Defense Genes Research Articles

Deciphering key factors in pathogen-suppressive microbiome assembly in the rhizosphere.

In a plant-microbe symbiosis, the host plant plays a key role in promoting the association of beneficial microbes and maintaining microbiome homeostasis through microbe-associated molecular patterns (MAMPs). The associated microbes provide an additional layer of protection for plant immunity and help in nutrient acquisition. Despite identical MAMPs in pathogens and commensals, the plant distinguishes between them and promotes the enrichment of beneficial ones while defending against the pathogens. The rhizosphere is a narrow zone of soil surrounding living plant roots. Hence, various biotic and abiotic factors are involved in shaping the rhizosphere microbiome responsible for pathogen suppression. Efforts have been devoted to modifying the composition and structure of the rhizosphere microbiome. Nevertheless, systemic manipulation of the rhizosphere microbiome has been challenging, and predicting the resultant microbiome structure after an introduced change is difficult. This is due to the involvement of various factors that determine microbiome assembly and result in an increased complexity of microbial networks. Thus, a comprehensive analysis of critical factors that influence microbiome assembly in the rhizosphere will enable scientists to design intervention techniques to reshape the rhizosphere microbiome structure and functions systematically. In this review, we give highlights on fundamental concepts in soil suppressiveness and concisely explore studies on how plants monitor microbiome assembly and homeostasis. We then emphasize key factors that govern pathogen-suppressive microbiome assembly. We discuss how pathogen infection enhances plant immunity by employing a cry-for-help strategy and examine how domestication wipes out defensive genes in plants experiencing domestication syndrome. Additionally, we provide insights into how nutrient availability and pH determine pathogen suppression in the rhizosphere. We finally highlight up-to-date endeavors in rhizosphere microbiome manipulation to gain valuable insights into potential strategies by which microbiome structure could be reshaped to promote pathogen-suppressive soil development.

Trichoderma harzianum Cellulase Gene thph2 Affects Trichoderma Root Colonization and Induces Resistance to Southern Leaf Blight in Maize.

Trichoderma, widely distributed all over the world, is commonly found in soil and root ecosystems. It is a group comprising beneficial fungi that improve plant disease resistance and promote plant growth. Studies have shown that Trichoderma cellulases can also improve plant disease resistance. Based on previous studies, we reported that a C6 zinc finger protein (Thc6) regulates two cellulase genes, thph1 and thph2, to induce ISR responses in plants. Therefore, in this study, we focused on the role of thph2 in the colonization of maize roots by T. harzianum and the induction of systemic resistance against southern leaf blight. The results showed that thph2 had a positive regulatory effect on the Trichoderma colonization of maize roots. After the root was treated with Trichoderma, the leaf defense genes AOS, LOX5, HPL, and OPR1 were expressed to resist the attack of Cochliobolus heterostrophus. The pure Thph2 protein also resulted in a similar induction activity of the AOS, LOX5, HPL, and OPR1 expression in maize roots, further demonstrating that thph2 can induce plant defense responses and that signal transduction occurs mainly through the JA signaling pathway.

Heterologous expression of influenza haemagglutinin leads to early and transient activation of the unfolded protein response in Nicotiana benthamiana.

The unfolded protein response (UPR) allows cells to cope with endoplasmic reticulum (ER) stress induced by accumulation of misfolded proteins in the ER. Due to its sensitivity to Agrobacterium tumefaciens, the model plant Nicotiana benthamiana is widely employed for transient expression of recombinant proteins of biopharmaceutical interest, including antibodies and virus surface proteins used for vaccine production. As such, study of the plant UPR is of practical significance, since enforced expression of complex secreted proteins often results in ER stress. After 6 days of expression, we recently reported that influenza haemagglutinin H5 induces accumulation of UPR proteins. Since up-regulation of corresponding UPR genes was not detected at this time, accumulation of UPR proteins was hypothesized to be independent of transcriptional induction, or associated with early but transient UPR gene up-regulation. Using time course sampling, we here show that H5 expression does result in early and transient activation of the UPR, as inferred from unconventional splicing of NbbZIP60 transcripts and induction of UPR genes with varied functions. Transient nature of H5-induced UPR suggests that this response was sufficient to cope with ER stress provoked by expression of the secreted protein, as opposed to an antibody that triggered stronger and more sustained UPR activation. As up-regulation of defence genes responding to H5 expression was detected after the peak of UPR activation and correlated with high increase in H5 protein accumulation, we hypothesize that these immune responses, rather than the UPR, were responsible for onset of the necrotic symptoms on H5-expressing leaves.

