Nonexpressor Of Pathogenesis-related Genes Research Articles

Role of NPR1 in Systemic Acquired Stomatal Immunity.

Stomatal immunity is the primary gate of the plant pathogen defense system. Non-expressor of Pathogenesis Related 1 (NPR1) is the salicylic acid (SA) receptor, which is critical for stomatal defense. SA induces stomatal closure, but the specific role of NPR1 in guard cells and its contribution to systemic acquired resistance (SAR) remain largely unknown. In this study, we compared the response to pathogen attack in wild-type Arabidopsis and the npr1-1 knockout mutant in terms of stomatal movement and proteomic changes. We found that NPR1 does not regulate stomatal density, but the npr1-1 mutant failed to close stomata when under pathogen attack, resulting in more pathogens entering the leaves. Moreover, the ROS levels in the npr1-1 mutant were higher than in the wild type, and several proteins involved in carbon fixation, oxidative phosphorylation, glycolysis, and glutathione metabolism were differentially changed in abundance. Our findings suggest that mobile SAR signals alter stomatal immune response possibly by initiating ROS burst, and the npr1-1 mutant has an alternative priming effect through translational regulation.

Non-specific LIPID TRANSFER PROTEIN 1 enhances immunity against tobacco mosaic virus in Nicotiana benthamiana.

Plant non-specific lipid transfer proteins (nsLTPs) are small, cysteine-rich proteins that play significant roles in biotic and abiotic stress responses; however, the molecular mechanism of their functions against viral infections remains unclear. In this study, we employed virus-induced gene-silencing and transgenic overexpression to functionally analyse a type-I nsLTP in Nicotiana benthamiana, NbLTP1, in the immunity response against tobacco mosaic virus (TMV). NbLTP1 was inducible by TMV infection, and its silencing increased TMV-induced oxidative damage and the production of reactive oxygen species (ROS), compromised local and systemic resistance to TMV, and inactivated the biosynthesis of salicylic acid (SA) and its downstream signaling pathway. The effects of NbLTP1-silencing were partially restored by application of exogenous SA. Overexpressing NbLTP1 activated genes related to ROS scavenging to increase cell membrane stability and maintain redox homeostasis, confirming that an early ROS burst followed by ROS suppression at the later phases of pathogenesis is essential for resistance to TMV infection. The cell-wall localization of NbLTP1 was beneficial to viral resistance. Overall, our results showed that NbLTP1 positively regulates plant immunity against viral infection through up-regulating SA biosynthesis and its downstream signaling component, NONEXPRESSOR OF PATHOGENESIS-RELATED 1 (NPR1), which in turn activates pathogenesis-related genes, and by suppressing ROS accumulation at the later phases of viral pathogenesis.

Genome-Wide Identification of the NPR1-like Gene Family in Solanum tuberosum and Functional Characterization of StNPR1 in Resistance to Ralstonia solanacearum.

The NPR1 (nonexpressor of pathogenesis-related genes 1) gene is an activator of the systemic acquisition of resistance (SAR) in plants and is one of the central factors in their response to pathogenic bacterial infestation, playing an important role in plant disease resistance. Potato (Solanum tuberosum) is a crucial non-grain crop that has been extensively studied. However, the identification and analysis of the NPR1-like gene within potato have not been understood well. In this study, a total of six NPR1-like proteins were identified in potato, and phylogenetic analysis showed that the six NPR1-like proteins in Solanum tuberosum could be divided into three major groups with NPR1-related proteins from Arabidopsis thaliana and other plants. Analysis of the exon-intron patterns and protein domains of the six NPR1-like genes from potato showed that the exon-intron patterns and protein domains of the NPR1-like genes belonging to the same Arabidopsis thaliana subfamily were similar. By performing quantitative real-time PCR (qRT-PCR) analysis, we found that six NPR1-like proteins have different expression patterns in different potato tissues. In addition, the expression of three StNPR1 genes was significantly downregulated after being infected by Ralstonia solanacearum (RS), while the difference in the expression of StNPR2/3 was insignificant. We also established potato StNPR1 overexpression lines that showed a significantly increased resistance to R. solanacearum and elevated activities of chitinase, β-1,3-glucanase, and phenylalanine deaminase. Increased peroxidase (POD), superoxide dismutase (SOD), and catalase (CAT) activities, as well as decreased hydrogen peroxide, regulated the dynamic balance of reactive oxygen species (ROS) in the StNPR1 overexpression lines. The transgenic plants activated the expression of the genes associated with the Salicylic acid (SA) defense response but suppressed the expression of the genes associated with Jasmonic acid (JA) signaling. This resulted in resistance to Ralstonia solanacearum.

