biosynthesis of JA is α-linolenic acid. The activity of the 13-lipoxygenase (LOX), allene oxide synthase (AOS), and allene oxide cyclase (AOC) enzymes converts α-linolenic acid to cis-(+)-12-oxophytodienoic acid (OPDA). OPDA REDUCTASE3 catalyzes the reduction of OPDA (and dinor-OPDA) to oxo-pentenyl-cycloheptane-octanoic acid, which, in turn, undergoes three rounds of β-oxidation leading to jasmonyl-CoA formation. Jasmonyl-CoA is then cleaved by a putative thioesterase yielding (+)-7-iso-JA, which equilibrates to the more stable (−)-JA (Wasternack and Kombrink, 2010).The exogenous application of jasmonates on plants and the existence of mutant and/or transgenic plants altered in JA biosynthesis or signaling have led to altered susceptibility or resistance to pathogens. Impaired JA biosynthesis or signaling is generally associated with decreased levels of defensive compounds, including VOCs, and reduced plant biomass and/or fitness under insect attack (Howe et al., 1996; Halitschke and Baldwin, 2004). For example, Arabidopsis (Arabidopsis thaliana) mutants defective in JA perception (e.g. coronatine-insensitive1 [coi1]) or biosynthesis (e.g. aos and defective in anther dehiscence1) are susceptible to pathogen infections (Feys et al., 1994; Xie et al., 1998; Park et al., 2002; Turner et al., 2002). In contrast, mutants (e.g. constitutive expression of vegetative storage protein1 and Arabidopsis Ser/Thr phosphatase of type 2C1) with constitutive or wound-induced activation of the JA pathway exhibit enhanced resistance to fungal pathogens and pests and phenotypes characteristic of JA-treated plants (Ellis and Turner, 2001; Ellis et al., 2002; Schweighofer et al., 2007).Sweet orange (Citrus sinensis) is a perennial tree species that is exposed to recurrent biotic and abiotic challenges during its decades of growth in orchards. Orange fruits undergo a nonclimacteric maturation process in which the biochemistry, physiology, and structure of the organ are altered to complete the release of mature seeds. These changes typically include fruit growth and texture modification; color change through the degradation of chlorophylls and a parallel induction of carotenogenesis in the peel (flavedo) and pulp; flavonoid accumulation in the pulp; increases and decreases in the sugar and acid contents, respectively; and global accumulation and selective emission of volatile terpenoids (Spiegel-Roy and Goldschmidt, 1996). In nature, d-limonene accumulates gradually in the oil glands of the peel during fruit development and reaches its maximum level shortly before the breaker stage, followed by a steady decline during maturation (Attaway et al., 1967; Kekelidze et al., 1989; Rodríguez et al., 2011b). The high amount of d-limonene that accumulates in orange peels has a tremendous metabolic cost, suggesting an important biological role for this terpene and other related compounds in the interactions between fruits and the biotic environment.Previously, we examined the biological role of d-limonene by manipulating oil gland chemistry via the antisense (AS) overexpression of a d-limonene synthase gene from Satsuma mandarin (Citrus unshiu) in orange fruits. Compared with empty vector (EV) controls, fruit peels from AS transformants showed a dramatic reduction in d-limonene accumulation; decreased levels of other monoterpenes, sesquiterpenes, and monoterpene aldehydes; and increased levels of monoterpene alcohols. When challenged with the necrotroph fungus Penicillium digitatum, the causal agent of green mold rot, AS-transformed fruits were highly resistant to fungal infection. Full susceptibility to P. digitatum infection was restored when AS fruits were supplemented with d-limonene but not other monoterpene alcohols, indicating that d-limonene accumulation in the orange peel was required for the successful progress of this plant-pathogen interaction (Rodríguez et al., 2011a, 2011b). Green mold rot is the most important postharvest disease of citrus fruit worldwide, accounting for up to 60% to 80% of total losses during postharvest life of the fruit. P. digitatum is considered to be a specialist pathogen of citrus fruits that efficiently infects the peel through injuries in which ubiquitous fungal spores germinate and rapidly colonize the surrounding areas (Droby et al., 2008). The control of this pathogen relies heavily on the use of synthetic chemicals, but concerns regarding their potential negative effects on human health and also the generation of fungicide-resistant strains have encouraged finding alternatives, such as the generation of citrus trees with fruits that are genetically resistant to the pathogen.In this work, to better understand the mechanism underlying the constitutive resistance to P. digitatum conferred by the reduction of limonene in AS orange fruits, we analyzed the pattern of fruit growth and the morphological and biochemical developmental characteristics and performed a global analysis of gene expression using a 20K citrus microarray. The study is supplemented by examining the possible involvement of key hormone signals and isoprenoid precursors in the fruit peel. We report here that the reduced level of d-limonene in AS fruits is tightly associated with the constitutive activation of defense response signaling cascades. Our results establish, to our knowledge for the first time, a correlation between increased volatile terpene content and the decline of defense responses in a fleshy fruit during maturation, which would facilitate necrotroph fungal infections in citrus fruits.     

