OPDA, more than just a jasmonate precursor.

The oxylipin 12-oxo-phytodienoic acid (OPDA) is known as a biosynthetic precursor of the important plant hormone jasmonic acid. However, OPDA is also a signaling molecule with functions independent of jasmonates. OPDA involvement in diverse biological processes, from plant defense and stress responses to growth regulation and development, has been documented across plant species. OPDA is synthesized in the plastids from alpha-linolenic acid, and OPDA binding to plastidial cyclophilins activates TGA transcription factors upstream of genes associated with stress responses. Here, we summarize what is known about OPDA metabolism and signaling while briefly discussing its jasmonate dependent and independent roles. We also describe open questions, such as the OPDA protein interactome and biological roles of OPDA conjugates.

Ligand diversity contributes to the full activation of the jasmonate pathway in Marchantia polymorpha.

In plants, jasmonate signaling regulates a wide range of processes from growth and development to defense responses and thermotolerance. Jasmonates, such as jasmonic acid (JA), (+)-7-iso-jasmonoyl-l-isoleucine (JA-Ile), 12-oxo-10,15(Z)-phytodienoic acid (OPDA), and dinor-12-oxo-10,15(Z)-phytodienoic acid (dn-OPDA), are derived from C18 (18 Carbon atoms) and C16 polyunsaturated fatty acids (PUFAs), which are found ubiquitously in the plant kingdom. Bryophytes are also rich in C20 and C22 long-chain polyunsaturated fatty acids (LCPUFAs), which are found only at low levels in some vascular plants but are abundant in organisms of other kingdoms, including animals. The existence of bioactive jasmonates derived from LCPUFAs is currently unknown. Here, we describe the identification of an OPDA-like molecule derived from a C20 fatty acid (FA) in the liverwort Marchantia polymorpha (Mp), which we term (5Z,8Z)-10-(4-oxo-5-((Z)-pent-2-en-1-yl)cyclopent-2-en-1-yl)deca-5,8-dienoic acid (C20-OPDA). This molecule accumulates upon wounding and, when applied exogenously, can activate known Coronatine Insensitive 1 (COI1) -dependent and -independent jasmonate responses. Furthermore, we identify a dn-OPDA-like molecule (Δ4-dn-OPDA) deriving from C20-OPDA and demonstrate it to be a ligand of the jasmonate coreceptor (MpCOI1-Mp Jasmonate-Zinc finger inflorescence meristem domain [MpJAZ]) in Marchantia. By analyzing mutants impaired in the production of LCPUFAs, we elucidate the major biosynthetic pathway of C20-OPDA and Δ4-dn-OPDA. Moreover, using a double mutant compromised in the production of both Δ4-dn-OPDA and dn-OPDA, we demonstrate the additive nature of these molecules in the activation of jasmonate responses. Taken together, our data identify a ligand of MpCOI1 and demonstrate LCPUFAs as a source of bioactive jasmonates that are essential to the immune response of M. polymorpha.

The jasmonic acid-amino acid conjugates JA-Val and JA-Leu are involved in rice resistance to herbivores.

The phytohormone jasmonic acid (JA) plays a core role in plant defence against herbivores. When attacked by herbivores, JA and its bioactive derivatives are accumulated at the damage site, and subsequently perceived by the jasmonate co-receptors COI1 and JAZ proteins. The (+)-7-iso-jasmonoyl-L-isoleucine (JA-Ile) is known to be the main active JA derivative controlling vascular plant responses to herbivores as well as other JA-regulated processes. However, whether other endogenous JA-amino acid conjugates (JA-AAs) are involved in herbivore-induced defence responses remain unknown. Here, we investigated the role of herbivore-elicited JA-AAs in the crop plant rice. The levels of five JA-AAs were significantly increased under the armyworm, leaf folder and brown planthopper attack. Of the elicited JA derivatives, JA-Ile, JA-Val and JA-Leu could serve as ligands to promote the interaction between rice COI1 and JAZs, inducing OsJAZ4 degradation in vivo. JA-Val or JA-Leu treatment increased the expression of JA- and defence-related pathway genes but not JA-Ile levels, suggesting that these JA-AAs may directly function in JA signalling. Furthermore, the application of JA-Val or JA-Leu resulted in JA-mediated plant growth inhibition, while enhancing plant resistance to herbivore attack. This study uncovers that JA-Val and JA-Leu also play a role in rice defence against herbivores.

SARS-CoV-2 Fears Green: The Chlorophyll Catabolite Pheophorbide A Is a Potent Antiviral.