BZIP transcription factor FabR: Redox-dependent mechanism controlling docosahexaenoic acid biosynthesis and H2O2 stress response in Schizochytrium sp.

Schizochytrium sp. is an important industrial strain for commercial production of docosahexaenoic acid (DHA), which plays essential physiological roles in infant development and human health. The regulatory network for DHA biosynthesis and lipid accumulation in Schizochytrium remains poorly understood. FabR (fatty acid biosynthesis repressor), a basic leucine zipper (bZIP) transcription factor, was transcriptionally downregulated under low-nitrogen condition. Deletion of fabR gene (mutant ΔfabR) increased production of total lipids and DHA by 30.1% and 46.5%, respectively. ΔfabR displayed H2O2 stress resistance higher than that of parental strain or complementation strain CfabR. FabR bound specifically to 7-bp pseudo-palindromic sequence 5′-ATTSAAT-3′ in upstream regions and repressed transcription of fatty acid biosynthesis genes (acl, fas, pfa) and antioxidant defense genes (cat, sod1, sod2, gpx). DNA binding activity of FabR was regulated in a redox-dependent manner. Under oxidative condition, FabR forms intermolecular disulfide bonds between two Cys46 residues of dimers; its DNA binding activity is thereby lost, and the transcription of its target genes is enhanced through derepression. Our findings clarify the redox-dependent mechanism that modulates FabR activity governing lipid and DHA biosynthesis and H2O2 stress response in Schizochytrium.

Feeding Assay to Study the Effect of Phytocytokines on Direct and Indirect Defense in Maize.

Phytocytokines mediate defense against pests and pathogens. Many methods have been developed to study the physiological responses triggered by phytocytokines in dicot plants. Here, we describe a detailed peptide feeding protocol to study the effect of phytocytokines on direct and indirect anti-herbivore defense in maize. This method relies on peptide uptake by the excised maize seedling or leaves via the transpiration stream. The headspace volatiles from plant samples are then analyzed by proton transfer reaction time-of-flight mass spectrometry (PTR-TOF-MS), or by gas chromatography-mass spectrometry (GC-MS). The samples can also be further processed to evaluate phytocytokine-induced defense gene expression or phytohormone production.

Ropeginterferon Alfa-2b Blunts Hyperactive JAK2 Activity in Polycythemia Vera and Essential Thrombocythemia

