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Diterpenoid Phytoalexin Factor (DPF) is a key transcription factor involved in DP biosynthesis under non-stressed conditions in rice (Oryza sativa L.). Using CRISPR/Cas9, DPF knockout rice lines were generated. Treatments with abiotic stresses (copper chloride, ultraviolet light, and jasmonic acid) and biotic stress (blast fungus infection) to the knockout lines revealed that the DPF positively regulates stress-induced DP biosynthesis.

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Biosynthesis of the phytoalexins scopoletin and scopolin in Nicotiana species is regulated by upstream signals including jasmonate (JA), ethylene (ET) and NaWRKY3 in response to the necrotrophic fungus Alternaria alternata, which causes brown spot disease. However, how these signals are coordinated to regulate these phytoalexins remains unknown. By analyzing RNA sequencing data and RNA interference, we identified NaERF1B-like (NaERF1B-L) as a key player in Nicotiana attenuata during A. alternata infection by regulating the transcripts of Feruloyl-CoA 6'-hydroxylase 1 (NaF6'H1), encoding a key enzyme for scopoletin biosynthesis, and NaVS1-like (NaVS1-L), a putative biosynthetic gene of the phytoalexin solavetivone. We further demonstrated that the synergistic induction of these two genes by JA and ET signaling is mediated by NaERF1B-L. Additionally, we found that the two closely related proteins NaWRKY6 and NaWRKY3 physically interact to enhance NaERF1B-L expression by directly binding and activating the NaERF1B-L promoter. Collectively, our current results demonstrate that NaERF1B-L plays a positive role in resistance to A. alternata by modulating phytoalexins biosynthesis through the integration of JA/ET and NaWRKY6/3 signaling. Our findings reveal a fine-tuned transcriptional regulatory hierarchy mediated by NaERF1B-L for brown spot disease resistance in wild tobacco.

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Cultivated rice (Oryza sativa) produces a variety of diterpenoid-type phytoalexins. Diterpene synthase genes that are responsible for the biosynthesis of momilactones, phytocassanes, and oryzalexins have been identified in O. sativa cv. Nipponbare. OsKSL10 (Os12t0491800 in RAP and LOC_Os12g30824 in MSU) was previously identified as an enzyme catalyzing the conversion of ent-copalyl diphosphate to ent-sandaracopimaradiene for the production of oryzalexins A to F. Our previous study on Oryza rufipogon, a wild progenitor of Asian cultivated rice, showed that both OrKSL10 and OrKSL10ind from O. rufipogon accessions W1943 and W0106, respectively, closely related to the japonica and indica subspecies, converted ent-copalyl diphosphate to ent-miltiradiene. Thus, the functional conversion of ent-miltiradiene synthase into ent-sandaracopimaradiene synthase is implied to have occurred through natural amino acid mutations, the details of which have not been elucidated. In this study, we show that introduction of A654G substitution into OrKSL10 significantly alters its function into more closely resembling that of OsKSL10. Moreover, double substitution V546I/A654G almost completely converts the function of OrKSL10 into that of OsKSL10. On the other hand, the reversed substitution I546V/G654A was insufficient to convert the function of OsKSL10 into OrKSL10, indicating the introduction of additional substitution S522I is required for the functionality of OsKSL10. Lastly, point mutations at the 654A residue in OrKSL10 suggest that hydrophobic side chains at this position have a negative influence on the production of ent-sandaracopimaradiene.

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ß-Costic acid is a sesquiterpene phytoalexin with acaricidal activity against Varroa destructor and antitrypanosomal activity. A concise and efficient method was developed for the synthesis of ß-costic acid via the allylic oxidation of ß-selinene, a component of celery seed oil.

