Light and singlet oxygen in plant defense against pathogens: phototoxic phenalenone phytoalexins

Plants defend themselves from pathogen infections or mechanical injury by a number of mechanisms, including the induced biosynthesis of antimicrobial secondary metabolites. These compounds, termed phytoalexins, represent a very economical way to counteract hazard, because the carbon and energy resources are diverted to phytoalexin synthesis only at the early period of attack and only at its site. The occurrence of phenalenone chromophores in phytoalexins of plants originally nonphototoxic suggests that these plants respond to pathogen attacks by biosynthesizing singlet oxygen photosensitizers able to use solar energy for defense. This concept may have implications for the development of novel crop protection strategies.     

Plant Antimicrobial Agents and Their Effects on Plant and Human Pathogens

To protect themselves, plants accumulate an armoury of antimicrobial secondary metabolites. Some metabolites represent constitutive chemical barriers to microbial attack (phytoanticipins) and others inducible antimicrobials (phytoalexins). They are extensively studied as promising plant and human disease-controlling agents. This review discusses the bioactivity of several phytoalexins and phytoanticipins defending plants against fungal and bacterial aggressors and those with antibacterial activities against pathogens affecting humans such as Pseudomonas aeruginosa and Staphylococcus aureus involved in respiratory infections of cystic fibrosis patients. The utility of plant products as “antibiotic potentiators” and “virulence attenuators” is also described as well as some biotechnological applications in phytoprotection.     

Effect of soybean phytoalexins on the herbivorous insects mexican bean beetle and soybean looper

Effects of soybean phytoalexins on the feeding of the soybean looper and Mexican bean beetle were investigated to test the hypothesis that phytoalexins might be a defense mechanism of plants against insects as well as against pathogens. Short-term behavioral responses to the phytoalexins were analyzed using dual-choice tests with phytoalexin-rich and phytoalexin-poor (control) tissues. Phytoalexin production was elicited with ultraviolet radiation. Results from the dual-choice tests indicated that 6th instar soybean looper larvae fed equally on the control and phytoalexin-rich tissues. Feeding by adult and 4th instar Mexican bean beetles, however, was strongly deterred by the phytoalexins as evidenced by “single-bite” mandible scars on the phytoalexin-rich cotyledon discs. Nutritional effects of the isoflavonoid phytoalexin glyceollin on early instar soybean looper larvae were tested by incorporating the phytoalexin into an artificial medium at a level of 1% dry weight (0.15% fresh weight). The larvae were reared for 7 days from emergence on diets of control and glyceollin-containing media. Although survival on the glyceollin diets was initially less than on the control diets, under the experimental conditions glyceollin had no significant effect on the growth, development, or subsequent survival of the larvae. Efficiency of food utilization (ECI) was reduced, indicating that the phytoalexins may be a mild digestibility-reducing factor for the loopers. Implications of the results for host-plant resistance are discussed.     

Chemical Diversity and Defence Metabolism: How Plants Cope with Pathogens and Ozone Pollution

Chemical defences represent a main trait of the plant innate immune system. Besides regulating the relationship between plants and their ecosystems, phytochemicals are involved both in resistance against pathogens and in tolerance towards abiotic stresses, such as atmospheric pollution. Plant defence metabolites arise from the main secondary metabolic routes, the phenylpropanoid, the isoprenoid and the alkaloid pathways. In plants, antibiotic compounds can be both preformed (phytoanticipins) and inducible (phytoalexins), the former including saponins, cyanogenic glycosides and glucosinolates. Chronic exposure to tropospheric ozone (O₃) stimulates the carbon fluxes from the primary to the secondary metabolic pathways to a great extent, inducing a shift of the available resources in favour of the synthesis of secondary products. In some cases, the plant defence responses against pathogens and environmental pollutants may overlap, leading to the unspecific synthesis of similar molecules, such as phenylpropanoids. Exposure to ozone can also modify the pattern of biogenic volatile organic compounds (BVOC), emitted from plant in response to herbivore feeding, thus altering the tritrophic interaction among plant, phytophagy and their natural enemies. Finally, the synthesis of ethylene and polyamines can be regulated by ozone at level of S-adenosylmethionine (SAM), the biosynthetic precursor of both classes of hormones, which can, therefore, mutually inhibit their own biosynthesis with consequence on plant phenotype.     