Polycythemia vera (PV) and essential thrombocythemia (ET) are myeloproliferative neoplasms (MPNs) characterized by phenotype defining somatic mutations of JAK2, cMPL, and CALR genes. These mutations activate the JAK-STAT signaling pathway, leading to augmented cellular proliferation. A major cause of morbidity and mortality in PV and ET is thrombosis. Current therapy options include hydroxyurea (HU), interferon-α (IFN-α), and JAK2 inhibitors. IFN-α treatment normalizes blood cell counts, and reduces thrombotic complications in PV and ET and has been associated with a decrease in JAK2V617F variant allele frequency (VAF), indicating its selective targeting of JAK2V617F-mutated cells. IFN-α treatment also rescues the normal dormant HSCs, resulting in conversion of clonal to polyclonal myelopoiesis (Tashi, Leukemia, 2018). Ropeginterferon alfa (ropeg-IFNα)-2b is mono-pegylated IFNα-2b with greater tolerability and an extended half-life compared to Peg-IFNα-2α and is FDA approved for PV therapy. We recently discovered a patient with a novel germline JAK2R715T mutation, which causes erythrocytosis with PV-like features (Kim, ASH, 2022, Song under review in Blood). This patient had poor tolerance to HU and a JAK2 inhibitor therapy; although the latter normalized her blood cell counts, it was poorly tolerated. She has been on ropeg-IFNα-2b since Mar. 2023. Given that ropeg-IFNα-2b's known function is to reduce mutated cells (Verger, Blood Cancer J. 2018) and all her hematopoietic cells were bearing a JAK2 gain-of-function germline mutation, we did not expect a complete hematological response, but her blood counts completely normalized. This observation led us to hypothesize that ropeg-IFNα-2b induces remission by reduction of JAK2 activity. To measure JAK2 activity, we quantitated phosphorylated STAT5 (p-STAT5) levels in her erythroid progenitors (CD71 +/CD235a +) and observed a 20% reduction in these levels after ropeg-IFNα-2b treatment. This prompted us to investigate PV and ET patients with the typical JAK2V617F mutation treated with ropeg-IFNα-2b. We measured p-STAT5 in erythroid progenitors and platelets (CD61 +) of 8 PV and 5 ET patients treated with ropeg-IFNα-2b and 1 PV and 6 ET patients treated with HU. Levels of p-STAT5 were higher in patients treated with HU compared to those treated with Ropeg-IFNα-2b (Figure A and B). In healthy controls, levels of p-STAT5 were lower than those in HU treated patients but comparable to those in ropeg-IFNα-2b treated patients (Figure A and B). p-STAT5 levels showed a positive correlation with platelet counts (r=0.5235, p=0.0397) and neutrophil counts (r=0.6088, p=0.0235). p-STAT5 levels were then measured before and after ropeg-IFNα-2b treatment in 1 ET patient and showed a 20% reduction, which coincided with a decrease in platelet count from 746 x 10 9/L to 207 x 10 9/L. Peg-IFN-2α downregulates upregulated oxidative stress genes and upregulates downregulated antioxidative defense genes in MPNs (Skov, PLOS One, 2022), suggesting its potential role in reducing oxidative stress.We measured reactive oxygen species (ROS) in erythroid progenitors and found that ROS levels were significantly higher in PV and ET patients treated with HU compared to those treated with ropeg-IFNα-2b and to healthy controls (Figure C). JAK2V617F mutated cells accumulate more ROS compared to unmutated cells (Marty, Leukemia, 2013). We found a positive correlation between ROS levels and p-STAT5 levels (r=0.4778, p=0.0449), suggesting an interaction between JAK2 activity and oxidative stress in PV and ET. Our study demonstrated the impact of ropeg-IFNα-2b treatment on JAK2 activity in PV and ET. Ropeg-IFNα-2b effectively reduced JAK2 activity, leading to suppression of dysregulated cell proliferation, and decreased ROS accumulation. To minimize the influence of confounding factors that could independently affect JAK2 activity, we are now comparing samples from the same patients before and after ropeg-IFNα-2b treatment. These results suggest a novel molecular benefit of ropeg-IFNα-2b therapy in PV and ET.

Molecular characterization and functional analysis of a beta-1,3-glucan recognition protein from oriental fruit moth Grapholita molesta (Lepidoptera: Tortricidae).

The beta-1,3-glucan recognition protein (BGRP) is an important pattern recognition protein (PRP), which plays an important role in immune recognition and signaling pathway of insect innate immunity. Herein, a BGRP gene was obtained from the transcriptome of Grapholita molesta and its expression was verified by PCR. The full cDNA of the GmBGRP gene was 1691 bp encoding 486 amino acid residues. The calculated molecular mass of the mature protein was 54.83 kDa with an estimated pI of 6.14. The amino acid sequence of GmBGRP was highly homologous to BGRPs of other lepidopterans including Leguminivora glycinivorella BGRP-3. Expression profile of GmBGRP at different developmental stages and different tissues of 5th instar larvae showed that the expression level of this gene tends to slightly increase and then decrease at the adult stage, with the highest at the pupa stage; and mainly expressed in the epidermis, fat body and hemocytes compared with other tissues. In addition, we investigated the transcription levels of other immune-related genes, such as Serine-1, Serine-2, Serine-3, Serpin, SRCB (scavenger receptor gene), Toll, PPO (prophenoloxidase) upon GmBGRP gene silencing, indicating that GmBGRP expression is associated with immune responses of G. molesta. This was further supported by the upregulation of the mRNA level of GmBGRP following fungal infection. Taken together, these results provide experimental evidence for the role of GmBGRP gene in immune defense in G. molesta larvae.

