The Mh-miR393a-TIR1 module regulates Alternaria alternata resistance of Malus hupehensis mainly by modulating the auxin signaling.

miRNAs govern gene expression and regulate plant defense. Alternaria alternata is a destructive fungal pathogen that damages apple. The wild apple germplasm Malus hupehensis is highly resistant to leaf spot disease caused by this fungus. Herein, we elucidated the regulatory and functional role of miR393a in apple resistance against A. alternata by targeting Transport Inhibitor Response 1. Mature miR393 accumulation in infected M. hupehensis increased owing to the transcriptional activation of MIR393a, determined to be a positive regulator of A. alternata resistance to either 'Orin' calli or 'Gala' leaves. 5' RLM-RACE and co-transformation assays showed that the target of miR393a was MhTIR1, a gene encoding a putative F-box auxin receptor that compromised apple immunity. RNA-seq analysis of transgenic calli revealed that MhTIR1 upregulated auxin signaling gene transcript levels and influenced phytohormone pathways and plant-pathogen interactions. miR393a compromised the sensitivity of several auxin-signaling genes to A. alternata infection, whereas MhTIR1 had the opposite effect. Using exogenous indole-3-acetic acid or the auxin synthesis inhibitor L-AOPP, we clarified that auxin enhances apple susceptibility to this pathogen. miR393a promotes SA biosynthesis and impedes pathogen-triggered ROS bursts by repressing TIR1-mediated auxin signaling. We uncovered the mechanism underlying the miR393a-TIR1 module, which interferes with apple defense against A. alternata by modulating the auxin signaling pathway.

Multifunctionality of AsCFEM6 and AsCFEM12 effectors from the potato early blight pathogen Alternaria solani.

Plant pathogens secrete fungal-specific common in several fungal extracellular membrane (CFEM) effectors to manipulate host immunity and contribute to their virulence. Little is known about effectors and their functions in Alternaria solani, the necrotrophic fungal pathogen causing potato early blight. To identify candidate CFEM effector genes, we mined A. solani genome databases. This led to the identification of 12 genes encoding CFEM proteins (termed AsCFEM1-AsCFEM12) and 6 of them were confirmed to be putative secreted effectors. In planta expression revealed that AsCFEM6 and AsCFEM12 have elicitor function that triggers plant defense response including cell death in different botanical families. Targeted gene disruption of AsCFEM6 and AsCFEM12 resulted in a change in spore development, significant reduction of virulence on potato and eggplant susceptible cultivars, increased resistance to fungicide stress, variation in iron acquisition and utilization, and the involvement in 1,8-dihydroxynaphthalene (DHN) melanin biosynthesis pathway. Using maximum likelihood method, we found that positive selection likely caused the polymorphism within AsCFEM6 and AsCFEM12 homologs in different Alternaria spp. Site-directed mutagenesis analysis indicated that positive selection sites within their CFEM domains are required for cell death induction in Nicotiana benthamiana and are critical for response to abiotic stress in yeast. These results demonstrate that AsCFEM effectors possess additional functions beyond their roles in host plant immune response and pathogen virulence.

miR164-NAC21/22 module regulates the resistance of Malus hupehensis against Alternaria alternata by controlling jasmonic acid signaling.

Apple leaf spot disease caused by Alternaria alternata apple pathotype (A. alternata AP) is one of the most severe fungal diseases affecting apple cultivation. Transcription factors are involved in various disease-resistance responses, and many of them are regulated by miRNAs. Here, we performed RNA-Seq to investigate gene expression changes during the defense response of Malus hupehensis against A. alternata AP. NAC21/22 was induced upon A. alternata AP infection and silenced by miR164 via direct mRNA cleavage. Contrasting expression patterns were noted between mature miR164 and NAC21/22 during infection. Contrary to NAC21/22 silencing, transiently overexpressing NAC21/22 in M. hupehensis alleviated disease symptoms on 'gala' leaves, impeded A. alternata AP growth, and promoted jasmonic acid (JA) signaling-related gene expression. Importantly, transient miR164f overexpression in 'gala' leaves enhanced A. alternata AP sensitivity, due perhaps to NAC21/22 downregulation, whereas miR164 suppression produced an opposite effect. In summary, the miR164-NAC21/22 module plays a pivotal role in apple resistance against A. alternata AP by regulating JA signaling.

Acibenzolar-S-methyl activates phenylpropanoid pathway to enhance resistance against Alternaria alternata in pear fruit.