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Rice blast, caused by Magnaporthe oryzae (M. oryzae), is one of the most common and damaging diseases of rice that limits rice yield and quality. The mediator complex plays a vital role in promoting transcription by bridging specific transcription factors and RNA polymerase II. Here, we show that the rice mediator subunit OsMED16 is essential for full induction of the diterpenoid phytoalexin biosynthesis genes and resistance to the ascomycetous fungus M. oryzae. Mutants of Osmed16 show reduced expression of the DP biosynthesis genes and are markedly more susceptible to M. oryzae, while transgenic plants overexpressing OsMED16 increased the expression of the DP biosynthesis genes and significantly enhanced resistance to M. oryzae. Interestingly, OsMED16 is physically associated with the WRKY family transcription factor OsWRKY45, which interacts with the phytoalexin synthesis key regulator transcription factor OsWRKY62. Further, OsMED16-OsWRKY45-OsWRKY62 complex could bind to the promoter regions of phytoalexin synthesis-related genes and activate their gene expression. Our results show that OsMED16 may enhance rice tolerance to M. oryzae via directly manipulating phytoalexin de novo biosynthesis.

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Jasmonate (JA) and abscisic acid (ABA) are two major phytohormones involved in pathogen resistance. However, how their biosynthesis is regulated is not well understood. We silenced NaWRKY70 in wild tobacco Nicotiana attenuata and determined its role in regulating genes involved in the production of JA, ABA and the phytoalexin capsidiol in response to the fungal pathogen Alternaria alternata using techniques including electrophoretic mobility shift, chromatin immunoprecipitation, transient overexpression and virus-induced gene silencing. Silencing NaWRKY70 dramatically reduced both basal and A. alternata-induced jasmonoyl-isoleucine (JA-Ile) and ABA. Further evidence showed that NaWRKY70 directly binds to the W-boxes of the promoters of NaAOS and NaJAR4 (JA biosynthesis), NaNCED1 and NaXD1-like (ABA biosynthesis), and NaMPK4 (ABA signaling) to activate their expression, while binding but repressing the expression of NaCYP707A4-like3 (ABA degradation). Additionally, NaWRKY70 regulates capsidiol production through its key enzyme genes NaEASs and NaEAHs, and interacts with its regulator NaERF2-like to enhance their expression, whereas ABA negatively regulates capsidiol biosynthesis. Our results highlight the key role of NaWRKY70 in controlling both JA-Ile and ABA production, as well as capsidiol production, thus providing new insight into the defense mechanism of plant resistance to A. alternata.

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Glyceollins, a family of phytoalexins elicited in legume species, play crucial roles in environmental stress response (e.g., defending against pathogens) and human health. However, little is known about the genetic basis of glyceollin elicitation. In the present study, we employed a metabolite-based genome-wide association (mGWA) approach to identify candidate genes involved in glyceollin elicitation in genetically diverse and understudied wild soybeans subjected to soybean cyst nematode. In total, eight SNPs on chromosomes 3, 9, 13, 15, and 20 showed significant associations with glyceollin elicitation. Six genes fell into two gene clusters that encode glycosyltransferases in the phenylpropanoid pathway and were physically close to one of the significant SNPs (ss715603454) on chromosome 9. Additionally, transcription factors (TFs) genes such as MYB and WRKY were also found as promising candidate genes within close linkage to significant SNPs on chromosome 9. Notably, four significant SNPs on chromosome 9 show epistasis and a strong signal for selection. The findings describe the genetic foundation of glyceollin biosynthesis in wild soybeans; the identified genes are predicted to play a significant role in glyceollin elicitation regulation in wild soybeans. Additionally, how the epistatic interactions and selection influence glyceollin variation in natural populations deserves further investigation to elucidate the molecular mechanism of glyceollin biosynthesis.