New multifunctional sulfonamide-containing azobenzene chromophores: Synthesis and photochromic properties

The work presents synthesis and characterization of novel sulfadimethoxine and sulfabenzamide azo derivatives as well as the kinetic study of their trans-cis-trans isomerization. Spectral properties and kinetic constants were determined for the chromophores dispersed in poly(methyl methacrylate-co-butyl methacrylate) deposited on glass substrates in form of transparent, thin films. The maximum absorption bands were observed within wavelength in range of 446-457 nm for N-disubstituted aminoazobenzene derivatives and 361-362 nm for phenolic chromophores. For the phenolic azo derivatives of sulfadimethoxine and sulfabenzamide the photoisomerization induced with UV light followed the first order kinetics, while for other compounds the process combined of two parallel reactions with different rate constants was observed. The rate constants of thermal relaxation in the dark were usually few times smaller than those determined for photoinduced trans-cis isomerization. The change of real part of refractive index upon illumination determined by ellipsometry was within the range of 0.0043-0.0138.     

Synergistic Action of Antimicrobial Lung Proteins against Klebsiella pneumoniae

As key components of innate immunity, lung antimicrobial proteins play a critical role in warding off invading respiratory pathogens. Lung surfactant protein A (SP-A) exerts synergistic antimicrobial activity with the N-terminal segment of the SP-B proprotein (SP-B^N) against Klebsiella pneumoniae K2 in vivo. However, the factors that govern SP-A/SP-B^N antimicrobial activity are still unclear. The aim of this study was to identify the mechanisms by which SP-A and SP-B^N act synergistically against K. pneumoniae, which is resistant to either protein alone. The effect of these proteins on K. pneumoniae was studied by membrane permeabilization and depolarization assays and transmission electron microscopy. Their effects on model membranes of the outer and inner bacterial membranes were analyzed by differential scanning calorimetry and membrane leakage assays. Our results indicate that the SP-A/SP-B^N complex alters the ultrastructure of K. pneumoniae by binding to lipopolysaccharide molecules present in the outer membrane, forming packing defects in the membrane that may favor the translocation of both proteins to the periplasmic space. The SP-A/SP-B^N complex depolarized and permeabilized the inner membrane, perhaps through the induction of toroidal pores. We conclude that the synergistic antimicrobial activity of SP-A/SP-B^N is based on the capability of this complex, but not either protein alone, to alter the integrity of bacterial membranes.     

Drought Stress Acclimation Imparts Tolerance to Sclerotinia sclerotiorum and Pseudomonas syringae in Nicotiana benthamiana

Acclimation of plants with an abiotic stress can impart tolerance to some biotic stresses. Such a priming response has not been widely studied. In particular, little is known about enhanced defense capacity of drought stress acclimated plants to fungal and bacterial pathogens. Here we show that prior drought acclimation in Nicotiana benthamiana plants imparts tolerance to necrotrophic fungus, Sclerotinia sclerotiorum, and also to hemi-biotrophic bacterial pathogen, Pseudomonas syringae pv. tabaci. S. sclerotiorum inoculation on N. benthamiana plants acclimated with drought stress lead to less disease-induced cell death compared to non-acclimated plants. Furthermore, inoculation of P. syringae pv. tabaci on N. benthamiana plants acclimated to moderate drought stress showed reduced disease symptoms. The levels of reactive oxygen species (ROS) in drought acclimated plants were highly correlated with disease resistance. Further, in planta growth of GFPuv expressing P. syringae pv. tabaci on plants pre-treated with methyl viologen showed complete inhibition of bacterial growth. Taken together, these experimental results suggested a role for ROS generated during drought acclimation in imparting tolerance against S. sclerotiorum and P. syringae pv. tabaci. We speculate that the generation of ROS during drought acclimation primed a defense response in plants that subsequently caused the tolerance against the pathogens tested.     

