Understanding Burkholderia glumae BGR1 Virulence through the Application of Toxoflavin-Degrading Enzyme, TxeA.

Rice (Oryzae sativa cv. dongjin) is a cornerstone of global food security; however, Burkholderia glumae BGR1, which is responsible for bacterial panicle blight (BPB), threatens its productive output, with dire consequences for rice and other crops. BPB is primarily caused by toxoflavin, a potent phytotoxin that disrupts plant growth at various developmental stages. Therefore, understanding the mechanisms through which toxoflavin and BPB affect rice plants is critical. Toxoflavin biosynthesis in B. glumae BGR1 relies on the toxABCDE operon, with ToxA playing a central role. In response to this threat, our study explores a metagenome-derived toxoflavin-degrading enzyme, TxeA, as a potential defense mechanism against toxoflavin's destructive impact. TxeA-induced degradation of toxoflavin represents a potential strategy to mitigate crop damage. We introduce a groundbreaking approach: engineering transgenic rice plants to produce toxoflavin-degrading enzymes. These genetically modified plants, armed with TxeA, hold significant potential for combating toxoflavin-related crop losses. However, removal of toxoflavin, a major virulence factor in B. glumae BGR1, does not completely inhibit virulence. This innovative perspective offers a new shift from pathogen eradication to leveraging transgenic plants' power, offering a beacon of hope for crop protection and disease management. Our study offers insights into the intricate interplay between toxoflavin, BPB, and TxeA, providing a promising avenue to safeguard rice crops, ensure food security, and potentially enhance the resilience of various agricultural crops to B. glumae BGR1-induced diseases.

Salicylic Acid and Mobile Regulators of Systemic Immunity in Plants: Transport and Metabolism.

Systemic acquired resistance (SAR) occurs when primary infected leaves produce several SAR-inducing chemical or mobile signals that are transported to uninfected distal parts via apoplastic or symplastic compartments and activate systemic immunity. The transport route of many chemicals associated with SAR is unknown. Recently, it was demonstrated that pathogen-infected cells preferentially transport salicylic acid (SA) through the apoplasts to uninfected areas. The pH gradient and deprotonation of SA may lead to apoplastic accumulation of SA before it accumulates in the cytosol following pathogen infection. Additionally, SA mobility over a long distance is essential for SAR, and transpiration controls the partitioning of SA into apoplasts and cuticles. On the other hand, glycerol-3-phosphate (G3P) and azelaic acid (AzA) travel via the plasmodesmata (PD) channel in the symplastic route. In this review, we discuss the role of SA as a mobile signal and the regulation of SA transport in SAR.

Regulation of Salicylic Acid and N-Hydroxy-Pipecolic Acid in Systemic Acquired Resistance.

In plants, salicylic acid (SA) is a central immune signal that is involved in both local and systemic acquired resistance (SAR). In addition to SA, several other chemical signals are also involved in SAR and these include N-hydroxy-pipecolic acid (NHP), a newly discovered plant metabolite that plays a crucial role in SAR. Recent discoveries have led to a better understanding of the biosynthesis of SA and NHP and their signaling during plant defense responses. Here, I review the recent progress in role of SA and NHP in SAR. In addition, I discuss how these signals cooperate with other SAR-inducing chemicals to regulate SAR.

The Potential Effects of Light Irradiance in Glaucoma and Photobiomodulation Therapy.

Human vision is mediated by the retina, one of the most critical tissues in the central nervous system. Glaucoma is a complex retinal disease attributed to environmental, genetic, and stochastic factors, all of which contribute to its pathogenesis. Historically, glaucoma had been thought of primarily as a disease of the elderly; however, it is now becoming more problematic as the incidence rate increases among young individuals. In recent years, excessive light exposure has been suggested as contributing to the rise in glaucoma among the younger generation. Blue light induces mitochondrial apoptosis in retinal ganglion cells, causing optic damage; red light increases cytochrome c oxidase activity in the electron transport system, reducing inflammation and increasing antioxidant reactions to promote cell regeneration. In conclusion, the minimization of blue light exposure and the general application of red light treatment strategies are anticipated to show synergistic effects with existing treatments for retinal disease and glaucoma and should be considered a necessary prospect for the future. This review introduces the recent studies that support the relationship between light exposure and the onset of glaucoma and discusses new treatments, such as photobiomodulation therapy.

Biometabolites of Citrus unshiu Peel Enhance Intestinal Permeability and Alter Gut Commensal Bacteria.