BACKGROUND: Alternaria alternata is a causal agent of black spot rot of pear fruit after harvest. Acibenzolar-S-methyl (ASM) has been shown to be a potential elicitor of tolerance in several horticultural products. This work was performed to research the influence of ASM on black spot rot of Docteur Jules Guyot pears and vital enzyme activity and gene expression in the phenylpropanoid pathway. RESULTS: ASM remarkably decreased the lesion diameter of A. alternata-inoculated pears. ASM also increased phenylalanine ammonialyase, cinnamate 4-hydroxylase, cinnamyl alcohol dehydrogenase, peroxidase, polyphenol oxidase activities and gene expression, and enhanced 4-coumarate/coenzyme A ligase activity in pears. Moreover, ASM improved the content of phenylalanine, total phenolic compounds, caffeic acid, flavonoids, anthocyanin and lignin in pears. CONCLUSION: ASM could modulate vital enzyme activity and gene expression in the phenylpropanoid pathway to accelerate metabolite synthesis, thereby enhancing resistance against A. alternata in pears. © 2022 Society of Chemical Industry.

Comparative Analysis of Alternative Splicing in Two Contrasting Apple Cultivars Defense against Alternaria alternata Apple Pathotype Infection.

Alternaria blotch disease, caused by the Alternaria alternata apple pathotype (A. alternata AP), is one of the most serious fungal diseases in apples. Alternative splicing (AS), one of the pivotal post-transcriptional regulatory mechanisms, plays essential roles in various disease resistance responses. Here, we performed RNA-Seq for two apple cultivars (resistant cultivar 'Jonathan' (J) and susceptible cultivar 'Starking Delicious' (SD)) infected by A. alternata AP to further investigate their AS divergence. In total, 1454, 1780, 1367 and 1698 specifically regulated differential alternative splicing (DAS) events were detected in J36, J72, SD36 and SD72 groups, respectively. Retained intron (RI) was the dominant AS pattern. Conformably, 642, 764, 585 and 742 uniquely regulated differentially spliced genes (DSGs) were found during A. alternata AP infection. Comparative analysis of AS genes in differential splicing and expression levels suggested that only a small proportion of DSGs overlapped with differentially expressed genes (DEGs). Gene ontology (GO) enrichment analysis demonstrated that the DSGs were significantly enriched at multiple levels of gene expression regulation. Briefly, the specific AS was triggered in apple defense against A. alternata AP. Therefore, this study facilitates our understanding on the roles of AS regulation in response to A. alternata AP infection in apples.

Identification of effector CEP112 that promotes the infection of necrotrophic Alternaria solani.

BACKGROUND: Alternaria solani is a typical necrotrophic pathogen that can cause severe early blight on Solanaceae crops and cause ring disease on plant leaves. Phytopathogens produce secretory effectors that regulate the host immune response and promote pathogenic infection. Effector proteins, as specialized secretions of host-infecting pathogens, play important roles in disrupting host defense systems. At present, the role of the effector secreted by A. solani during infection remains unclear. We report the identification and characterization of AsCEP112, an effector required for A. solani virulence. RESULT: The AsCEP112 gene was screened from the transcriptome and genome of A. solani on the basis of typical effector signatures. Fluorescence quantification and transient expression analysis showed that the expression level of AsCEP112 continued to increase during infection. The protein localized to the cell membrane of Nicotiana benthamiana and regulated senescence-related genes, resulting in the chlorosis of N. benthamiana and tomato leaves. Moreover, comparative analysis of AsCEP112 mutant obtained by homologous recombination with wild-type and revertant strains indicated that AsCEP112 gene played an active role in regulating melanin formation and penetration in the pathogen. Deletion of AsCEP112 also reduced the pathogenicity of HWC-168. CONCLUSION: Our findings demonstrate that AsCEP112 was an important effector protein that targeted host cell membranes. AsCEP112 regulateed host senescence-related genes to control host leaf senescence and chlorosis, and contribute to pathogen virulence.

Integrated transcriptomic and transgenic analyses reveal potential mechanisms of poplar resistance to Alternaria alternata infection.