NPR1 Translocation from Chloroplast to Nucleus Activates Plant Tolerance to Salt Stress.

Chloroplasts play crucial roles in biotic and abiotic stress responses, regulated by nuclear gene expression through changes in the cellular redox state. Despite lacking the N-terminal chloroplast transit peptide (cTP), nonexpressor of pathogenesis-related genes 1 (NPR1), a redox-sensitive transcriptional coactivator was consistently found in the tobacco chloroplasts. Under salt stress and after exogenous application of H2O2 or aminocyclopropane-1-carboxylic acid, an ethylene precursor, transgenic tobacco plants expressing green fluorescent protein (GFP)-tagged NPR1 (NPR1-GFP) showed significant accumulation of monomeric nuclear NPR1, irrespective of the presence of cTP. Immunoblotting and fluorescence image analyses indicated that NPR1-GFP, with and without cTP, had similar molecular weights, suggesting that the chloroplast-targeted NPR1-GFP is likely translocated from the chloroplasts to the nucleus after processing in the stroma. Translation in the chloroplast is essential for nuclear NPR1 accumulation and stress-related expression of nuclear genes. An overexpression of chloroplast-targeted NPR1 enhanced stress tolerance and photosynthetic capacity. In addition, compared to the wild-type lines, several genes encoding retrograde signaling-related proteins were severely impaired in the Arabidopsis npr1-1 mutant, but were enhanced in NPR1 overexpression (NPR1-Ox) transgenic tobacco line. Taken together, chloroplast NPR1 acts as a retrograding signal that enhances the adaptability of plants to adverse environments.

Gold nanoparticles (AuNPs) can rapidly deliver artificial microRNA (AmiRNA)-ATG6 to silence ATG6 expression in Arabidopsis.

We establish a fast and efficient transient silencing system that facilitates functional studies of some genes, whose knockout leads to plant lethality. In plants, the generation of loss-of-function mutants is crucial for studying gene function. Artificial microRNA (AmiRNA) technology is a more targeted and effective tool for gene silencing. Gold nanoparticles (AuNPs) can bind nucleic acids and deliver them into animal cells. Here, AuNPs are used in combination with AmiRNA technology in plants. We found that AmiRNA-autophagy-related proteins (ATG6) can be delivered to cells by AuNPs to achieve the effect of ATG6 silencing. It is worth noting that on the 10th day there is still a silencing effect. Similar to the atg5 lines, silencing of ATG6 significantly reduced plant resistance to Pseudomonas syringae pv.maculicola (Psm) ES4326/AvrRpt2. Interestingly, ATG6 silencing and ATG5 mutation in NPR1-GFP (nonexpressor of pathogenesis-related genes) lines significantly reduced plant resistance to Psm ES4326/AvrRpt2, suggesting that autophagy is also involved in NPR1-regulated plant immune responses. In summary, we establish a fast and efficient transient silencing system that facilitates functional studies of some genes, whose knockout leads to plant lethality.

Chemical priming of plant defense responses to pathogen attacks.