Wheat stripe rust resistance locus YR63 is a hot spot for evolution of defence genes – a pangenome discovery

BackgroundStripe rust, caused by Puccinia striiformis f. sp. tritici (Pst), poses a threat to global wheat production. Deployment of widely effective resistance genes underpins management of this ongoing threat. This study focused on the mapping of stripe rust resistance gene YR63 from a Portuguese hexaploid wheat landrace AUS27955 of the Watkins Collection.ResultsYR63 exhibits resistance to a broad spectrum of Pst races from Australia, Africa, Asia, Europe, Middle East and South America. It was mapped to the short arm of chromosome 7B, between two single nucleotide polymorphic (SNP) markers sunCS_YR63 and sunCS_67, positioned at 0.8 and 3.7 Mb, respectively, in the Chinese Spring genome assembly v2.1. We characterised YR63 locus using an integrated approach engaging targeted genotyping-by-sequencing (tGBS), mutagenesis, resistance gene enrichment and sequencing (MutRenSeq), RNA sequencing (RNASeq) and comparative genomic analysis with tetraploid (Zavitan and Svevo) and hexaploid (Chinese Spring) wheat genome references and 10+ hexaploid wheat genomes. YR63 is positioned at a hot spot enriched with multiple nucleotide-binding and leucine rich repeat (NLR) and kinase domain encoding genes, known widely for defence against pests and diseases in plants and animals. Detection of YR63 within these gene clusters is not possible through short-read sequencing due to high homology between members. However, using the sequence of a NLR member we were successful in detecting a closely linked SNP marker for YR63 and validated on a panel of Australian bread wheat, durum and triticale cultivars.ConclusionsThis study highlights YR63 as a valuable source for resistance against Pst in Australia and elsewhere. The closely linked SNP marker will facilitate rapid introgression of YR63 into elite cultivars through marker-assisted selection. The bottleneck of this study reinforces the necessity for a long-read sequencing such as PacBio or Oxford Nanopore based techniques for accurate detection of the underlying resistance gene when it is part of a large gene cluster.

Case-Control Study of the Association between Single Nucleotide Polymorphisms of Genes Involved in Xenobiotic Detoxification and Antioxidant Protection with the Long-Term Influence of Organochlorine Pesticides on the Population of the Almaty Region.

The association of genetic polymorphisms with the individual sensitivity of humans to the action of pesticide pollution is being actively studied in the world. The aim of this study was a molecular epidemiological analysis of candidate polymorphisms of genes involved in pesticide metabolism, detoxification, and antioxidant protection. Some of the selected polymorphisms also relate to susceptibility to cancer and cardiovascular, respiratory, and immune system diseases in individuals exposed to pesticides for a long time. For a case-control study of a unique cohort of people exposed to organochlorine pesticides for 10 years or more were chosen, a control cohort was selected that matched with the experimental group by the main population characteristics. PCR-PRLF and genome-wide microarray genotyping (GWAS) methods were used. We identified 17 polymorphisms of xenobiotic detoxification genes and 27 polymorphisms of antioxidant defense genes, which had a significantly high statistical association with the negative impact of chronic pesticide intoxication on human health. We also found 17 polymorphisms of xenobiotic detoxification genes and 12 polymorphisms of antioxidant defense genes that have a protective effect. Data obtained added to the list of potential polymorphisms that define a group at high risk or resistant to the negative effects of pesticides.

Identification and immune analysis of antimicrobial peptides from the cigarette beetle (Lasioderma serricorne).

Antimicrobial peptides (AMPs) in insects are endogenous peptides that are effector components of the innate defense system of the insect. AMPs may serve as antimicrobial agents because of their small molecular weight and broad-spectrum antimicrobial activity. In this study, we performed transcriptome analysis of cigarette beetle (Lasioderma serricorne) larvae, parasitized by the ectoparasitic wasp, Anisopteromalus calandrae. Several AMP genes were significantly upregulated following A.calandrae parasitism, postulating the hypothesis that the parasitization enhanced the host's resistance against pathogenic microorganisms through the regulation of host AMP genes. Specifically, 3 AMP genes (LsDef1, LsDef2, and LsCole) were significantly upregulated and we studied their immune function in L.serricorne. Immune challenge and functional analysis showed that LsCole was responsible for the immune response against Gram-negative and Gram-positive bacteria, while LsDef1 and LsDef2 were involved in insect defense against Gram-positive bacteria. Purified recombinant LsCole exhibited antimicrobial activities against the Gram-negative bacterium Escherichia coli and the Gram-positive bacterium Staphylococcus aureus. LsDef2 showed an antibacterial effect against S.aureus. LsCole and LsDef2 exhibited antibiofilm activity against S.aureus. The 2 AMPs disrupted cell membranes and caused leakage of S.aureus cell contents. The results indicated that the 3 AMPs in L.serricorne are involved in the innate immunity of this pest insect. These AMPs may have potential as antimicrobial agents for bacterial infection chemotherapy. Hence, data are discussed in relation to new control strategies with greater biosafety against pest insects with use of microbial biocontrol agents in combination with RNA interference against the insect's defensive AMP genes.