BACKGROUND: Populus davidiana × P. bollena is a species of poplar from northeastern China that is characterized by cold resistance and fast growth but now suffers from pathogen infections. Leaf blight caused by Alternaria alternata has become a common poplar disease that causes serious economic impacts, but the molecular mechanisms of resistance to A. alternata in P. davidiana × P. bollena are still unclear. RESULTS: In this study, the transcriptomic response of P. davidiana × P. bollena to A. alternata infection was determined via RNA-Seq. Twelve cDNA libraries were generated from RNA isolated from three biological replicates at four time points (0, 2, 3, and 4 d post inoculation), and a total of 5,930 differentially expressed genes (DEGs) were detected (| log2 fold change |≥ 1 and FDR values < 0.05). Functional analysis revealed that the DEGs were mainly enriched for the "plant hormone signal transduction" pathway, followed by the "phenylpropanoid biosynthesis" pathway. In addition, DEGs that encode defense-related proteins and are related to ROS metabolism were also identified. Numerous transcription factors, such as the bHLH, WRKY and MYB families, were also induced by A. alternata infection. Among these DEGs, those related to JA biosynthesis and JA signal transduction were consistently activated. Therefore, the lipoxygenase gene PdbLOX2, which is involved in JA biosynthesis, was selected for functional characterization. Overexpression of PdbLOX2 enhanced the resistance of P. davidiana × P. bollena to A. alternata, whereas silencing this gene enhanced susceptibility to A. alternata infection. CONCLUSIONS: These results provide new insight into the molecular mechanisms of poplar resistance to A. alternata infection and provide candidate genes for breeding resistant cultivars using genetic engineering.

The regulation of Alternaria alternata resistance by LRR-RK4 through ERF109, defensin19 and phytoalexin scopoletin in Nicotiana attenuata.

Leucine-rich repeat receptor-like kinases (LRR-RKs), belonging to the largest subfamily of transmembrane receptor-like kinases in plants, are proposed to be involved in pathogen resistance. However, it is currently unknown whether LRR-RKs regulate Nicotiana attenuata resistance to Alternaria alternata, a notorious fungal pathogen causing tobacco brown disease. During transcriptome analysis, we identified a highly induced receptor kinase (NaLRR-RK4) in N. attenuata leaves after A. alternata inoculation. We speculated that this NaLRR-RK4 might be the resistance gene of tobacco to brown spot disease, and if so, what is its function and mechanism of action? Silencing of NaLRR-RK4 via virus-induced gene silencing (VIGS) lead to plants highly susceptible to A. alternata, and this result was further confirmed by two stable transformation lines (NaLRR-RK4-RNAi lines) generated by RNA interference technology. The susceptible of NaLRR-RK4-RNAi lines to A. alternata was associated with reduced levels of phytoalexin scopoletin and its key synthesis gene NaF6'H1. Further transcriptome analysis of leaves of WT and NaLRR-RK4-RNAi line after A. alternata inoculation revealed that NaLRR-RK4 regulated NaERF109 and NaDEF19. Silencing NaERF109 or NaDEF19 by VIGS lead to plants more susceptible to A.alternata, demonstrating their role in pathogen resistance. Interestingly, A.alternata-induced expression of NaF6'H1 and NaDEF19 were dramatically reduced in NaERF109-silenced VIGS plants. Taken all together, we identified LRR-RK4 as the first Leucine-rich repeat receptor-like kinases involved in A.alternata resistance in tobacco species, by regulating NaERF109, and subsequently NaDEF19 and NaF6'H1.

Ozone reduces the fruit decay of postharvest winter jujube by altering the microbial community structure on fruit surface.

Microbial community structure on fruit surface plays an important role in fruit decay during postharvest storage, although the underlying mechanism has not been fully elucidated. Winter jujube (Ziziphus jujuba Miller cv. Dongzao) is a unique fruit resource with high edible and commercial value in China, while postharvest decay has always been a severe problem leading to short shelf life and poor quality of fruit. Ozone treatment is regarded as one of the most effective means to control decay and extend shelf life because of its cost-effective and eco-friendly properties. In the present study, three concentrations of ozone (2.5, 5 and 10 µL L-1) were found to reduce significantly postharvest decay of winter jujube on days 10 and 15, which were produced from Huanghua City, Hebei, China. High-throughput sequencing revealed significant changes in the bacterial and fungal communities in response to the application of ozone treatment, while Didymella, Rhizopus, Alternaria, Phialemoniopsis and Mycosphaerella were found to be the most abundant in fungi, and Methylobacterium, Pseudomonas, Pantoea, Sphingomonas and Gluconobacter being the most abundant in bacteria. Results of linear discriminant analysis (LDA) effect size (LEfSe) indicated that ozone treatments considerably reduced the abundance of Rhizopus and Gluconobacter on the surface of winter jujube fruit. Furthermore, Pearson correlation analysis showed that Rhizopus was positively correlated with Gluconobacter (r = 0.97) while negatively correlated with Didymella (r = -0.96). By predicting the metabolic function, ozone may inhibit metabolic pathways including nucleoside and nucleotide biosynthesis, amino acid biosynthesis, fatty acid and lipid degradation, respiration, and electron transfer, thereby reducing the incidence of fruit decay and maintaining the firmness of winter jujube fruit.