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The natural variation of plant-specialized metabolites represents the evolutionary adaptation of plants to their environments. However, the molecular mechanisms that account for the diversification of the metabolic pathways have not been fully clarified. Rice plants resist attacks from pathogens by accumulating diterpenoid phytoalexins. It has been confirmed that the composition of rice phytoalexins exhibits numerous natural variations. Major rice phytoalexins (momilactones and phytocassanes) are accumulated in most cultivars, although oryzalactone is a cultivar-specific compound. Here, we attempted to reveal the evolutionary trajectory of the diversification of phytoalexins by analyzing the oryzalactone biosynthetic gene in Oryza species. The candidate gene, KSLX-OL, which accounts for oryzalactone biosynthesis, was found around the single-nucleotide polymorphisms specific to the oryzalactone-accumulating cultivars in the long arm of chromosome 11. The metabolite analyses in Nicotiana benthamiana and rice plants overexpressing KSLX-OL indicated that KSLX-OL is responsible for the oryzalactone biosynthesis. KSLX-OL is an allele of KSL8 that is involved in the biosynthesis of another diterpenoid phytoalexin, oryzalexin S and is specifically distributed in the AA genome species. KSLX-NOL and KSLX-bar, which encode similar enzymes but are not involved in oryzalactone biosynthesis, were also found in AA genome species. The phylogenetic analyses of KSLXs, KSL8s, and related pseudogenes (KSL9s) indicated that KSLX-OL was generated from a common ancestor with KSL8 and KSL9 via gene duplication, functional differentiation, and gene fusion. The wide distributions of KSLX-OL and KSL8 in AA genome species demonstrate their long-term coexistence beyond species differentiation, suggesting a balancing selection between the genes.

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Resveratrol (3, 5, 4'-trihydroxystilbene) is a polyphenolic derivative with herbal origin. It has attracted considerable attention in recent decades. Many studies have revealed the benefits of Resveratrol over several human disease models, including heart and neurological diseases, nephroprotective, immune regulation, antidiabetic, anti-obesity, age-related diseases, antiviral, and anticancer in experimental and clinical conditions. Recently, the antioxidant and anti-inflammatory activities of Resveratrol have been observed, and it has been shown that Resveratrol reduces inflammatory biomarkers, such as tissue degradation factor, cyclooxygenase 2, nitric oxide synthase, and interleukins. All of these activities appear to be dependent on its structural properties, such as the number and position of the hydroxyl group, which regulates oxidative stress, cell death, and inflammation. Resveratrol is well tolerated and safe even at higher pharmacological doses and desirably affects cardiovascular, neurological, and diabetic diseases. Consequently, it is plausible that Resveratrol can be regarded as a beneficial nutritional additive and a complementary drug, particularly for therapeutic applications. The present review provides an overview of currently available investigations on preventive and therapeutic characteristics and the main molecular mechanisms of Resveratrol and its potent derivatives in various diseases. Thus, this review would enhance knowledge and information about Resveratrol and encourage researchers worldwide to consider it as a pharmaceutical drug to struggle with future health crises against different human disorders.

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Production of the phytoalexins scopoletin and scopolin is regulated by jasmonate (JA) and ethylene signaling in Nicotiana species in response to Alternaria alternata, the necrotrophic fungal pathogen that causes brown spot disease. However, how these two signaling pathways are coordinated to control this process remains unclear. In this study, we found that the levels of these two phytoalexins and transcripts of their key enzyme gene, feruloyl-CoA 6'-hydroxylase 1 (NaF6'H1), were synergistically induced in Nicotiana attenuata by co-treatment with methyl jasmonate (MeJA) and ethephon. By combination of RNA sequencing and virus-induced gene silencing, we identified a WRKY transcription factor, NaWRKY70, which had a similar expression pattern to NaF6'H1 and was responsible for A. alternata-induced NaF6'H1 expression. Further evidence from stable transformed plants with RNA interference, knock out and overexpression of NaWRKY70 demonstrated that it is a key player in the synergistic induction of phytoalexins and plant resistance to A. alternata. Electrophoretic mobility shift, chromatin immunoprecipitation-quantitative PCR, and dual-luciferase assays revealed that NaWRKY70 can bind directly to the NaF6'H1 promoter and activate its expression. Furthermore, the key regulator of the ethylene pathway, NaEIN3-like1, can directly bind to the NaWRKY70 promoter and activate its expression. Meanwhile, NaMYC2s, important JA pathway transcription factors, also indirectly regulate the expression of NaWRKY70 and NaF6'H1 to control scopoletin and scopolin production. Our data reveal that these phytoalexins are synergistically induced by JA and ethylene signaling during A. alternata infection, which is largely mediated by NaWRKY70, thus providing new insights into the defense responses against A. alternata in Nicotiana species.