Silencing an E3 Ubiquitin Ligase Gene OsJMJ715 Enhances the Resistance of Rice to a Piercing-Sucking Herbivore by Activating ABA and JA Signaling Pathways

The RING-type E3 ubiquitin ligases play an important role in plant growth, development, and defense responses to abiotic stresses and pathogens. However, their roles in the resistance of plants to herbivorous insects remain largely unknown. In this study, we isolated the rice gene OsJMJ715, which encodes a RING-domain containing protein, and investigated its role in rice resistance to brown planthopper (BPH, Nilaparvata lugens). OsJMJ715 is a nucleus-localized E3 ligase whose mRNA levels were upregulated by the infestation of gravid BPH females, mechanical wounding, and treatment with JA or ABA. Silencing OsJMJ715 enhanced BPH-elicited levels of ABA, JA, and JA-Ile as well as the amount of callose deposition in plants, which in turn increased the resistance of rice to BPH by reducing the feeding of BPH and the hatching rate of BPH eggs. These findings suggest that OsJMJ715 negative regulates the BPH-induced biosynthesis of ABA, JA, and JA-Ile and that BPH benefits by enhancing the expression of OsJMJ715.     

Maternal and Environmental Effects on Symbiont-Mediated Antimicrobial Defense

Bacteria produce a remarkable diversity of bioactive molecules with antimicrobial properties. Despite the importance of such compounds for human medicine, little is known about the factors influencing antibiotic production in natural environments. Recently, several insects have been found to benefit from symbiont-produced antimicrobial compounds for defense against pathogenic microbes. In the European beewolf, Philanthus triangulum (Hymenoptera, Crabronidae), bacteria of the genus Streptomyces provide protection against pathogens by producing antimicrobials on the larval cocoon during hibernation, thereby significantly enhancing the survival probability of the beewolf larva. To investigate the effects of abiotic and biotic factors on antibiotic production, we exposed beewolf cocoons to different environmental conditions and quantified the amount of Streptomyces-produced antibiotics by using gas chromatography/mass spectrometry (GC/MS). The results revealed no significant influence of temperature, humidity, or pathogen load on the antibiotic amount, indicating that antibiotic production is not affected by current environmental conditions but rather may be optimized to serve as a reliable long-term protection during the unpredictable phase of beewolf hibernation. However, the amount of antibiotics was positively correlated with the symbiont population size on the cocoon, which in turn is affected by the number of Streptomyces cells provided by the mother into the brood cell. Additionally, we found a positive correlation between the amount of hydrocarbons and the number and length of bacterial cells in the antennal gland secretion, suggesting that maternal investment affects symbiont growth and, thus, antibiotic production on the larval cocoon.     

Plant Secondary Metabolites with an Overview of Populus

Populus trees meet continuous difficulties from the environment through their life cycle. To warrant their durability and generation, Populus trees exhibit various types of defenses, including the production of secondary metabolites. Syntheses derived from the shikimate-phenylpropanoid pathway are a varied and plentiful class of secondary metabolites manufactured in Populus. Amongst other main classes of secondary metabolites in Populus are fatty acid and terpenoid-derivatives. Many of the secondary metabolites made by Populus trees have been functionally described. Any others have been associated with particular ecological or biological processes, such as resistance against pests and microbial pathogens or acclimatization to abiotic stresses. Still, the functions of many Populus secondary metabolites are incompletely understood. Furthermore, many secondary metabolites have therapeutic effects, leading to more studies of secondary metabolites and their biosynthesis. This paper reviews the biosynthetic pathways and therapeutic impacts of secondary metabolites in Populus using a genomics approach. Compared with bacteria, fewer known pathways produce secondary metabolites in Populus despite P. trichocarpa having had its genome sequenced.     