Utilization of the antagonistic yeast, Wickerhamomyces anomalus, combined with UV-C to manage postharvest rot of potato tubers caused by Alternaria tenuissima.

Postharvest rot of potato tubers caused by fungal pathogens is the main cause of significant economic losses, while also raising potential food safety issues. Integrated disease management, utilizing bio-safe and eco-friendly methods, represents a sustainable strategy for reducing postharvest losses in crops, including potato. In the current study, the application of the antagonistic yeast, Wickerhamomyces anomalus, combined with a UV-C treatment was evaluated for the management of postharvest Alternaria rot of potato tubers, caused by Alternaria tenuissima. Both W. anomalus and UV-C as individual treatments reduced the size of A. tenuissima infections on potato tubers, relative to the control, while the combined treatment of W. anomalus and UV-C exhibited the highest level of inhibition. W. anomalus or UV-C alone, and especially when used in combination, induced the expression of defense-related genes, including polyphenol oxidase, peroxidase, and ß-1,3-glucanase, and also increased the level of flavonoids and lignin in potato tubers. Our findings indicate that the mechanism of action by which UV-C enhances the biocontrol effect of W. anomalus against postharvest Alternaria rot includes the activation of defense-related response in potato tubers. The integration of biocontrol agents and physical treatments (e.g., UV-C) represents an effective, eco-friendly hurdle technology for managing postharvest rot in potato.

The elemental defense effect of cadmium on Alternaria brassicicola in Brassica juncea.

BACKGROUND: The elemental defense hypothesis states a new defensive strategy that hyperaccumulators defense against herbivores or pathogens attacks by accumulating heavy metals. Brassica juncea has an excellent ability of cadmium (Cd) accumulation. However, the elemental defense effect and its regulation mechanism in B. juncea remain unclear. RESULTS: In this study, we profiled the elemental defense effect and the molecular regulatory mechanism in Cd-accumulated B. juncea after Alternaria brassicicola infection. B. juncea treated with 180 mg Kg- 1 DW CdCl2 2.5H2O exhibited obvious elemental defense effect after 72 h of infection with A. brassicicola. The expression of some defense-related genes including BjNPR1, BjPR12, BjPR2, and stress-related miRNAs (miR156, miR397, miR398a, miR398b/c, miR408, miR395a, miR395b, miR396a, and miR396b) were remarkably elevated during elemental defense in B. juncea. CONCLUSIONS: The results indicate that Cd-accumulated B. juncea may defend against pathogens by coordinating salicylic acid (SA) and jasmonic acid (JA) mediated systemic acquired resistance (SAR) and elemental defense in a synergistic joint effect. Furthermore, the expression of miRNAs related to heavy metal stress response and disease resistance may regulate the balance between pathogen defense and heavy metal stress-responsive in B. juncea. The findings provide experimental evidence for the elemental defense hypothesis in plants from the perspectives of phytohormones, defense-related genes, and miRNAs.

Alternaria alternata-induced airway epithelial signaling and inflammatory responses via protease-activated receptor-2 expression.

Inhalation of the fungus Alternaria alternata is associated with an increased risk of allergic asthma development and exacerbations. Recent work in acute exposure animal models suggests that A. alternata-induced asthma symptoms, which include inflammation, mucus overproduction and airway hyperresponsiveness, are due to A. alternata proteases that act via protease-activated receptor-2 (PAR2). However, because other active components present in A. alternata may be contributing to asthma pathophysiology through alternative signaling, the specific role PAR2 plays in asthma initiation and maintenance remains undefined. Airway epithelial cells provide the first encounter with A. alternata and are thought to play an important role in initiating the physiologic response. To better understand the role for PAR2 airway epithelial signaling we created a PAR2-deficient human bronchial epithelial cell line (16HBEPAR-/-) from a model bronchial parental line (16HBE14o-). Comparison of in vitro physiologic responses in these cell lines demonstrated a complete loss of PAR2 agonist (2at-LIGRL-NH2) response and significantly attenuated protease (trypsin and elastase) and A. alternata responses in the 16HBEPAR-/- line. Apical application of A. alternata to 16HBE14o- and 16HBEPAR2-/- grown at air-liquid interface demonstrated rapid, PAR2-dependent and independent, inflammatory cytokine, chemokine and growth factor basolateral release. In conclusion, the novel human PAR2-deficient cell line allows for direct in vitro examination of the role(s) for PAR2 in allergen challenge with polarized human airway epithelial cells.