转基因改良植物抗真菌病害的策略及其进展

随着近几年来生物技术的迅猛发展,一系列新的广谱持久性抗病基因已经被鉴定、克隆并应用于转基因改良抗病性的实践。概述目前植物抗真菌病害的基因工程策略及其研究进展：（1）基于寄主－病原菌相互识别和信号传导体系的基因工程策略;（2）基于抗真菌蛋白的基因工程策略。     

Fungal Pathogens: The Battle for Plant Infection

The attempted infection of a plant by a pathogen, such as a fungus or an Oomycete, may be regarded as a battle whose major weapons are proteins and smaller chemical compounds produced by both organisms. Indeed, plants produce an astonishing plethora of defense compounds that are still being discovered at a rapid pace. This pattern arose from a multi-million year, ping-pong?type co-evolution, in which plant and pathogen successively added new chemical weapons in this perpetual battle. As each defensive innovation was established in the host, new ways to circumvent it evolved in the pathogen. This complex co-evolution process probably explains not only the exquisite specificity observed between many pathogens and their hosts, but also the ineffectiveness or redundancy of some defensive genes which often encode enzymes with overlapping activities. Plants evolved a complex, multi-level series of structural and chemical barriers that are both constitutive or preformed and inducible. These defenses may involve strengthening of the cell wall, hypersensitive response (HR), oxidative burst, phytoalexins and pathogenesis-related (PR) proteins. The pathogen must successfully overcome these obstacles before it succeeds in causing disease. In some cases, it needs to modulate or modify plant cell metabolism to its own benefit and/or to abolish defense reactions. Central to the activation of plant responses is timely perception of the pathogen by the plant. A crucial role is played by elicitors which, depending on their mode of action, are broadly classified into nonspecific elicitors and highly specific elicitors or virulence effector/avirulence factors. A protein battle for penetration is then initiated, marking the pathogen attempted transition from extracellular to invasive growth before parasitism and disease can be established. Three major types of defense responses may be observed in plants: non-host resistance, host resistance, and host pathogenesis. Plant innate immunity may comprise a continuum from non-host resistance involving the detection of general elicitors to host-specific resistance involving detection of specific elicitors by R proteins. It was generally assumed that non-host resistance was based on passive mechanisms and that nonspecific rejection usually arose as a consequence of the non-host pathogen failure to breach the first lines of plant defense. However, recent evidence has blurred the clear-cut distinction among non-host resistance, host-specific resistance and disease. The same obstacles are also serious challenges for host pathogens, reducing their success rate significantly in causing disease. Indeed, even susceptible plants mount a (insufficient) defense response upon recognition of pathogen elicited molecular signals. Recent evidence suggests the occurrence of significant overlaps between the protein components and signalling pathways of these types of resistance, suggesting the existence of both shared and unique features for the three branches of plant innate immunity.     

Resistance and Susceptibility of Plants to Fungal Pathogens

Plants are under continuous threat of infection by pathogens endowed with diverse strategies to colonize their host. Comprehensive biochemical and genetic approaches are now starting to reveal the complex signaling pathways that mediate plant disease resistance. Initiation of defense signaling often involves specific recognition of invading pathogens by the products of specialized host resistance (R) genes. Potential resistance signaling components have been identified by mutational analyses to be required for specific resistance in the model Arabidopsis and some crop species. Strikingly, many of the components share similarity to that of innate immune systems in animals. Evidence is also accumulating that plant pathogens have a number of ways to evade host defenses during the early stages of infection, similar to animal pathogens. These strategies are becoming much better understood in a number of plant–pathogen interactions. In this review, we focus on the current knowledge of host factors that control plant resistance and susceptibility to fungal pathogens. The knowledge accumulated in these studies will serve a fundamental basis for combating diseases in strategic molecular agriculture.     