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BACKGROUND: Resveratrol's structural similarity to commercialized anti-breast cancer medications such as Tamoxifen underlines its potential as a promising option for developing successful anti-breast cancer drugs. However, the pharmacokinetic issues associated with resveratrol, such as its low bioavailability, have piqued the attention of researchers in developing novel derivatives. METHODS: A novel phytoalexin derivative, RsvD1, was successfully synthesized using resveratrol extracted from green grape peels as a precursor to investigate its anti-breast cancer efficacy on Estrogen receptor (ER) positive and negative breast cancer cells. RESULTS: The comparative analysis revealed that RsvD1 exhibited remarkable radical scavenging ability (IC50 = 2.21 µg/mL), surpassing the control, Trolox (IC50 = 6.3 µg/mL). Furthermore, RsvD1 demonstrated enhanced and selective antiproliferative activity against ER-positive MCF-7 cells (IC50 = 20.09 µg/mL) compared to resveratrol, the parent molecule (IC50 = 30.90 µg/mL). Further investigations unveiled that RsvD1 induced apoptosis and DNA damage in MCF-7 cells, leading to cell cycle arrest at the G0/G1 phase after 24 hours of incubation. RTqPCR gene expression analysis indicated that RsvD1 down-regulated the CAXII (ER-dependent) genes. In silico predictions demonstrated that RsvD1 possesses promising potential as a drug candidate due to its drug-like characteristics and favourable ADMET profile. Moreover, molecular docking studies provided insights into the theoretical binding mode between RsvD1 and ERα protein. CONCLUSION: The study highlights the therapeutic potential of the synthesized resveratrol derivative, RsvD1, positioning it as a promising scaffold for developing novel analogues with improved therapeutic properties and selectivity, specifically targeting ER+ breast cancer cells. Moreover, the compound's non-cytotoxic yet antiproliferative properties, coupled with its capability to induce programmed cell death and cell cycle arrest, enhance its potential as a highly effective drug candidate. As a result, this paves a promising path for the development of innovative and selective inhibitors targeting ER+ breast cancer with enhanced efficacy.

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With the aim of discovering novel and effective antifungal agents derived from natural sources, a series of new biphenyls based on natural biphenyl phytoalexins were designed, synthesized and evaluated for their antifungal activities against four invasive fungi. By modifying the two benzene rings of noraucuparin, a well-known biphenyl phytoantitoxin, some promising compounds with remarkable antifungal activity were discovered. Notably, compounds 23a, 23e and 23h exhibited potent activities and a broad antifungal spectrum with low MICs of 0.25-16 µg/mL, which were 8-256-fold more potent than that of the lead compound noraucuparin. Particularly, they displayed comparable potency to the positive control amphotericin B against Cryptococcus neoformans. Some interesting structure-activity relationships have also been discussed. Preliminary mechanism studies revealed that compound 23h might achieve its rapid fungicidal activity by disrupting the fungal cell membrane. Moreover, compound 23h exhibited significant inhibition against some virulence factors of Cryptococcus neoformans, low toxicity to normal human cells, as well as favorable pharmacokinetic and drug-like properties. The above results evidenced that the development of new antifungal candidates derived from natural phytoalexins was a bright and promising strategy.