Distinct and essential roles of bZIP transcription factors in the stress response and pathogenesis in Alternaria alternata.

The ability to cope with environmental abiotic stress and biotic stress is crucial for the survival of plants and microorganisms, which enable them to occupy multiple niches in the environment. Previous studies have shown that transcription factors play crucial roles in regulating various biological processes including multiple stress tolerance and response in eukaryotes. This work identified multiple critical transcription factor genes, metabolic pathways and gene ontology (GO) terms related to abiotic stress response were broadly activated by analyzing the transcriptome of phytopathogenic fungus Alternaria alternata under metal ions stresses, oxidative stress, salt stresses, and host-pathogen interaction. We investigated the biological functions and regulatory roles of the bZIP transcriptional factor (TF) genes in the phytopathogenic fungus A.alternata by analyzing targeted gene disrupted mutants. Morphological analysis provides evidence that the bZIP transcription factors (Gcn4, MeaB, Atf1, the ER stress regulator Hac1, and the all development altered-1 gene Ada1) are required for morphogenesis as the colony morphology of these gene deletion mutants was significantly different from that of the wild-type. In addition, bZIPs are involved in the resistance to multiple stresses such as oxidative stress (Ada1, Yap1, MetR) and virulence (Hac1, MetR, Yap1, Ada1) at varying degrees. Transcriptome data demonstrated that the inactivation of bZIPs (Hac1, Atf1, Ada1 and Yap1) significantly affected many genes in multiple critical metabolism pathways and gene ontology (GO) terms. Moreover,the ΔHac1 mutants displayed reduced aerial hypha and are hypersensitivity to endoplasmic reticulum disruptors such as tunicamycin and dithiothreitol. Transcriptome analysis showed that inactivation of Hac1 significantly affected the proteasome process and its downstream unfolded protein binding, indicating that Hac1 participates in the endoplasmic reticulum stress response through the conserved unfolded protein response. Taken together, our findings reveal that bZIP transcription factors function as key regulators of fungal morphogenesis, abiotic stress response and pathogenesis, and expand our understanding of how microbial pathogens utilize these genes to deal with environmental stresses and achieve successful infection in the host plant.

Antibacterial and antioxidant activities of endophytic fungi and nettle (Urtica dioica L.) leaves as their host.

Nettle (Urtica dioica L), as a plant rich in biologically active compounds, is one of the most important plants used in herbal medicine. Studies have shown that this plant has antioxidant, antiplatelet, hypoglycemic and hypocholesterolemia effects. In this study, we characterized three Alternaria endophytic fungi isolated from their host U. dioica. We hypothesized that these endophytic fungi can produce new bioactive metabolites, which may possess the bioactive property with potential application in the medical and pharmaceutical industries. The antibacterial activity was evaluated against reference and isolated strains, including Methicillin-Resistant Staphylococcus aureus. A wide range of antimicrobial activities similar to those measured in nettle leaves was detected especially for Alternaria sorghi. Furthermore, the highest antioxidant activity detected with DPPH free radical scavenging was measured for A. sorghi and nettle leaves ethyl acetate extracts. In addition, whereas catalase activity was similar in the three isolated fungi and nettle leaves, total thiol content and superoxide dismutase activity were significantly higher in leaves. A. sorghi showed the best activities compared to other isolated fungi. The characterization and further production of bioactive compounds produced by this endophyte should be investigated to fight bacteria and especially those that develop drug multi-resistance.

Estrogen Receptor ß Participates in Alternariol-Induced Oxidative Stress in Normal Prostate Epithelial Cells.