NaCl-Induced Elicitation Alters Physiology and Increases Accumulation of Phenolic Compounds in Melissa officinalis L.

In nature, plants usually produce secondary metabolites as a defense mechanism against environmental stresses. Different stresses determine the chemical diversity of plant-specialized metabolism products. In this study, we applied an abiotic elicitor, i.e., NaCl, to enhance the biosynthesis and accumulation of phenolic secondary metabolites in Melissa officinalis L. Plants were subjected to salt stress treatment by application of NaCl solutions (0, 50, or 100 mM) to the pots. Generally, the NaCl treatments were found to inhibit the growth of plants, simultaneously enhancing the accumulation of phenolic compounds (total phenolics, soluble flavonols, anthocyanins, phenolic acids), especially at 100 mM NaCl. However, the salt stress did not disturb the accumulation of photosynthetic pigments and proper functioning of the PS II photosystem. Therefore, the proposed method of elicitation represents a convenient alternative to cell suspension or hydroponic techniques as it is easier and cheaper with simple application in lemon balm pot cultivation. The improvement of lemon balm quality by NaCl elicitation can potentially increase the level of health-promoting phytochemicals and the bioactivity of low-processed herbal products.     

Protease Inhibitors from Plants with Antimicrobial Activity

Antimicrobial proteins (peptides) are known to play important roles in the innate host defense mechanisms of most living organisms, including plants, insects, amphibians and mammals. They are also known to possess potent antibiotic activity against bacteria, fungi, and even certain viruses. Recently, the rapid emergence of microbial pathogens that are resistant to currently available antibiotics has triggered considerable interest in the isolation and investigation of the mode of action of antimicrobial proteins (peptides). Plants produce a variety of proteins (peptides) that are involved in the defense against pathogens and invading organisms, including ribosome-inactivating proteins, lectins, protease inhibitors and antifungal peptides (proteins). Specially, the protease inhibitors can inhibit aspartic, serine and cysteine proteinases. Increased levels of trypsin and chymotrypsin inhibitors correlated with the plants resistance to the pathogen. Usually, the purification of antimicrobial proteins (peptides) with protease inhibitor activity was accomplished by salt-extraction, ultrafiltration and C₁₈ reverse phase chromatography, successfully. We discuss the relation between antimicrobial and anti-protease activity in this review. Protease inhibitors from plants potently inhibited the growth of a variety of pathogenic bacterial and fungal strains and are therefore excellent candidates for use as the lead compounds for the development of novel antimicrobial agents.     

Sulfur-Containing Secondary Metabolites from Arabidopsis thaliana and other Brassicaceae with Function in Plant Immunity

Biosynthesis of antimicrobial secondary metabolites in response to microbial infection is one of the features of the plant immune system. Particular classes of plant secondary metabolites involved in plant defence are often produced only by species belonging to certain phylogenetic clades. Brassicaceae plants have evolved the ability to synthesise a wide range of sulfur-containing secondary metabolites, including glucosinolates and indole-type phytoalexins. A subset of these compounds is produced by the model plant Arabidopsis thaliana. Genetic tools available for this species enabled verification of immune functions of glucosinolates and camalexin (A. thaliana phytoalexin), as well as characterisation of their respective biosynthetic pathways. Current knowledge of the biosynthesis of Brassicaceae sulfur-containing metabolites suggests that the key event in the evolution of these compounds is the acquisition of biochemical mechanisms originating from detoxification pathways into secondary metabolite biosynthesis. Moreover, it is likely that glucosinolates and Brassicaceae phytoalexins, traditionally considered as separate groups of compounds, have a common evolutionary origin and are interconnected on the biosynthetic level. This suggests that the diversity of Brassicaceae sulfur-containing phytochemicals reflect phylogenetic clade-specific branches of an ancient biosynthetic pathway.     