Flavanones in Citrus unshiu peel (CUP) have been used as therapeutic agents to reduce intestinal inflammation; however, the anti-inflammatory effects of their biometabolites remain ambiguous. Here, we identified aglycone-type flavanones, such as hesperetin and naringenin, which were more abundant in the bioconversion of the CUP than in the ethanol extracts of the CUP. We found that the bioconversion of the CUP induced the canonical nuclear factor-κB pathway via degradation of IκB in Caco-2 cells. To check the immune suppressive capacity of the aglycones of the CUP in vivo, we orally administered the bioconversion of the CUP (500 mg/kg) to mice for two weeks prior to the 3% dextran sulfate sodium treatment. The CUP-pretreated group showed improved body weight loss, colon length shortage, and intestinal inflammation than the control mice. We also found a significant decrease in the population of lamina propria Th17 cells in the CUP-pretreated group following dextran sodium sulfate (DSS) treatment and an increase in mRNA levels of occludin in CUP-treated Caco-2 cells. Pyrosequencing analysis revealed a decreased abundance of Alistipes putredinis and an increased abundance of Muribaculum intestinale in the feces of the CUP-pretreated mice compared to those of the control mice. Overall, these findings suggest that the pre-administration of CUP biometabolites may inhibit the development of murine colitis by modulating intestinal permeability and the gut microbiome.

Isolation and Functional Characterization of Soybean BES1/BZR1 Homolog 3-Like 1 (GmBEH3L1) Associated with Dehydration Sensitivity and Brassinosteroid Signaling in Arabidopsis thaliana

Brassinosteroid (BR) is an important steroid hormone that regulates plant development, abscisic acid (ABA) signaling, and responses to abiotic stress. We previously demonstrated that BEH3 (BES1/BZR1 Homolog 3) of Arabidopsis thaliana regulates dehydration and ABA responses by mediating proline metabolism. Furthermore, BEH3 negatively regulates BR-mediated hypocotyl elongation in dark-grown seedlings. However, the roles of BEH3 ortholog genes in the osmotic stress response of plants have remained largely unknown. Here, GmBEH3L1 (Glycine max BEH3-Like 1), a soybean (G. max) ortholog of the BEH3 gene of A. thaliana, was isolated and functionally characterized. GmBEH3L1 is induced by ABA, dehydration, and drought conditions. The GmBEH3L1-overexpressing transgenic lines (GmBEH3L1-OE/beh3) with the beh3 mutant background have ABA- and dehydration-sensitive phenotypes during early seedling growth, implying that GmBEH3L1 is involved in both osmotic stress and ABA sensitivity as a negative regulator in A. thaliana. Consistent with these results, GmBEH3L1-OE/beh3 complemental lines exhibit decreased expression levels of ABA- or dehydration-inducible genes. Under darkness, GmBEH3L1-OE/beh3 complemental lines display a short hypocotyl length compared to the beh3 mutant, indicating that GmBEH3L1 is linked to BR signaling. Together, our data suggest that GmBEH3L1 participates negatively in ABA and dehydration responses through BR signaling.

Förster Resonance Energy Transfer-Based Single-Cell Imaging Reveals Piezo1-Induced Ca2+ Flux Mediates Membrane Ruffling and Cell Survival.

A mechanosensitive ion channel, Piezo1 induces non-selective cation flux in response to various mechanical stresses. However, the biological interpretation and underlying mechanisms of cells resulting from Piezo1 activation remain elusive. This study elucidates Piezo1-mediated Ca2+ influx driven by channel activation and cellular behavior using novel Förster Resonance Energy Transfer (FRET)-based biosensors and single-cell imaging analysis. Results reveal that extracellular Ca2+ influx via Piezo1 requires intact caveolin, cholesterol, and cytoskeletal support. Increased cytoplasmic Ca2+ levels enhance PKA, ERK, Rac1, and ROCK activity, which have the potential to promote cancer cell survival and migration. Furthermore, we demonstrate that Piezo1-mediated Ca2+ influx upregulates membrane ruffling, a characteristic feature of cancer cell metastasis, using spatiotemporal image correlation spectroscopy. Thus, our findings provide new insights into the function of Piezo1, suggesting that Piezo1 plays a significant role in the behavior of cancer cells.

Phased small RNA-mediated systemic signaling in plants.