Alternaria toxins are considered as emerging mycotoxins, however their toxicity has not been fully evaluated in humans. Alternariol (AOH), the most prevalent Alternaria mycotoxin, was previously reported to be genotoxic and to affect hormonal balance in cells; however, its direct molecular mechanism is not known. The imbalance in androgen/estrogen ratio as well as chronic inflammation are postulated as factors in prostate diseases. The environmental agents affecting the hormonal balance might participate in prostate carcinogenesis. Thus, this study evaluated the effect of two doses of AOH on prostate epithelial cells. We observed that AOH in a dose of 10 µM induces oxidative stress, DNA damage and cell cycle arrest and that this effect is partially mediated by estrogen receptor ß (ERß) whereas the lower tested dose of AOH (0.1 µM) induces only oxidative stress in cells. The modulation of nuclear erythroid-related factor 2 (Nrf2) was observed in response to the higher dose of AOH. The use of selective estrogen receptor ß (ERß) inhibitor PHTPP revealed that AOH-induced oxidative stress in both tested doses is partially dependent on activation of ERß, but lack of its activation did not protect cells against AOH-induced ROS production or DNA-damaging effect in case of higher dose of AOH (10 µM). Taken together, this is the first study reporting that AOH might affect basic processes in normal prostate epithelial cells associated with benign and malignant changes in prostate tissue.

CCR -NB-LRR proteins MdRNL2 and MdRNL6 interact physically to confer broad-spectrum fungal resistance in apple (Malus × domestica).

Apple leaf spot, a disease caused by Alternaria alternata f. sp. mali and other fungal species, leads to severe defoliation and results in tremendous losses to the apple (Malus × domestica) industry in China. We previously identified three RPW8, nucleotide-binding, and leucine-rich repeat domain CCR -NB-LRR proteins (RNLs), named MdRNL1, MdRNL2, and MdRNL3, that contribute to Alternaria leaf spot (ALT1) resistance in apple. However, the role of NB-LRR proteins in resistance to fungal diseases in apple remains poorly understood. We therefore used MdRNL1/2/3 as baits to screen ALT1-inoculated leaves for interacting proteins and identified only MdRNL6 (another RNL) as an interactor of MdRNL2. Protein interaction assays demonstrated that MdRNL2 and MdRNL6 interact through their NB-ARC domains. Transient expression assays in apple indicated that complexes containing both MdRNL2 and MdRNL6 are necessary for resistance to Alternaria leaf spot. Intriguingly, the same complexes were also required to confer resistance to Glomerella leaf spot and Marssonina leaf spot in transient expression assays. Furthermore, stable transgenic apple plants with suppressed expression of MdRNL6 showed hypersensitivity to Alternaria leaf spot, Glomerella leaf spot, and Marssonina leaf spot; these effects were similar to the effects of suppressing MdRNL2 expression in transgenic apple plantlets. The identification of these novel broad-spectrum fungal resistance genes will facilitate breeding for fungal disease resistance in apple.

Mesenchymal Stem Cells Suppress Severe Asthma by Directly Regulating Th2 Cells and Type 2 Innate Lymphoid Cells.

Patients with severe asthma have unmet clinical needs for effective and safe therapies. One possibility may be mesenchymal stem cell (MSC) therapy, which can improve asthma in murine models. However, it remains unclear how MSCs exert their beneficial effects in asthma. Here, we examined the effect of human umbilical cord blood-derived MSCs (hUC-MSC) on two mouse models of severe asthma, namely, Alternaria alternata-induced and house dust mite (HDM)/diesel exhaust particle (DEP)-induced asthma. hUC-MSC treatment attenuated lung type 2 (Th2 and type 2 innate lymphoid cell) inflammation in both models. However, these effects were only observed with particular treatment routes and timings. In vitro co-culture showed that hUC-MSC directly downregulated the interleukin (IL)-5 and IL-13 production of differentiated mouse Th2 cells and peripheral blood mononuclear cells from asthma patients. Thus, these results showed that hUC-MSC treatment can ameliorate asthma by suppressing the asthmogenic cytokine production of effector cells. However, the successful clinical application of MSCs in the future is likely to require careful optimization of the route, dosage, and timing.

Overexpression of LYK4, a lysin motif receptor with non-functional kinase domain, enhances tolerance to Alternaria brassicicola and increases trichome density in Brassica juncea.