Plants can acquire an improved resistance against pathogen attacks by exogenous application of natural or artificial compounds. In a process called chemical priming, application of these compounds causes earlier, faster and/or stronger responses to pathogen attacks. The primed defense may persist over a stress-free time (lag phase) and may be expressed also in plant organs that have not been directly treated with the compound. This review summarizes the current knowledge on the signaling pathways involved in chemical priming of plant defense responses to pathogen attacks. Chemical priming in induced systemic resistance (ISR) and systemic acquired resistance (SAR) is highlighted. The roles of the transcriptional coactivator NONEXPRESSOR OF PR1 (NPR1), a key regulator of plant immunity, induced resistance (IR) and salicylic acid signaling during chemical priming are underlined. Finally, we consider the potential usage of chemical priming to enhance plant resistance to pathogens in agriculture.

Expression of Arabidopsis NPR1 (AtNPR1) in Brassica juncea var Varuna confers significant resistance against Sclerotinia sclerotiorum and Alternaria brassicae

The productivity of Brassica juncea, a major oilseed crop grown widely in South Asia, is limited by two major fungal pathogens: Sclerotinia sclerotiorum, and Alternaria brassicae. Currently, no effective resistance against these pathogens has been identified in the B. juncea germplasm. To improve the resistance against these fungal pathogens, Non-expressor of PR1 (NPR1) from Arabidopsis was expressed in the mega variety Varuna via Agrobacterium-mediated transformation. A total of 129, independent transgenic events were generated. Segregation analysis for Basta resistance and subsequent Southern hybridization identified six single-copy transgenic lines (PN23, 42, 60, 72, HN30, and HN32) which were taken forward for disease assays over two generations. Transgenic lines challenged with S. sclerotiorum exhibited a significant reduction in stem lesions ranging from 13 to 42% in the T3 generation and 4 to 31% in the T4 generation. We also observed a reduced infection against A. brassicae in the transgenic lines which developed smaller and fewer necrotic lesions as compared to untransformed lines. A correlation between the upregulation of defense marker genes and enhanced tolerance was observed in the transgenic lines upon S. sclerotiorum infection.

The SmNPR4-SmTGA5 module regulates SA-mediated phenolic acid biosynthesis in Salvia miltiorrhiza hairy roots.

Phenolic acids are the main bioactive compounds in Salvia miltiorrhiza, which can be increased by salicylic acid (SA) elicitation. However, the specific molecular mechanism remains unclear. The nonexpresser of PR genes 1 (NPR1) and its family members are essential components of the SA signaling pathway. Here, we report an NPR protein, SmNPR4, that showed strong expression in hairy root after SA treatment, acting as a negative moderator of SA-induced phenolic acid biosynthesis in S. miltiorrhiza (S. miltiorrhiza). Moreover, a basic leucine zipper family transcription factor SmTGA5 was identified and was found to interact with SmNPR4. SmTGA5 activates the expression of phenolic acid biosynthesis gene SmTAT1 through binding to the as-1 element. Finally, a series of biochemical assays and dual gene overexpression analysis demonstrated that the SmNPR4 significantly inhibited the function of SmTGA5, and SA can alleviate the inhibitory effect of SmNPR4 on SmTGA5. Overall, our results reveal the molecular mechanism of salicylic acid regulating phenolic acid biosynthesis in S. miltiorrhiza and provide new insights for SA signaling to regulate secondary metabolic biosynthesis.

Rice microRNA156/529-SQUAMOSA PROMOTER BINDING PROTEIN-LIKE7/14/17 modules regulate defenses against bacteria.