Systemic acquired resistance (SAR) involves the generation of systemically transported signal that arms distal plant parts against secondary infections. We show that two phased 21-nucleotide (nt) trans-acting small interfering RNA3a RNAs (tasi-RNA) derived from TAS3a and synthesized within 3 hours of pathogen infection are the early mobile signal in SAR. TAS3a undergoes alternate polyadenylation, resulting in the generation of 555- and 367-nt transcripts. The 555-nt transcripts likely serves as the sole precursor for tasi-RNAs D7 and D8, which cleave Auxin response factors (ARF) 2, 3, and 4 to induce SAR. Conversely, increased expression of ARF3 represses SAR. Knockout mutations in TAS3a or RNA silencing components required for tasi-RNA biogenesis compromise SAR without altering levels of known SAR-inducing chemicals. Both tasi-ARFs and the 367-nt transcripts are mobile and transported via plasmodesmata. Together, we show that tasi-ARFs are the early mobile signal in SAR.

Upregulation of the MYB2 Transcription Factor Is Associated with Increased Accumulation of Anthocyanin in the Leaves of Dendrobium bigibbum.

Orchids with colorful leaves and flowers have significant ornamental value. Here, we used γ-irradiation-based mutagenesis to produce a Dendrobium bigibbum mutant that developed purple instead of the normal green leaves. RNA sequencing of the mutant plant identified 2513 differentially expressed genes, including 1870 up- and 706 downregulated genes. The purple leaf color of mutant leaves was associated with increased expression of genes that encoded key biosynthetic enzymes in the anthocyanin biosynthetic pathway. In addition, the mutant leaves also showed increased expression of several families of transcription factors including the MYB2 gene. Transient overexpression of D. biggibumMYB2 in Nicotiana benthamiana was associated with increased expression of endogenous anthocyanin biosynthesis genes. Interestingly, transient overexpression of orthologous MYB2 genes from other orchids did not upregulate expression of endogenous anthocyanin biosynthesis genes. Together, these results suggest that the purple coloration of D. biggibum leaves is at least associated with increased expression of the MYB2 gene, and the MYB2 orthologs from orchids likely function differently, regardless of their high level of similarity.

Dramatic Increase in Content of Diverse Flavonoids Accompanied with Down-Regulation of F-Box Genes in a Chrysanthemum (Chrysanthemum × morifolium (Ramat.) Hemsl.) Mutant Cultivar Producing Dark-Purple Ray Florets.

Anthocyanins (a subclass of flavonoids) and flavonoids are crucial determinants of flower color and substances of pharmacological efficacy, respectively, in chrysanthemum. However, metabolic and transcriptomic profiling regarding flavonoid accumulation has not been performed simultaneously, thus the understanding of mechanisms gained has been limited. We performed HPLC-DAD-ESI-MS (high-performance liquid chromatography coupled with photodiode array detection and electrospray ionization mass spectrometry) and transcriptome analyses using "ARTI-Dark Chocolate" (AD), which is a chrysanthemum mutant cultivar producing dark-purple ray florets, and the parental cultivar "Noble Wine" for metabolic characterization and elucidation of the genetic mechanism determining flavonoid content. Among 26 phenolic compounds identified, three cyanidins and eight other flavonoids were detected only in AD. The total amounts of diverse flavonoids were 8.0 to 10.3 times higher in AD. Transcriptome analysis showed that genes in the flavonoid biosynthetic pathway were not up-regulated in AD at the early flower stage, implying that the transcriptional regulation of the pathway did not cause flavonoid accumulation. However, genes encoding post-translational regulation-related proteins, especially F-box genes in the mutated gene, were enriched among down-regulated genes in AD. From the combination of metabolic and transcriptomic data, we suggest that the suppression of post-translational regulation is a possible mechanism for flavonoid accumulation in AD. These results will contribute to research on the regulation and manipulation of flavonoid biosynthesis in chrysanthemum.

The plant cuticle regulates apoplastic transport of salicylic acid during systemic acquired resistance.

The plant cuticle is often considered a passive barrier from the environment. We show that the cuticle regulates active transport of the defense hormone salicylic acid (SA). SA, an important regulator of systemic acquired resistance (SAR), is preferentially transported from pathogen-infected to uninfected parts via the apoplast. Apoplastic accumulation of SA, which precedes its accumulation in the cytosol, is driven by the pH gradient and deprotonation of SA. In cuticle-defective mutants, increased transpiration and reduced water potential preferentially routes SA to cuticle wax rather than to the apoplast. This results in defective long-distance transport of SA, which in turn impairs distal accumulation of the SAR-inducer pipecolic acid. High humidity reduces transpiration to restore systemic SA transport and, thereby, SAR in cuticle-defective mutants. Together, our results demonstrate that long-distance mobility of SA is essential for SAR and that partitioning of SA between the symplast and cuticle is regulated by transpiration.

Transcriptome analysis to identify candidate genes associated with the yellow-leaf phenotype of a Cymbidium mutant generated by γ-irradiation.