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In recent years, petal blight disease caused by pathogens has become increasingly epidemic in Rhododendron. Breeding disease-resistant rhododendron is considered to be a more environmentally friendly strategy than is the use of chemical reagents. In this study, we aimed to investigate the response mechanisms of rhododendron varieties to petal blight, using transcriptomics and metabolomics analyses. Specifically, we monitored changes in gene expression and metabolite accumulation in Rhododendron 'Xiaotaohong' petals infected with the Alternaria sp. strain (MR-9). The infection of MR-9 led to the development of petal blight and induced significant changes in gene transcription. Differentially expressed genes (DEGs) were predominantly enriched in the plant-pathogen interaction pathway. These DEGs were involved in carrying out stress responses, with genes associated with H2O2 production being up-regulated during the early and late stages of infection. Correspondingly, H2O2 accumulation was detected in the vicinity of the blight lesions. In addition, defense-related genes, including PR and FRK, exhibited significant up-regulated expression during the infection by MR-9. In the late stage of the infection, we also observed significant changes in differentially abundant metabolites (DAMs), including flavonoids, alkaloids, phenols, and terpenes. Notably, the levels of euscaphic acid, ganoderol A, (-)-cinchonidine, and theophylline in infected petals were 21.8, 8.5, 4.5, and 4.3 times higher, respectively, compared to the control. Our results suggest that H2O2, defense-related genes, and DAM accumulation are involved in the complex response mechanisms of Rhododendron 'Xiaotaohong' petals to MR-9 infection. These insights provide a deeper understanding of the pathogenesis of petal blight disease and may have practical implications for developing disease-resistant rhododendron varieties.

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Crops experience herbivory by arthropods and microbial infections. In the interaction between plants and chewing herbivores, lepidopteran larval oral secretions (OS) and plant-derived damage-associated molecular patterns (DAMPs) trigger plant defense responses. However, the mechanisms underlying anti-herbivore defense, especially in monocots, have not been elucidated. The receptor-like cytoplasmic kinase Broad-Spectrum Resistance 1 (BSR1) of Oryza sativa L. (rice) mediates cytoplasmic defense signaling in response to microbial pathogens and enhances disease resistance when overexpressed. Here, we investigated whether BSR1 contributes to anti-herbivore defense responses. BSR1 knockout suppressed rice responses triggered by OS from the chewing herbivore Mythimna loreyi Duponchel (Lepidoptera: Noctuidae) and peptidic DAMPs OsPeps, including the activation of genes required for biosynthesis of diterpenoid phytoalexins (DPs). BSR1-overexpressing rice plants exhibited hyperactivation of DP accumulation and ethylene signaling after treatment with simulated herbivory and acquired enhanced resistance to larval feeding. As the biological significance of herbivory-induced accumulation of rice DPs remains unexplained, their physiological activities in M. loreyi were analyzed. The addition of momilactone B, a rice DP, to the artificial diet suppressed the growth of M. loreyi larvae. Altogether, this study revealed that BSR1 and herbivory-induced rice DPs are involved in the defense against chewing insects, in addition to pathogens.

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Phytoalexins are antimicrobial plant metabolites elicited by microbial attack or abiotic stress. We investigated phytoalexin profiles after foliar abiotic elicitation in the crucifer Barbarea vulgaris and interactions with the glucosinolate-myrosinase system. The treatment for abiotic elicitation was a foliar spray with CuCl2 solution, a usual eliciting agent, and three independent experiments were carried out. Two genotypes of B. vulgaris (G-type and P-type) accumulated the same three major phytoalexins in rosette leaves after treatment: phenyl-containing nasturlexin D and indole-containing cyclonasturlexin and cyclobrassinin. Phytoalexin levels were investigated daily by UHPLC-QToF MS and tended to differ among plant types and individual phytoalexins. In roots, phytoalexins were low or not detected. In treated leaves, typical total phytoalexin levels were in the range 1-10 nmol/g fresh wt. during three days after treatment while typical total glucosinolate (GSL) levels were three orders of magnitude higher. Levels of some minor GSLs responded to the treatment: phenethylGSL (PE) and 4-substituted indole GSLs. Levels of PE, a suggested nasturlexin D precursor, were lower in treated plants than controls. Another suggested precursor GSL, 3-hydroxyPE, was not detected, suggesting PE hydrolysis to be a key biosynthetic step. Levels of 4-substituted indole GSLs differed markedly between treated and control plants in most experiments, but not in a consistent way. The dominant GSLs, glucobarbarins, are not believed to be phytoalexin precursors. We observed statistically significant linear correlations between total major phytoalexins and the glucobarbarin products barbarin and resedine, suggesting that GSL turnover for phytoalexin biosynthesis was unspecific. In contrast, we did not find correlations between total major phytoalexins and raphanusamic acid or total glucobarbarins and barbarin. In conclusion, two groups of phytoalexins were detected in B. vulgaris, apparently derived from the GSLs PE and indol-3-ylmethylGSL. Phytoalexin biosynthesis was accompanied by depletion of the precursor PE and by turnover of major non-precursor GSLs to resedine. This work paves the way for identifying and characterizing genes and enzymes in the biosyntheses of phytoalexins and resedine.