Rice (Oryza sativa L.) microRNA156/529-SQUAMOSA PROMOTER BINDING PROTEIN-LIKE7/14/17 (miR156/529-SPL7/14/17) modules have pleiotropic effects on many biological pathways. OsSPL7/14 can interact with DELLA protein SLENDER RICE1 (SLR1) to modulate gibberellin acid (GA) signal transduction against the bacterial pathogen Xanthomonas oryzae pv. oryzae. However, whether the miR156/529-OsSPL7/14/17 modules also regulate resistance against other pathogens is unclear. Notably, OsSPL7/14/17 functioning as transcriptional activators, their target genes, and the corresponding downstream signaling pathways remain largely unexplored. Here, we demonstrate that miR156/529 play negative roles in plant immunity and that miR156/529-regulated OsSPL7/14/17 confer broad-spectrum resistance against 2 devastating bacterial pathogens. Three OsSPL7/14/17 proteins directly bind to the promoters of rice Allene Oxide Synthase 2 (OsAOS2) and NONEXPRESSOR OF PATHOGENESIS-RELATED GENES1 (OsNPR1) and activate their transcription, regulating jasmonic acid (JA) accumulation and the salicylic acid (SA) signaling pathway, respectively. Overexpression of OsAOS2 or OsNPR1 impairs the susceptibility of the osspl7/14/17 triple mutant. Exogenous application of JA enhances resistance of the osspl7/14/17 triple mutant and the miR156 overexpressing plants. In addition, genetic evidence confirms that bacterial pathogen-activated miR156/529 negatively regulate pathogen-associated molecular pattern (PAMP)-triggered immunity (PTI) responses, such as pattern recognition receptor Xa3/Xa26-initiated PTI. Our findings demonstrate that bacterial pathogens modulate miR156/529-OsSPL7/14/17 modules to suppress OsAOS2-catalyzed JA accumulation and the OsNPR1-promoted SA signaling pathway, facilitating pathogen infection. The uncovered miR156/529-OsSPL7/14/17-OsAOS2/OsNPR1 regulatory network provides a potential strategy to genetically improve rice disease resistance.

Selection of reference genes for RT-qPCR analysis of rice with Rhizoctonia solani infection and biocontrol PGPR/KSi application.

Rhizoctonia solani (AG1 IA) is an important pathogen of rice (Oryza sativa L.) that causes rice sheath blight (RSB). Since control of RSB by breeding and fungicides have had limited success, novel strategies like biocontrol with plant growth-promoting rhizobacteria (PGPR) can be an effective alternative. Seven commonly used reference genes (RGs), 18SrRNA, ACT1, GAPDH2, UBC5, RPS27, eIF4a and CYP28, were evaluated for their stability in rice-R. solani-PGPR interaction for real-time quantitative PCR (RT-qPCR) analysis. Different algorithms were examined, Delta Ct, geNorm, NormFinder, BestKeeper, and comprehensive ranking by RefFinder, to evaluate RT-qPCR of rice in tissues infected with R. solani and treated with the PGPR strains, Pseudomonas saponiphilia and Pseudomonas protegens, with potassium silicate (KSi) alone or in combination with each PGPR strain. RG stability was affected for each treatment and treatment-specific RG selection was suggested. Validation analysis was done for nonexpressor of PR-1(NPR1) for each treatment. Overall, ACT1 was the most stable RG with R. solani infection alone, GAPDH2 with R. solani infection plus KSi, UBC5 with R. solani infection plus P. saponiphilia, and eIF4a with R. solani infection plus P. protegens. Both ACT1 and RPS27 were the most stable with the combination of KSi and P. saponiphilia, while RPS27 was the most stable with the combination of KSi and P. protegens.

Exogenous application of salicylic acid improves freezing stress tolerance in alfalfa.