SARS-CoV-2 pandemic is having devastating consequences worldwide. Although vaccination advances at good pace, effectiveness against emerging variants is unpredictable. The virus has displayed a remarkable resistance to treatments and no drugs have been proved fully effective against COVID-19. Thus, despite the international efforts, there is still an urgent need for new potent and safe antivirals against SARS-CoV-2. Here, we exploited the enormous potential of plant metabolism using the bryophyte Marchantia polymorpha L. and identified a potent SARS-CoV-2 antiviral, following a bioactivity-guided fractionation and mass-spectrometry approach. We found that the chlorophyll derivative Pheophorbide a (PheoA), a porphyrin compound similar to animal Protoporphyrin IX, has an extraordinary antiviral activity against SARS-CoV-2, preventing infection of cultured monkey and human cells, without noticeable cytotoxicity. We also show that PheoA targets the viral particle, interfering with its infectivity in a dose- and time-dependent manner. Besides SARS-CoV-2, PheoA also displayed a broad-spectrum antiviral activity against enveloped RNA viral pathogens such as HCV, West Nile, and other coronaviruses. Our results indicate that PheoA displays a remarkable potency and a satisfactory therapeutic index, which together with its previous use in photoactivable cancer therapy in humans, suggest that it may be considered as a potential candidate for antiviral therapy against SARS-CoV-2.

Omega hydroxylated JA-Ile is an endogenous bioactive jasmonate that signals through the canonical jasmonate signaling pathway.

Jasmonates are fatty acid derivatives that control several plant processes including growth, development and defense. Despite the chemical diversity of jasmonates, only jasmonoyl-L-isoleucine (JA-Ile) has been clearly characterized as the endogenous ligand of the jasmonate co-receptors (COI1-JAZs) in higher plants. Currently, it is accepted that ω-hydroxylation of JA-Ile leads to inactivation of the molecule. This study shows that ω-hydroxylated JA-Ile (12-OH-JA-Ile) retains bioactivity and signals through the canonical JA-pathway. The results suggest that 12-OH-JA-Ile differentially activates a subset of JA-Ile co-receptors that may control and/or modulate particular jasmonate dependent responses. It is proposed that after a strong immune response mediated by JA-Ile, the ω-hydroxylated form modulates JA-Ile activated processes thereby improving plant resilience.

Flower-specific jasmonate signaling regulates constitutive floral defenses in wild tobacco.

Optimal defense (OD) theory predicts that within a plant, tissues are defended in proportion to their fitness value and risk of predation. The fitness value of leaves varies greatly and leaves are protected by jasmonate (JA)-inducible defenses. Flowers are vehicles of Darwinian fitness in flowering plants and are attacked by herbivores and pathogens, but how they are defended is rarely investigated. We used Nicotiana attenuata, an ecological model plant with well-characterized herbivore interactions to characterize defense responses in flowers. Early floral stages constitutively accumulate greater amounts of two well-characterized defensive compounds, the volatile (E)-α-bergamotene and trypsin proteinase inhibitors (TPIs), which are also found in herbivore-induced leaves. Plants rendered deficient in JA biosynthesis or perception by RNA interference had significantly attenuated floral accumulations of defensive compounds known to be regulated by JA in leaves. By RNA-seq, we found a JAZ gene, NaJAZi, specifically expressed in early-stage floral tissues. Gene silencing revealed that NaJAZi functions as a flower-specific jasmonate repressor that regulates JAs, (E)-α-bergamotene, TPIs, and a defensin. Flowers silenced in NaJAZi are more resistant to tobacco budworm attack, a florivore. When the defensin was ectopically expressed in leaves, performance of Manduca sexta larvae, a folivore, decreased. NaJAZi physically interacts with a newly identified NINJA-like protein, but not the canonical NINJA. This NINJA-like recruits the corepressor TOPLESS that contributes to the suppressive function of NaJAZi on floral defenses. This study uncovers the defensive function of JA signaling in flowers, which includes components that tailor JA signaling to provide flower-specific defense.

A succinct access to ω-hydroxylated jasmonates via olefin metathesis.

In higher plants, jasmonates are lipid-derived signaling molecules that control many physiological processes, including responses to abiotic stress, defenses against insects and pathogens, and development. Among jasmonates, ω-oxidized compounds form an important subfamily. The biological roles of these ω-modified derivatives are not fully understood, largely due to their limited availability. Herein, a brief (two-step), simple and efficient (>80% yield), versatile, gram-scalable, and environmentally friendly synthetic route to ω-oxidized jasmonates is described. The approach utilizes olefin cross-metathesis as the key step employing inexpensive, commercially available substrates and catalysts.