OsJAR1 Contributes Mainly to Biosynthesis of the Stress-Induced Jasmonoyl-Isoleucine Involved in Defense Responses in Rice

Jasmonate plays key roles in plant growth and stress responses, as in defense against pathogen attack. Jasmonoyl-isoleucine (JA-Ile), a major active form of jasmonates, is thought to play a pivotal role in plant defense responses, but the involvement of JA-Ile in rice defense responses, including phytoalexin production, remains largely unknown. Here we found that OsJAR1 contributes mainly to stress-induced JA-Ile production by the use of an osjar1 Tos17 mutant. The osjar1 mutant was impaired in JA-induced expression of JA-responsive genes and phytoalexin production, and these defects were restored genetically. Endogenous JA-Ile was indispensable to the production of a flavonoid phytoalexin, sakuranetin, but not to that of diterpenoid phytoalexins in response to heavy metal stress and the rice blast fungus. The osjar1 mutant was also found to be more susceptible to the blast fungus than the parental wild type. These results suggest that JA-Ile production makes a contribution to rice defense responses with a great impact on stress-induced sakuranetin production.     

Correction to: A New Combination with D-Cateslytin to Eradicate Root Canal Pathogens

International Journal of Peptide Research and Therapeutics - The article A New Combination with D?Cateslytin to Eradicate Root Canal Pathogens, written by Claire Ehlinger, Pauline Dartevelle,...     

Host-Pathogen Interactions : XXXIII. A Plant Protein Converts a Fungal Pathogenesis Factor into an Elicitor of Plant Defense Responses

This paper describes the effect of a plant-derived polygalacturonase-inhibiting protein (PGIP) on the activity of endopolygalacturonases isolated from fungi. PGIP's effect on endopolygalacturonases is to enhance the production of oligogalacturonides that are active as elicitors of phytoalexin (antibiotic) accumulation and other defense reactions in plants. Only oligogalacturonides with a degree of polymerization higher than nine are able to elicit phytoalexin synthesis in soybean cotyledons. In the absence of PGIP, a 1-minute exposure of polygalacturonic acid to endopolygalacturonase resulted in the production of elicitor-active oligogalacturonides. However, the enzyme depolymerized essentially all of the polygalacturonic acid substrate to elicitor-inactive oligogalacturonides within 15 minutes. When the digestion of polygalacturonic acid was carried out with the same amount of enzyme but in the presence of excess PGIP, the rate of production of elicitor-active oligogalacturonides was dramatically altered. The amount of elicitor-active oligogalacturonide steadily increased for 24 hours. It was only after about 48 hours that the enzyme converted the polygalacturonic acid into short, elicitor-inactive oligomers. PGIP is a specific, reversible, saturable, high-affinity receptor for endopolygalacturonase. Formation of the PGIP-endopolygalacturonase complex results in increased concentrations of oligogalacturonides that activate plant defense responses. The interaction of the plant-derived PGIP with fungal endopolygalacturonases may be a mechanism by which plants convert endopolygalacturonase, a factor important for the virulence of pathogens, into a factor that elicits plant defense mechanisms.     

A Model Sheet Mineral System to Study Fungal Bioweathering of Mica

The general objective of this research was to examine fungal interactions with silicate minerals within the context of their roles in bioweathering. To achieve this, we used muscovite, a phyllosilicate mineral (KAl₂[(OH)₂|AlSi₃O₁₀]), in the form of a mineral sheet model system for ease of experimental manipulation and microscopic examination. It was found that test fungal species successfully colonized and degraded the surface of muscovite sheets in both laboratory and field experiments. After colonization by the common soil fungus Aspergillus niger, a network of hyphae covered the surface of the muscovite, and mineral dissolution or degradation was clearly evidenced by a network of fungal “footprints” that reflected coverage by the mycelium. For natural soil incubations, microorganisms associated with muscovite sheet material included biofilms of fungi and bacteria on the surface, while mineral encrustation or adhesion to microbial structures was also observed. Our results show that muscovite sheet is a good model mineral system for examination of microbial colonization and degradation, and this was demonstrated using laboratory and field systems, providing more evidence for the bioweathering significance of fungal activities in the context of silicate degradation and soil formation and development. The approach is also clearly applicable to other rock and mineral-based substrates and a variety of free-living and symbiotic microbial systems.     