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Phytoalexin is the main chemical weapon against pathogens in plants. Rice (Oryza sativa L.) produces a number of phytoalexins to defend against pathogens, most of which belong to the class of diterpenoid phytoalexins. Three biosynthetic gene clusters (BGCs) and a few non-BGC genes are responsible for rice diterpenoid phytoalexin biosynthesis. The corresponding regulatory mechanism of these phytoalexins in response to pathogen challenges still remains unclear. Here we identified a transcription factor, OsWRKY10, which positively regulates rice diterpenoid phytoalexin biosynthesis. Knockout mutants of OsWRKY10 obtained by CRISPR/Cas9 technology are more susceptible to Magnaporthe oryzae infection, while overexpression of OsWRKY10 enhances resistance to rice blast. Further analysis revealed that overexpression of OsWRKY10 increases accumulation of multiple rice diterpenoid phytoalexins and expression of genes in three BGCs and non-BGC genes in response to M. oryzae infection. Knockout of OsWRKY10 impairs upregulation of rice diterpenoid phytoalexin biosynthesis gene expression by blast pathogen and CuCl2 treatment. OsWRKY10 directly binds to the W-boxes or W-box-like elements (WLEs) of rice diterpenoid phytoalexin biosynthesis gene promoters to regulate gene expression. This study identified an extensive regulator (OsWRKY10) with broad transcriptional regulatory effects on rice diterpenoid phytoalexin biosynthesis genes, providing insight into the regulation of chemical defense to improve disease resistance in rice.

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WRKY transcription factors are involved in plant defense against pathogens. No WRKYs have been reported to be involved in resistance to tobacco brown spot disease caused by Alternaria alternata. Here, we found that NaWRKY3 plays a critical role in Nicotiana attenuata defense against A. alternata. NaWRKY3 bound and regulated many defense genes, including: lipoxygenase 3, ACC synthase 1, and ACC oxidase 1, three jasmonate- and ethylene-biosynthetic genes; feruloyl-CoA 6'-hydroxylase 1 (NaF6'H1), the biosynthetic gene for the phytoalexins scopoletin and scopolin; and three A. alternata resistance genes, the long non-coding RNA L2, NADPH oxidase (NaRboh D), and berberine bridge-like (NaBBL28). Silencing L2 reduced jasmonate concentrations and NaF6'H1 expression. NaRboh D-silenced plants were severely impaired in reactive oxygen species production and stomatal closure responses. NaBBL28 was the first A. alternata resistance BBL identified and was involved in the hydroxylation of 17-hydroxygeranyllinalool diterpene glycosides. NaWRKY3 bound to its own promoter but repressed its expression. Thus, we demonstrated that NaWRKY3 is a fine-tuned master regulator of the defense network against A. alternata in N. attenuata by regulating several signaling pathways and defense metabolites. This is the first time such an important WRKY has been identified in Nicotiana species, providing new insights into defense against A. alternata.