LHP1-mediated epigenetic buffering of subgenome diversity and defense responses confers genome plasticity and adaptability in allopolyploid wheat

Polyploidization is a major driver of genome diversification and environmental adaptation. However, the merger of different genomes may result in genomic conflicts, raising a major question regarding how genetic diversity is interpreted and regulated to enable environmental plasticity. By analyzing the genome-wide binding of 191 trans-factors in allopolyploid wheat, we identified like heterochromatin protein 1 (LHP1) as a master regulator of subgenome-diversified genes. Transcriptomic and epigenomic analyses of LHP1 mutants reveal its role in buffering the expression of subgenome-diversified defense genes by controlling H3K27me3 homeostasis. Stripe rust infection releases latent subgenomic variations by eliminating H3K27me3-related repression. The simultaneous inactivation of LHP1 homoeologs by CRISPR–Cas9 confers robust stripe rust resistance in wheat seedlings. The conditional repression of subgenome-diversified defenses ensures developmental plasticity to external changes, while also promoting neutral-to-non-neutral selection transitions and adaptive evolution. These findings establish an LHP1-mediated buffering system at the intersection of genotypes, environments, and phenotypes in polyploid wheat. Manipulating the epigenetic buffering capacity offers a tool to harness cryptic subgenomic variations for crop improvement.

β-Aminobutyric Acid and Powdery Mildew Infection Enhanced the Activation of Defense-Related Genes and Salicylic Acid in Cucumber (Cucumis sativus L.).

Powdery mildew disease, caused by Sphaerotheca fusca, is a major disease affecting cucumbers cultivated in greenhouses. This study was conducted to find defense genes induced by β-aminobutyric acid (BABA) and powdery mildew in cucumber. Disease severities of 25% and 5% were exhibited by the 2000 and 5000 mg/L BABA-treated cucumber, respectively. BABA did not affect the spore germination of the powdery mildew pathogen, showing that BABA is not an antifungal agent against the pathogen. In quantitative real-time PCR analysis, BABA-treated cucumber upregulated the transcriptional levels of the defense genes CsPAL, CsPR3, CsPR1, CsLOX1, CsLOX23, Cs LecRK6.1, CsWRKY20, and Cupi4 in cucumber to maximum levels at 48 h, whereas CsLecRK6.1 reached maximum expression after 24 h, and further, salicylic acid (SA) levels were significantly increased in BABA-treated cucumber plants. In addition, the cucumber infected with powdery mildew underwent a 1.6- to 47.3-fold enhancement in the defense genes PAL, PR3, PR1, Lox1, Lox 23, LecRK6.1, WRKY20, and Cupi4 compared to heathy cucumber. These results suggest that the BABA-induced defense response is associated with SA signaling pathway-dependent systemic acquired resistance (SAR) in cucumber, which is involved in plant resistance mechanisms.

Molecular Regulatory Mechanism of Exogenous Hydrogen Sulfide in Alleviating Low-Temperature Stress in Pepper Seedlings.