Lysin motif receptor-like kinases (LYKs) are involved in the recognition of chitin and activation of plant immune response. In this study, we found LYK4 to be strongly induced in resistant Sinapis alba compared with susceptible Brassica juncea on challenge with Alternaria brassicicola. In silico analysis and in vitro kinase assay revealed that despite the presence of canonical protein kinase fold, B.juncea LYK4 (BjLYK4) lacks several key residues of a prototype protein kinase which renders it catalytically inactive. Transient expression analysis confirmed that fluorescently tagged BjLYK4 localizes specifically to the plasma membrane. Overexpression (OE) of BjLYK4 in B. juncea enhanced tolerance against A. brassicicola. Interestingly, the OE lines also exhibited a novel trichome dense phenotype and increased jasmonic acid (JA) responsiveness. We further showed that many chitin responsive WRKY transcription factors and JA biosynthetic genes were strongly induced in the OE lines on challenge with the pathogen. Moreover, several JA inducible trichome developmental genes constituting the WD-repeat/bHLH/MYB activator complex were also upregulated in the OE lines compared with vector control and RNA interference line. These results suggest that BjLYK4 plays an essential role in chitin-dependent activation of defense response and chitin independent trichome development likely by influencing the JA signaling pathway.

Overexpression of SlGSNOR impairs in vitro shoot proliferation and developmental architecture in tomato but confers enhanced disease resistance.

The pervasive presence of nitric oxide (NO) in cells and its role in modifying cystein residues through protein S-nitrosylation is a remarkable redox based signalling mechanism regulating a variety of cellular processes. S-NITROSOGLUTATHIONE REDUCTASE (GSNOR) governs NO bioavailability by the breakdown of S-nitrosoglutathione (GSNO), fine-tunes NO signalling and controls total cellular S-nitrosylated proteins. Most of the published data on GSNOR functional analysis is based on the model plant Arabidopsis with no previous report for its effect on in vitro regeneration of tissue cultured plants. Moreover, the effect of GSNOR overexpression (O.E) on tomato growth, development and disease resistance remains enigmatic. Here we show that SlGSNOR O.E in tomato alters multiple developmental programs from in vitro culture establishment to plant growth and fruit set. Moreover, constitutive SlGSNOR O.E in tomato showed enhanced resistance against early blight (EB) disease caused by Alternaria solani and reduction in hypersensitive response (HR)-mediated cell death after Pseudomonas syringae pv. tomato DC3000 (Pst DC3000) infiltrations. High GSNOR transcript levels led to the inhibition of in vitro shoot proliferation in transformed explants as revealed by the fluorescence microscopy after YFP labelling. Transgenic tomato lines overexpressing SlGSNOR showed defective phenotypes exhibiting stunted plant growth and bushy-type plants due to loss of apical dominance, along with reduced seed germination and delayed flowering. Furthermore, SlGSNOR O.E plants exhibited altered leaf arrangement, fruit shape and modified locules number in tomato fruit. These findings give a novel insight into a multifaceted regulatory role of SlGSNOR in tomato plant development, reproduction and response to pathogens.

Suppression of Alternaria brassicicola infection by volatile compounds from spent mushroom substrates.

Most commercially circulating mushrooms are produced via cultivation using artificially produced mushroom substrates. However, after mushroom harvesting, the disposal of spent mushroom substrates (SMSs) is a serious problem for the mushroom industry owing to the need for a disposal site and the cost involved. Thus, in view of the possibility of recycling SMSs as a soil modifier, we examined the effect of soil mixed with SMSs on the infection of Arabidopsis leaves by Alternaria brassicicola, the causal agent of cabbage leaf spot. The mixing of SMSs used for Hypsizygus marmoreus, Pholiota microspora, Lyophyllum decastes, and Auricularia polytricha into culture soil suppressed the lesion formation caused by A. brassicicola. The defense responses of Arabidopsis were not induced by the culturing of these seedlings in soils containing SMSs. Suppressed lesion formation was observed after the seedlings were treated with volatiles emitted from SMSs that were incubated with soil for 7 days and used for H. marmoreus, P. microspora, L. decastes, A. polytricha, Lentinula edodes, and Cyclocybe cylindracea. The volatiles from the SMSs reduced the elongation of A. brassicicola hyphae. GC-MS analyses of extracts from the SMS containing soils led to the detection of various volatile compounds; among these, skatole, 2,4-di-tert-butylphenol, γ-dodecalactone, butyric acid, guaiacol, 6-amyl-2-pyrone, and 1-octen-3-ol were examined for inhibitory activity on A. brassicicola and found to suppress hyphae elongation. These findings indicate that the antifungal volatile compounds emitted by the SMSs suppress A. brassicicola infection.