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Enhanced Disease Susceptibility 1 (EDS1), a key component of microbe-triggered immunity and effector-triggered immunity in most higher plants, forms functional heterodimeric complexes with its homologs Phytoalexin Deficient 4 (PAD4) or Senescence-associated Gene 101 (SAG101). Here, the crystal structure of VvEDS1Nterm , the N-terminal domain of EDS1 from Vitis vinifera, is reported, representing the first structure of an EDS1 entity beyond the model plant Arabidopsis thaliana. VvEDS1Nterm has an α/ß-hydrolase fold, is similar to the N-terminal domain of A. thaliana EDS1 and forms stable homodimers in solution as well as in crystals. These VvEDS1Nterm homodimers are spatially incompatible with heterodimers with PAD4 or SAG101, they explain why VvEDS1Nterm does not interact with V. vinifera PAD4 according to gel filtration, and they serve as a guide to develop a plausible, albeit experimentally not verified model of full-length EDS1. VvEDS1Nterm is a splicing variant comprising two of three exons of the VvEDS1 gene. It originates from a naturally occurring mRNA, in which the first of two introns was removed while the second one containing a stop codon close to the exon/intron border was retained. This is a potential case of intron retention and the first report of this phenomenon in the context of EDS1. Its biological significance has not yet been clarified, nor has the question if a VvEDS1Nterm protein with a specific function can occur under physiological conditions.

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Glycosyltransferases are nature's versatile tools to tailor the functionalities of proteins, carbohydrates, lipids, and small molecules by transferring sugars. Prominent substrates are hydroxycoumarins such as scopoletin, which serve as natural plant protection agents. Similarly, C13-apocarotenoids, which are oxidative degradation products of carotenoids/xanthophylls, protect plants by repelling pests and attracting pest predators. We show that C13-apocarotenoids interact with the plant glycosyltransferase NbUGT72AY1 and induce conformational changes in the enzyme catalytic center ultimately reducing its inherent UDP-α-d-glucose glucohydrolase activity and increasing its catalytic activity for productive hydroxycoumarin substrates. By contrast, C13-apocarotenoids show no effect on the catalytic activity toward monolignol lignin precursors, which are competitive substrates. In vivo studies in tobacco plants (Nicotiana benthamiana) confirmed increased glycosylation activity upon apocarotenoid supplementation. Thus, hydroxycoumarins and apocarotenoids represent specialized damage-associated molecular patterns, as they each provide precise information about the plant compartments damaged by pathogen attack. The molecular basis for the C13-apocarotenoid-mediated interplay of two plant protective mechanisms and their function as allosteric enhancers opens up potential applications of the natural products in agriculture and pharmaceutical industry.

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Pancreatic cancer (PC) is one of the deadliest malignancies, with an increasing incidence and limited response to current therapeutic options. Therefore, more effective and low-toxic agents are needed to improve PC patients' outcomes. Resveratrol (RSV) is a natural polyphenol with multiple biological properties, including anticancer effects. In this study, we explored the antiproliferative activities of newly synthetized RSV analogues in a panel of PC cell lines and evaluated the physicochemical properties of the most active compound. This derivative exhibited marked antiproliferative effects in PC cells through mechanisms involving DNA damage, apoptosis induction, and interference in cell cycle progression, as assessed using flow cytometry and immunoblot analysis of cell cycle proteins, PARP cleavage, and H2AX phosphorylation. Notably, the compound induced a consistent reduction in the PC cell subpopulation with a CD133+EpCAM+ stem-like phenotype, paralleled by dramatic effects on cell clonogenicity. Moreover, the RSV derivative had negligible toxicity against normal HFF-1 cells and, thus, good selectivity index values toward PC cell lines. Remarkably, its higher lipophilicity and stability in human plasma, as compared to RSV, might ensure a better permeation along the gastrointestinal tract. Our results provide insights into the mechanisms of action contributing to the antiproliferative activity of a synthetic RSV analogue, supporting its potential value in the search for effective and safe agents in PC treatment.

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