Leaf color is an important agronomic trait in flowering plants, including orchids. However, factors underlying leaf phenotypes in plants remain largely unclear. A mutant displaying yellow leaves was obtained by the γ-ray-based mutagenesis of a Cymbidium orchid and characterized using RNA sequencing. A total of 144,918 unigenes obtained from over 25 million reads were assigned to 22 metabolic pathways in the Kyoto Encyclopedia of Genes and Genomes database. In addition, gene ontology was used to classify the predicted functions of transcripts into 73 functional groups. The RNA sequencing analysis identified 2,267 differentially expressed genes between wild-type and mutant Cymbidium sp. Genes involved in the chlorophyll biosynthesis and degradation, as well as ion transport, were identified and assayed for their expression levels in wild-type and mutant plants using quantitative real-time profiling. No critical expression changes were detected in genes involved in chlorophyll biosynthesis. In contrast, seven genes involved in ion transport, including two metal ion transporters, were down-regulated, and chlorophyllase 2, associated with chlorophyll degradation, was up-regulated. Together, these results suggest that alterations in chlorophyll metabolism and/or ion transport might contribute to leaf color in Cymbidium orchids.

The analogous and opposing roles of double-stranded RNA-binding proteins in bacterial resistance.

The Arabidopsis plasma membrane-localized resistance protein RPM1 is degraded upon the induction of the hypersensitive response (HR) triggered in response to its own activation or that of other unrelated resistance (R) proteins. We investigated the role of RPM1 turnover in RPM1-mediated resistance and showed that degradation of RPM1 is not associated with HR or resistance mediated by this R protein. Likewise, the runaway cell death phenotype in the lsd1 mutant was not associated with RPM1 degradation and did not alter RPM1-derived resistance. RPM1 stability and RPM1-mediated resistance were dependent on the double-stranded RNA binding (DRB) proteins 1 and 4. Interestingly, the function of DRB1 in RPM1-mediated resistance was not associated with its role in pre-miRNA processing. The DRB3 and DRB5 proteins negatively regulated RPM1-mediated resistance and a mutation in these completely or partially restored resistance in the drb1, drb2, and drb4 mutant backgrounds. Conversely, plants overexpressing DRB5 showed attenuated RPM1-mediated resistance. A similar role for DRBs in basal and R-mediated resistance suggests that these proteins play a general role in bacterial resistance.

Pipecolic acid confers systemic immunity by regulating free radicals.

Pipecolic acid (Pip), a non-proteinaceous product of lysine catabolism, is an important regulator of immunity in plants and humans alike. In plants, Pip accumulates upon pathogen infection and has been associated with systemic acquired resistance (SAR). However, the molecular mechanisms underlying Pip-mediated signaling and its relationship to other known SAR inducers remain unknown. We show that in plants, Pip confers SAR by increasing levels of the free radicals, nitric oxide (NO), and reactive oxygen species (ROS), which act upstream of glycerol-3-phosphate (G3P). Plants defective in NO, ROS, G3P, or salicylic acid (SA) biosynthesis accumulate reduced Pip in their distal uninfected tissues although they contain wild-type-like levels of Pip in their infected leaves. These data indicate that de novo synthesis of Pip in distal tissues is dependent on both SA and G3P and that distal levels of SA and G3P play an important role in SAR. These results also suggest a unique scenario whereby metabolites in a signaling cascade can stimulate each other's biosynthesis depending on their relative levels and their site of action.

COP1, a negative regulator of photomorphogenesis, positively regulates plant disease resistance via double-stranded RNA binding proteins.

The E3 ubiquitin ligase COP1 (Constitutive Photomorphogenesis 1) is a well known component of the light-mediated plant development that acts as a repressor of photomorphogenesis. Here we show that COP1 positively regulates defense against turnip crinkle virus (TCV) and avrRPM1 bacteria by contributing to stability of resistance (R) protein HRT and RPM1, respectively. HRT and RPM1 levels and thereby pathogen resistance is significantly reduced in the cop1 mutant background. Notably, the levels of at least two double-stranded RNA binding (DRB) proteins DRB1 and DRB4 are reduced in the cop1 mutant background suggesting that COP1 affects HRT stability via its effect on the DRB proteins. Indeed, a mutation in either drb1 or drb4 resulted in degradation of HRT. In contrast to COP1, a multi-subunit E3 ligase encoded by anaphase-promoting complex (APC) 10 negatively regulates DRB4 and TCV resistance but had no effect on DRB1 levels. We propose that COP1-mediated positive regulation of HRT is dependent on a balance between COP1 and negative regulators that target DRB1 and DRB4.