Pepper (Capsicum annuum L.) is sensitive to low temperatures, with low-temperature stress affecting its plant growth, yield, and quality. In this study, we analyzed the effects of exogenous hydrogen sulfide (H2S) on pepper seedlings subjected to low-temperature stress. Exogenous H2S increased the content of endogenous H2S and its synthetase activity, enhanced the antioxidant capacity of membrane lipids, and protected the integrity of the membrane system. Exogenous H2S also promoted the Calvin cycle to protect the integrity of photosynthetic organs; enhanced the photosynthetic rate (Pn), stomatal conductance (Gs), transpiration rate (Tr), and photosynthesis; and reduced the intercellular CO2 concentration (Ci). Moreover, the activities of superoxide dismutase, peroxidase, catalase, and anti-cyclic glutathione (ASA-GSH) oxidase were improved to decompose excess reactive oxygen species (ROS), enhance the oxidative stress and detoxification ability of pepper seedlings, and improve the resistance to low-temperature chilling injury in 'Long Yun2' pepper seedlings. In addition, the H2S scavenger hypotaurine (HT) aggravated the ROS imbalance by reducing the endogenous H2S content, partially eliminating the beneficial effects of H2S on the oxidative stress and antioxidant defense system, indicating that H2S can effectively alleviate the damage of low temperature on pepper seedlings. The results of transcriptome analysis showed that H2S could induce the MAPK-signaling pathway and plant hormone signal transduction; upregulate the expression of transcription factors WRKY22 and PTI6; induce defense genes; and activate the ethylene and gibberellin synthesis receptors ERF1, GDI2, and DELLA, enhancing the resistance to low-temperature chilling injury of pepper seedlings. The plant-pathogen interaction was also significantly enriched, suggesting that exogenous H2S also promotes the expression of genes related to plant-pathogen interaction. The results of this study provide novel insights into the molecular mechanisms and genetic modifications of H2S that mitigate the hypothermic response.

Comparative proteomic analysis of the moderately resistant and susceptible peanut cultivars during infestation by Cylindrocladium parasiticum

Cylindrocladium parasiticum Crous, Wingfield & Alfenas, the causal agent of Cylindrocladium black rot of peanut, has led to economic losses in China. However, the response of peanut plant to C. parasiticum is poorly understood. Here, two contrasting peanut cultivars, T09 (C. parasiticum-moderately resistant) and P562 (C. parasiticum-susceptible) were used for comparative analysis of protein profiles in root segment of plant tissues in responses to C. parasiticum infection. Proteomic profiling identified 1647 and 391 differentially accumulated proteins (DAPs) in P562 and T09 relative to uninoculated control, respectively. A total of 350 and 1095 DAPs were identified between P562 and T09 after 9 day(s) post infection (dpi), respectively. Functional categorization showed that C. parasiticum-responsive proteins were mainly involved in catalytic activity and binding functions. KEGG pathway analysis indicated both moderately resistant and susceptible peanut cultivars can regulate gene expression in the phenylpropanoid pathway, terpenoid backbone biosynthesis, SA, and JA pathways to induce defensive genes and protein expression which enhances plant defence or susceptibility capacity. However, MAPK signal pathway was more pronounced in moderately resistant peanut cultivar T09. We also observed an increase of CYP73A100 involved in phenylpropanoid and flavonoid biosynthesis pathways in susceptible peanut ecotype P562, while decrease in moderately resistant interaction. Additionally, there was a marked activation of brassinosteroid biosynthesis in T09, which indicated a possible involvement of activation of plant immune response in moderately resistant responses of peanut to C. parasiticum. This study provides some insights into molecular networks involved in cellular and physiological responses to C. parasiticum in peanut.

Eliciting transcriptomic and antioxidant defensive responses against Rhizoctonia root rot of sorghum using the endophyte Aspergillus oryzae YRA3

Environmental pollution due to the improper use of the chemical fungicides represents a vital ecological problem, which affects human and animal health, as well as the microbial biodiversity and abundance in the soil. In this study, an endophytic fungus Aspergillus oryzae YRA3, isolated from the wild plant Atractylis carduus (Forssk.) C.Chr, was tested for its biocontrol activity against Rhizoctonia root rot of sorghum. The antagonistic potential of A. oryzae YRA3 was tested against Rhizoctonia solani in vitro. A full inhibition in the growth of R. solani was recorded indicating a strong antagonistic potential for this endophyte. To investigate the chemical composition of its metabolites, GC/MS analysis was used and thirty-two compounds in its culture filtrate were identified. Among these metabolites, some compounds with an antifungal background were detected including palmitic acid, 2-heptanone, and 2,3-butanediol. To these antifungal metabolites the antagonistic activity of A. oryzae YRA3 can be attributed. In the greenhouse experiment, treating of the infected sorghum plants with A. oryzae YRA3 significantly reduced severity of the Rhizoctonia root rot by 73.4%. An upregulation of the defensive genes (JERF3), (POD) and (CHI II) was recorded in sorghum roots when were inoculated with A. oryzae YRA3. In addition, an increment in the activity of peroxidase and polyphenol oxidase, as well as the total phenolic content in the sorghum roots was also recorded. Furthermore, the results obtained from the greenhouse experiment revealed a growth-promoting effect for inoculating the sorghum plants with A. oryzae YRA3. It can be concluded that A. oryzae YRA3 can be a probable biological agent to control this disease in sorghum. However, its evaluation under field conditions is highly needed in the future studies.