Yeast α-Glucosidase Inhibitory Phenolic Compounds Isolated from Gynura medica Leaf

Gynura medica leaf extract contains significant amounts of flavonols and phenolic acids and exhibits powerful hypoglycemic activity against diabetic rats in vivo. However, the hypoglycemic active constituents that exist in the plant have not been fully elaborated. The purpose of this study is to isolate and elaborate the hypoglycemic activity compounds against inhibition the yeast α-glucosidase in vitro. Seven phenolic compounds including five flavonols and two phenolic acids were isolated from the leaf of G. medica. Their structures were identified by the extensive NMR and mass spectral analyses as: kaempferol (1), quercetin (2), kaempferol-3-O-β-D-glucopyranoside (3), kaempferol-3-O-rutinoside (4), rutin (5), chlorogenic acid (6) and 3,5-dicaffeoylquinic acid methyl ester (7). All of the compounds except 1 and 3 were isolated for the first time from G. medica. Compounds 1–7 were also assayed for their hypoglycemic activity against yeast α-glucosidase in vitro. All of the compounds except 1 and 6 showed good yeast α-glucosidase inhibitory activity with the IC₅₀ values of 1.67 mg/mL, 1.46 mg/mL, 0.38 mg/mL, 0.10 mg/mL and 0.53 mg/mL, respectively.     

Stress-Responsive Expression,Subcellular Localization and Protein–Protein Interactions of the Rice Metacaspase Family

Metacaspases, a class of cysteine-dependent proteases like caspases in animals, are important regulators of programmed cell death (PCD) during development and stress responses in plants. The present study was focused on comprehensive analyses of expression patterns of the rice metacaspase (OsMC) genes in response to abiotic and biotic stresses and stress-related hormones. Results indicate that members of the OsMC family displayed differential expression patterns in response to abiotic (e.g., drought, salt, cold, and heat) and biotic (e.g., infection by Magnaporthe oryzae, Xanthomonas oryzae pv. oryzae and Rhizoctonia solani) stresses and stress-related hormones such as abscisic acid, salicylic acid, jasmonic acid, and 1-amino cyclopropane-1-carboxylic acid (a precursor of ethylene), although the responsiveness to these stresses or hormones varies to some extent. Subcellular localization analyses revealed that OsMC1 was solely localized and OsMC2 was mainly localized in the nucleus. Whereas OsMC3, OsMC4, and OsMC7 were evenly distributed in the cells, OsMC5, OsMC6, and OsMC8 were localized in cytoplasm. OsMC1 interacted with OsLSD1 and OsLSD3 while OsMC3 only interacted with OsLSD1 and that the zinc finger domain in OsMC1 is responsible for the interaction activity. The systematic expression and biochemical analyses of the OsMC family provide valuable information for further functional studies on the biological roles of OsMCs in PCD that is related to abiotic and biotic stress responses.     

Gene Expression of Type VI Secretion System Associated with Environmental Survival in Acidovorax avenae subsp. avenae by Principle Component Analysis

Valine glycine repeat G (VgrG) proteins are regarded as one of two effectors of Type VI secretion system (T6SS) which is a complex multi-component secretion system. In this study, potential biological roles of T6SS structural and VgrG genes in a rice bacterial pathogen, Acidovorax avenae subsp. avenae (Aaa) RS-1, were evaluated under seven stress conditions using principle component analysis of gene expression. The results showed that growth of the pathogen was reduced by H₂O₂ and paraquat-induced oxidative stress, high salt, low temperature, and vgrG mutation, compared to the control. However, pathogen growth was unaffected by co-culture with a rice rhizobacterium Burkholderia seminalis R456. In addition, expression of 14 T6SS structural and eight vgrG genes was significantly changed under seven conditions. Among different stress conditions, high salt, and low temperature showed a higher effect on the expression of T6SS gene compared with host infection and other environmental conditions. As a first report, this study revealed an association of T6SS gene expression of the pathogen with the host infection, gene mutation, and some common environmental stresses. The results of this research can increase understanding of the biological function of T6SS in this economically-important pathogen of rice.