Fatty Acid- and Lipid-Mediated Signaling in Plant Defense.

Fatty acids and lipids, which are major and essential constituents of all plant cells, not only provide structural integrity and energy for various metabolic processes but can also function as signal transduction mediators. Lipids and fatty acids can act as both intracellular and extracellular signals. In addition, cyclic and acyclic products generated during fatty acid metabolism can also function as important chemical signals. This review summarizes the biosynthesis of fatty acids and lipids and their involvement in pathogen defense.

Transport of chemical signals in systemic acquired resistance.

Systemic acquired resistance (SAR) is a form of broad-spectrum resistance induced in response to local infections that protects uninfected parts against subsequent secondary infections by related or unrelated pathogens. SAR signaling requires two parallel branches, one regulated by salicylic acid (SA), and the other by azelaic acid (AzA) and glycerol-3-phosphate (G3P). AzA and G3P function downstream of the free radicals nitric oxide (NO) and reactive oxygen species (ROS). During SAR, SA, AzA and G3P accumulate in the infected leaves, but only a small portion of these is transported to distal uninfected leaves. SA is preferentially transported via the apoplast, whereas phloem loading of AzA and G3P occurs via the symplast. The symplastic transport of AzA and G3P is regulated by gating of the plasmodesmata (PD). The PD localizing proteins, PDLP1 and PDLP5, regulate SAR by regulating PD gating as well as the subcellular partitioning of a SAR-associated protein.

Role of plasmodesmata and plasmodesmata localizing proteins in systemic immunity.

Systemic acquired resistance (SAR) is a highly desirable form of resistance that protects against a broad-spectrum of pathogens. SAR involves the generation of a mobile signal at the site of primary infection, which arms distal portions of a plant against subsequent secondary infections. A number of diverse chemical signals contributing to SAR have been isolated and characterized. Among these, salicylic acid (SA) functions in parallel to azelaic acid (AzA) and glycerol-3-phosphate (G3P), and both AzA and G3P function downstream of the free radicals nitric oxide and reactive oxygen species. We now show that phloem loading of AzA and G3P occurs via the symplast, whereas that of SA occurs via the apoplast. The symplastic transport of AzA and G3P is regulated by plasmodesmata localizing protein (PDLP) 5, which together with PDLP1 also plays a signaling role in SAR. Together, these results reveal the transport routes of SAR associated chemical signals, and the regulatory role of PDLPs in SAR.

Plasmodesmata Localizing Proteins Regulate Transport and Signaling during Systemic Acquired Immunity in Plants.

Systemic acquired resistance (SAR) in plants is mediated by the signaling molecules azelaic acid (AzA), glycerol-3-phosphate (G3P), and salicylic acid (SA). Here, we show that AzA and G3P transport occurs via the symplastic route, which is regulated by channels known as plasmodesmata (PD). In contrast, SA moves via the extracytosolic apoplast compartment. We found that PD localizing proteins (PDLP) 1 and 5 were required for SAR even though PD permeability in pdlp1 and 5 mutants was comparable to or higher than wild-type plants, respectively. Furthermore, PDLP function was required in the recipient cell, suggesting regulatory function in SAR. Interestingly, overexpression of PDLP5 drastically reduced PD permeability, yet also impaired SAR. PDLP1 interacted with AZI1 (lipid transfer-like protein required for AzA- and G3P-induced SAR) and contributed to its intracellular partitioning. Together, these results reveal the transport routes of SAR chemical signals and highlight the regulatory role of PD-localizing proteins in SAR.

RNA silencing components mediate resistance signaling against turnip crinkle virus.

Species-specific immunity is induced when an effector protein from a specific pathogen strain is perceived by a cognate resistance (R) protein in the plant. In Arabidopsis, the R protein HRT, which confers resistance to turnip crinkle virus (TCV), is activated upon recognition of the TCV coat-protein (CP), a potent suppressor of host RNA silencing. Recognition by HRT does not require RNA silencing suppressor function of CP and is not associated with the accumulation of TCV-specific small-RNA. However, several components of the host RNA silencing pathway participate in HRT-mediated defense against TCV. For example, the double stranded RNA binding protein (DRB) 4 interacts with the plasma membrane localized HRT, and is required for its stability. Intriguingly, TCV infection promotes the cytosolic accumulation of the otherwise primarily nuclear DRB4, and this in turn inhibits HRT-DRB4 interaction. These data together with differential localization of DRB4 in plants inoculated with avirulent and virulent viruses, suggests that sub-cellular compartmentalization of DRB4 plays an important role in activation of HRT.