Potency of CRISPR-Cas Antifungals Is Enhanced by Cotargeting DNA Repair and Growth Regulatory Machinery at the Genetic Level.

The emergence of virulent, resistant, and rapidly evolving fungal pathogens poses a significant threat to public health, agriculture, and the environment. Targeting cellular processes with standard small-molecule intervention may be effective but requires long development times and is prone to antibiotic resistance. To overcome the current limitations of antibiotic development and treatment, this study harnesses CRISPR-Cas systems as antifungals by capitalizing on their adaptability, specificity, and efficiency in target design. The conventional design of CRISPR-Cas antimicrobials, based on induction of DNA double-strand breaks (DSBs), is potentially less effective in fungi due to robust eukaryotic DNA repair machinery. Here, we report a novel design principle to formulate more effective CRISPR-Cas antifungals by cotargeting essential genes with DNA repair defensive genes that remove the fungi's ability to repair the DSB sites of essential genes. By evaluating this design on the model fungus Saccharomyces cerevisiae, we demonstrated that essential and defensive gene cotargeting is more effective than either essential or defensive gene targeting alone. The top-performing CRISPR-Cas antifungals performed as effectively as the antibiotic Geneticin. A gene cotargeting interaction analysis revealed that cotargeting essential genes with RAD52 involved in homologous recombination (HR) was the most synergistic combination. Fast growth kinetics of S. cerevisiae induced resistance to CRISPR-Cas antifungals, where genetic mutations mostly occurred in defensive genes and guide RNA sequences.

Rice foliar-adapted Pantoea species: Promising microbial biostimulants enhancing rice resilience against foliar pathogens, Magnaporthe oryzae and Xanthomonas oryzae pv. oryzae

Foliar fungal blast and bacterial leaf blight have significant impacts on rice production, and their management through host resistance and agrochemicals has proven inadequate. To achieve their sustainable management, innovative approaches like leveraging the foliar microbiome, which collaborates with plants and competes against pathogens, are essential. In our study, we isolated three Pantoea strains (P. agglomerans Os-Ep-PPA-1b, P. vagans Os-Ep-PPA-3b, and P. deleyi Os-Ep-VPA-9a) from the rice phylloplane. These isolates exhibited antimicrobial action through their metabolome and volatilome, while also promoting rice growth. Our analysis, using Gas Chromatography-Mass Spectrometry (GC-MS), revealed the presence of various antimicrobial compounds such as esters and fatty acids produced by these Pantoea isolates. Inoculating rice seedlings with P. agglomerans and P. vagans led to increased root and shoot growth. Additionally, bacterized seedlings displayed enhanced immunocompetence, as evidenced by upregulated expressions of defense genes (OsEDS1, OsFLS2, OsPDF2.2, OsACO4, OsICS OsPR1a, OsNPR1.3, OsPAD4, OsCERK1.1), along with heightened activities of defense enzymes like Polyphenol Oxidase and Peroxidase. These plants also exhibited elevated levels of total phenols. In field trials, the Pantoea isolates contributed to improved plant growth, exemplified by increased flag-leaf length, panicle number, and grains per panicle, while simultaneously reducing the incidence of chaffy grains. Hypersensitivity assays performed on a model plant, tobacco, confirmed the non-pathogenic nature of these Pantoea isolates. In summary, our study underscores the potential of Pantoea bacteria in combatting rice foliar diseases. Coupled with their remarkable growth-promoting and biostimulant capabilities, these findings position Pantoea as promising agents for enhancing rice cultivation.

WRKY Transcriptional Factor IlWRKY70 from Iris laevigata Enhances Drought and Salinity Tolerances in Nicotiana tabacum.