JA-Ile-macrolactones uncouple growth and defense in wild tobacco.

Small molecules capable of uncoupling growth-defense in plants are currently not known. In this study, for the first time, semi-synthetic analogues of the phytohormone JA-Ile are employed to uncouple growth and defense responses in wild tobacco. The JA-Ile analogues are easily synthesized from inexpensive substrates via olefin metathesis.

Auxin Is Rapidly Induced by Herbivore Attack and Regulates a Subset of Systemic, Jasmonate-Dependent Defenses.

Plant responses to herbivore attack are regulated by phytohormonal networks. To date, the role of the auxin indole-3-acetic acid (IAA) in this context is not well understood. We quantified and manipulated the spatiotemporal patterns of IAA accumulation in herbivore-attacked Nicotiana attenuata plants to unravel its role in the regulation of plant secondary metabolism. We found that IAA is strongly, rapidly, and specifically induced by herbivore attack. IAA is elicited by herbivore oral secretions and fatty acid conjugate elicitors and is accompanied by a rapid transcriptional increase of auxin biosynthetic YUCCA-like genes. IAA accumulation starts 30 to 60 s after local induction and peaks within 5 min after induction, thereby preceding the jasmonate (JA) burst. IAA accumulation does not require JA signaling and spreads rapidly from the wound site to systemic tissues. Complementation and transport inhibition experiments reveal that IAA is required for the herbivore-specific, JA-dependent accumulation of anthocyanins and phenolamides in the stems. In contrast, IAA does not affect the accumulation of nicotine or 7-hydroxygeranyllinalool diterpene glycosides in the same tissue. Taken together, our results uncover IAA as a rapid and specific signal that regulates a subset of systemic, JA-dependent secondary metabolites in herbivore-attacked plants.

The Aggregation Pheromone of Phyllotreta striolata (Coleoptera: Chrysomelidae) Revisited.

Aggregations of the striped flea beetle Phyllotreta striolata on their crucifer host plants are mediated by volatiles emitted from feeding males. The male-specific sesquiterpene, (6R,7S)-himachala-9,11-diene (compound A), was shown previously to be physiologically and behaviorally active, but compound A was attractive only when combined with unnaturally high doses of the host plant volatile allyl isothiocyanate (AITC) in field trapping experiments. This indicated that our understanding of the chemical communication in this species is incomplete. Another male-specific sesquiterpenoid, (3S,9R,9aS)-3-hydroxy-3,5,5,9-tetramethyl-5,6,7,8,9,9a-hexahydro-1H-benzo[7]annulen-2(3H)-one (compound G), has been reported from an American P. striolata population. We confirmed the presence of compound G, and investigated its interaction with compound A and AITC in a P. striolata population in Taiwan. Compound G was attractive to Taiwanese P. striolata in laboratory bioassays, but significantly more beetles were attracted to a blend of compounds A and G. Under the same conditions, P. striolata showed no preference for the blend of A and G combined with a range of doses of AITC over the sesquiterpenoid blend alone. The sesquiterpenoid blend was tested further in field trapping experiments and attracted significantly more beetles than traps baited with compound A and ecologically relevant amounts of AITC. We conclude that A and G are components of the male-specific aggregation pheromone of P. striolata in Taiwan, and that the attractiveness of the pheromone is not reliant on the presence of AITC. Our results further indicate that the male-specific sesquiterpenoid blends differ qualitatively between the Taiwanese and American populations of P. striolata.

Synthesis, metabolism and systemic transport of a fluorinated mimic of the endogenous jasmonate precursor OPC-8:0.

Jasmonates (JAs) are fatty acid derivatives that mediate many developmental processes and stress responses in plants. Synthetic jasmonate derivatives (commonly isotopically labeled), which mimic the action of the endogenous compounds are often employed as internal standards or probes to study metabolic processes. However, stable-isotope labeling of jasmonates does not allow the study of spatial and temporal distribution of these compounds in real time by positron emission tomography (PET). In this study, we explore whether a fluorinated jasmonate could mimic the action of the endogenous compound and therefore, be later employed as a tracer to study metabolic processes by PET. We describe the synthesis and the metabolism of (Z)-7-fluoro-8-(3-oxo-2-(pent-2-en-1-yl)cyclopentyl)octanoic acid (7F-OPC-8:0), a fluorinated analog of the JA precursor OPC-8:0. Like endogenous jasmonates, 7F-OPC-8:0 induces the transcription of marker jasmonate responsive genes (JRG) and the accumulation of jasmonates after its application to Arabidopsis thaliana plants. By using UHPLC-MS/MS, we could show that 7F-OPC-8:0 is metabolized in vivo similarly to the endogenous OPC-8:0. Furthermore, the fluorinated analog was successfully employed as a probe to show its translocation to undamaged systemic leaves when it was applied to wounded leaves. This result suggests that OPC-8:0 - and maybe other oxylipins - may contribute to the mobile signal which triggers systemic defense responses in plants. We highlight the potential of fluorinated oxylipins to study the mode of action of lipid-derived molecules in planta, either by conventional analytical methods or fluorine-based detection techniques.