Regulation of Plant Defense Response to Fungal Pathogens: Two Types of Protein Kinases in the Reversible Phosphorylation of the Host Plasma Membrane H+-ATPase

The role of reversible phosphorylation of the host plasma membrane H+-ATPase in signal transduction during the incompatible interaction between tomato cells and the fungal pathogen Cladosporium fulvum was investigated. Tomato cells (with the Cf-5 resistance gene) or isolated plasma membranes from Cf-5 cells treated with elicitor preparations from race 2.3 or 4 of C. fulvum (containing the avr5 gene product) showed a marked dephosphorylation of plasma membrane H+-ATPase. Similar treatment with elicitor preparations from races 5 and 2.4.5.9.11 (lacking the avr5 gene product) showed no change in dephosphorylation. Elicitor (race 4) treatment of cells, but not of isolated plasma membranes, for 2 hr resulted in rephosphorylation of the ATPase via Ca2+-dependent protein kinases. The initial (first hour) rephosphorylation was enhanced by protein kinase C (PKC) activators and was prevented by PKC inhibitors. Activity of a second kinase appeared after 1 hr and was responsible for the continuing phosphorylation of the H+-ATPase. This latter Ca2+-dependent kinase was inhibited by a calmodulin (CaM) antagonist and by an inhibitor of Ca2+/CaM-dependent protein kinase II. The activation of the Ca2+/CaM-dependent protein kinase depended on the prior activation of the PKC-like kinase.     

植物病原真菌对二甲酰亚胺类杀菌剂的抗性分子机制

综述了近年来国内外植物病原真菌对二甲酰亚胺类杀菌剂抗性机制研究的主要成果,包括：二甲酰亚胺类杀菌剂（DCFs）的杀菌机制、植物病原菌对DCFs抗药性的产生现状、促分裂原活化蛋白激酶（MAPK）途径和依赖环化腺苷酸（cAMP）的蛋白激酶途径在抗药性产生中的可能作用及相关的分子生物学研究进展。     

Fungal volatiles: an environmentally friendly tool to control pathogenic microorganisms in plants

Fungi are an extraordinary and immensely diverse group of microorganisms that colonize many habitats even competing with other microorganisms. Fungi have received recognition for interesting metabolic activities that have an enormous variety of biotechnological applications. Previously, volatile organic compounds produced by fungi (FVOCs) have been demonstrated to have a great capacity for use as antagonist products against plant pathogens. However, in recent years, FVOCs have been received attention as potential alternatives to the use of traditional pesticides and, therefore, as important eco-friendly biotechnological tools to control plant pathogens. Therefore, highlighting the current state of knowledge of these fascinating FVOCs, the actual detection techniques and the bioactivity against plant pathogens is essential to the discovery of new products that can be used as biopesticides.     

Root Endodermis and Exodermis: Structure, Function, and Responses to the Environment

Roots of virtually all vascular plants have an endodermis with a Casparian band, and the majority of angiosperm roots tested also have an exodermis with a Casparian band. Both the endodermis and exodermis may develop suberin lamellae and thick, tertiary walls. Each of these wall modifications has its own function(s). The endodermal Casparian band prevents the unimpeded movement of apoplastic substances into the stele and also prevents the backflow of ions that have moved into the stele symplastically and then were released into its apoplast. In roots with a mature exodermis, the barrier to apoplastic inflow of ions occurs near the root surface, but prevention of backflow of ions from the stele remains a function of the endodermis. The suberin lamellae protect against pathogen invasion and possibly root drying during times of stress. Tertiary walls of the endodermis and exodermis are believed to function in mechanical support of the root, but this idea remains to be tested. During stress, root growth rates decline, and the endodermis and exodermis develop closer to the root tip. In two cases, stress is known to induce the formation of an exodermis, and in several other cases to accelerate the development of both the exodermis and endodermis. The responses of the endodermis and exodermis to drought, exposure to moist air, flooding, salinity, ion deficiency, acidity, and mechanical impedance are discussed.     

Plant Root Responses to Three Abundant Soil Minerals: Silicon,Aluminum and Iron

Silicon (Si), aluminum (Al), and iron (Fe) are the three most abundant minerals in soil; however, their effects on plants differ because they are beneficial, toxic, and essential to plant growth, respectively. High accumulation of silicon in the shoots helps some plants to overcome a range of biotic and abiotic stresses. However, plants vary in their ability to take up Si from the soil and load it into the xylem and so the accumulation of silicon varies greatly between plant species. Aluminum toxicity is characterized by a rapid inhibition of root elongation but some species and even genotypes within species can tolerate Al toxicity better than others. While the mechanisms controlling this tolerance in most of the more resistant species are poorly understood, some plants are able to detoxify Al externally and/or internally by complexation with ligands or by pH changes in the rhizosphere. Iron is taken up from the soil by two efficient mechanisms called Strategy I and Strategy II, which operate in distinct phylogenic groups. Strategy I plants increase soil Fe solubility by releasing protons and reductants/chelators, such as organic acids and phenolics, into the rhizosphere, while Strategy II plants are characterized by the secretion of ferric chelating substances (phytosiderophores) coupled with a specific Fe³⁺: chelate uptake system. In this review, the molecular mechanisms underlying root response to Si, Al, and Fe are described.