Drought and high salinity greatly affect plant growth and development. WRKY transcription factors play a key role in plant tolerance to abiotic stress, but the functions of WRKYs in the ornamental monocotyledon Iris laevigata remain largely unexplored. In this study, we cloned IlWRKY70 and found that it is a Group III WRKY localized in the nucleus. The expression of IlWRKY70 was induced by NaCl and PEG-6000, which reached peaks (4.38 and 5.65 times) after 3 h and 1 h, respectively. The exogenous overexpression of IlWRKY70 in N. tabacum significantly improved the resistance under NaCl and drought treatments, as evidenced by higher germination rates, longer root lengths, and increased fresh weights compared to those of control plants. In addition, transgenic seedlings showed significantly reduced wilting, higher photosynthetic performance, higher Fv/Fm and chlorophyll content, and lower stomatal conductance. Moreover, transgenic lines showed higher antioxidant enzymatic activities, lower reactive oxygen species (ROS), and lower malondialdehyde contents. Accordingly, we also found higher expressions of antioxidant defense genes, including SOD, CAT, and POD, in transgenic lines compared to controls under salt and drought stresses. Thus, IlWRKY70 enhances the abilities of salt and drought tolerances in plants, at least partially, via ROS regulation and can be used for breeding I. laevigata possessing enhanced salt and drought resistances.

The intricate role of Sir2 in oxidative stress response during the post-diauxic phase in Saccharomyces cerevisiae.

Silent information regulator 2 (Sir2) is a conserved NAD+-dependent histone deacetylase crucial for regulating cellular stress response and the aging process in Saccharomyces cerevisiae. In this study, we investigated the molecular mechanism underlying how the absence of Sir2 can lead to altered stress susceptibilities in S. cerevisiae under different environmental and physiological conditions. In a glucose-complex medium, the sir2Δ strain showed increased sensitivity to H2O2 compared to the wild-type strain during the post-diauxic phase. In contrast, it displayed increased resistance during the exponential growth phase. Transcriptome analysis of yeast cells in the post-diauxic phase indicated that the sir2Δ mutant expressed several oxidative defense genes at lower levels than the wild-type, potentially accounting for its increased susceptibility to H2O2. Interestingly, however, the sir2Δras2Δ double mutant exhibited greater resistance to H2O2 than the ras2Δ single mutant counterpart. We found that the expression regulation of the cytoplasmic catalase encoded by CTT1 was critical for the increased resistance to H2O2 in the sir2Δras2Δ strain. The expression of the CTT1 gene was influenced by the combined effect of RAS2 deletion and the transcription factor Azf1, whose level was modulated by Sir2. These findings provide insights into the importance of understanding the intricate interactions among various factors contributing to cellular stress response.

The expression of the NPR1-dependent defense response pathway genes in Persea americana (Mill.) following infection with Phytophthora cinnamomi

A plant’s defense against pathogens involves an extensive set of phytohormone regulated defense signaling pathways. The salicylic acid (SA)-signaling pathway is one of the most well-studied in plant defense. The bulk of SA-related defense gene expression and the subsequent establishment of systemic acquired resistance (SAR) is dependent on the nonexpressor of pathogenesis-related genes 1 (NPR1). Therefore, understanding the NPR1 pathway and all its associations has the potential to provide valuable insights into defense against pathogens. The causal agent of Phytophthora root rot (PRR), Phytophthora cinnamomi, is of particular importance to the avocado (Persea americana) industry, which encounters considerable economic losses on account of this pathogen each year. Furthermore, P. cinnamomi is a hemibiotrophic pathogen, suggesting that the SA-signaling pathway plays an essential role in the initial defense response. Therefore, the NPR1 pathway which regulates downstream SA-induced gene expression would be instrumental in defense against P. cinnamomi. Thus, we identified 92 NPR1 pathway-associated orthologs from the P. americana West Indian pure accession genome and interrogated their expression following P. cinnamomi inoculation, using RNA-sequencing data. In total, 64 and 51 NPR1 pathway-associated genes were temporally regulated in the partially resistant (Dusa®) and susceptible (R0.12) P. americana rootstocks, respectively. Furthermore, 42 NPR1 pathway-associated genes were differentially regulated when comparing Dusa® to R0.12. Although this study suggests that SAR was established successfully in both rootstocks, the evidence presented indicated that Dusa® suppressed SA-signaling more effectively following the induction of SAR. Additionally, contrary to Dusa®, data from R0.12 suggested a substantial lack of SA- and NPR1-related defense gene expression during some of the earliest time-points following P. cinnamomi inoculation. This study represents the most comprehensive investigation of the SA-induced, NPR1-dependent pathway in P. americana to date. Lastly, this work provides novel insights into the likely mechanisms governing P. cinnamomi resistance in P. americana.