Synthesis, structural characterization and biological activity of two diastereomeric JA-Ile macrolactones.

Jasmonates are phytohormones involved in a wide range of plant processes, including growth, development, senescence, and defense. Jasmonoyl-L-isoleucine (JA-Ile, 2), an amino acid conjugate of jasmonic acid (JA, 1), has been identified as a bioactive endogenous jasmonate. However, JA-Ile (2) analogues trigger different responses in the plant. ω-Hydroxylation of the pentenyl side chain leads to the inactive 12-OH-JA-Ile (3) acting as a "stop" signal. On the other hand, a lactone derivative of 12-OH-JA (5) (jasmine ketolactone, JKL) occurs in nature, although with no known biological function. Inspired by the chemical structure of JKL (6) and in order to further explore the potential biological activities of 12-modified JA-Ile derivatives, we synthesized two macrolactones (JA-Ile-lactones (4a) and (4b)) derived from 12-OH-JA-Ile (3). The biological activity of (4a) and (4b) was tested for their ability to elicit nicotine production, a well-known jasmonate dependent secondary metabolite. Both macrolactones showed strong biological activity, inducing nicotine accumulation to a similar extent as methyl jasmonate does in Nicotiana attenuata leaves. Surprisingly, the highest nicotine contents were found in plants treated with the JA-Ile-lactone (4b), which has (3S,7S) configuration at the cyclopentanone not known from natural jasmonates. Macrolactone (4a) is a valuable standard to explore for its occurrence in nature.

Neomycin inhibition of (+)-7-iso-jasmonoyl-L-isoleucine accumulation and signaling.

The majority of plant defenses against insect herbivores are coordinated by jasmonate (jasmonic acid, JA; (+)-7-iso-jasmonoyl-L-isoleucine, JA-Ile)-dependent signaling cascades. Insect feeding and mimicking herbivory by application of oral secretions (OS) from the insect induced both cytosolic Ca(2+) and jasmonate-phytohormone elevation in plants. Here it is shown that in Arabidopsis thaliana upon treatment with OS from lepidopteran Spodoptera littoralis larvae, the antibiotic neomycin selectively blocked the accumulation of OS-induced Ca(2+) elevation and level of the bioactive JA-Ile, in contrast to JA level. Furthermore, neomycin treatment affected the downstream expression of JA-Ile-responsive genes, VSP2 and LOX2, in Arabidopsis. The neomycin-dependent reduced JA-Ile level is partially due to increased CYP94B3 expression and subsequent JA-Ile turn-over to12-hydroxy-JA-Ile. It is neither due to the inhibition of the enzymatic conjugation process nor to substrate availability. Thus, blocking Ca(2+) elevation specifically controls JA-Ile accumulation and signaling, offering an insight into role of calcium in defense against insect herbivory.

Foliar endophytic fungi as potential protectors from pathogens in myrmecophytic Acacia plants.

In defensive ant-plant interactions myrmecophytic plants express reduced chemical defense in their leaves to protect themselves from pathogens, and it seems that mutualistic partners are required to make up for this lack of defensive function. Previously, we reported that mutualistic ants confer plants of Acacia hindsii protection from pathogens, and that the protection is given by the ant-associated bacteria. Here, we examined whether foliar endophytic fungi may potentially act as a new partner, in addition to mutualistic ants and their bacteria inhabitants, involved in the protection from pathogens in myrmecophytic Acacia plants. Fungal endophytes were isolated from the asymptomatic leaves of A. hindsii plants for further molecular identification of 18S rRNA gene. Inhibitory effects of fungal endophytes were tested against Pseudomonas plant pathogens. Our findings support a potential role of fungal endophytes in pathogen the protection mechanisms against pathogens in myrmecophytic plants and provide the evidence of novel fungal endophytes capable of biosynthesizing bioactive metabolites.