Freezing stress is one of the most detrimental environmental factors that can seriously impact the growth, development, and distribution of alfalfa (Medicago sativa L.). Exogenous salicylic acid (SA) has been revealed as a cost-effective method of improving defense against freezing stress due to its predominant role in biotic and abiotic stress resistance. However, how the molecular mechanisms of SA improve freezing stress resistance in alfalfa is still unclear. Therefore, in this study, we used leaf samples of alfalfa seedlings pretreatment with 200 μM and 0 μM SA, which were exposed to freezing stress (-10°C) for 0, 0.5, 1, and 2h and allowed to recover at normal temperature in a growth chamber for 2 days, after which we detect the changes in the phenotypical, physiological, hormone content, and performed a transcriptome analysis to explain SA influence alfalfa in freezing stress. The results demonstrated that exogenous SA could improve the accumulation of free SA in alfalfa leaves primarily through the phenylalanine ammonia-lyase pathway. Moreover, the results of transcriptome analysis revealed that the mitogen-activated protein kinase (MAPK) signaling pathway-plant play a critical role in SA alleviating freezing stress. In addition, the weighted gene co-expression network analysis (WGCNA) found that MPK3, MPK9, WRKY22 (downstream target gene of MPK3), and TGACG-binding factor 1 (TGA1) are candidate hub genes involved in freezing stress defense, all of which are involved in the SA signaling pathway. Therefore, we conclude that SA could possibly induce MPK3 to regulate WRKY22 to participate in freezing stress to induced gene expression related to SA signaling pathway (NPR1-dependent pathway and NPR1-independent pathway), including the genes of non-expresser of pathogenesis-related gene 1 (NPR1), TGA1, pathogenesis-related 1 (PR1), superoxide dismutase (SOD), peroxidase (POD), ascorbate peroxidase (APX), glutathione-S-transferase (GST), and heat shock protein (HSP). This enhanced the production of antioxidant enzymes such as SOD, POD, and APX, which increases the freezing stress tolerance of alfalfa plants.

GIGANTEA supresses wilt disease resistance by down-regulating the jasmonate signaling in Arabidopsis thaliana.

GIGANTEA (GI) is a plant-specific nuclear protein that plays a pleiotropic role in the growth and development of plants. GI's involvement in circadian clock function, flowering time regulation, and various types of abiotic stress tolerance has been well documented in recent years. Here, the role of GI in response to Fusarium oxysporum (F. oxysporum) infection is investigated at the molecular level comparing Col-0 WT with the gi-100 mutant in Arabidopsis thaliana. Disease progression, photosynthetic parameters, and comparative anatomy confirmed that the spread and damage caused by pathogen infection were less severe in gi-100 than in Col-0 WT plants. F. oxysporum infection induces a remarkable accumulation of GI protein. Our report showed that it is not involved in flowering time regulation during F. oxysporum infection. Estimation of defense hormone after infection showed that jasmonic acid (JA) level is higher and salicylic acid (SA) level is lower in gi-100 compared to Col-0 WT. Here, we show that the relative transcript expression of CORONATINE INSENSITIVE1 (COI1) and PLANT DEFENSIN1.2 (PDF1.2) as a marker of the JA pathway is significantly higher while ISOCHORISMATE SYNTHASE1 (ICS1) and NON-EXPRESSOR OF PATHOGENESIS-RELATED GENES1 (NPR1), the markers of the SA pathway, are downregulated in the gi-100 mutants compared to Col-0 plants. The present study convincingly suggests that the GI module promotes susceptibility to F. oxysporum infection by inducing the SA pathway and inhibiting JA signaling in A. thaliana.

SlNPR1 modulates chilling stress resistance in tomato plant by alleviating oxidative damage and affecting the synthesis of ferulic acid

• NPR1 gene and protein content are dramatically induced by chilling stress. • CRISPR/Cas9-mediated SlNPR1 inhibition aggravated the symptoms of chilling injury. • SlNPR1 enhances tolerance to chilling stress by alleviating oxidative damage. • SlNPR1 enhances tolerance to chilling stress by affecting the synthesis of FA. Non-expresser of pathogenesis-related genes 1 (NPR1) has been demonstrated to be a master transcription factor during resistance against pathogens, the mechanisms by which regulates chilling stress, however, remain unclear. Our results revealed that the expression and protein content of SlNPR1 were dramatically induced by low temperature (4 °C). CRISPR/Cas9-mediated SlNPR1 mutagenesis aggravated the symptoms of chilling injury in tomato plant, which was accompanied with the accumulation of malonic dialdehyde (MDA), hydrogen peroxide (H 2 O 2 ) and superoxide anion (O 2 − ), and the decrease of soluble protein content, proline content as well as antioxidant enzymes activity. In addition, slnpr1 mutants showed lower expression of SlICE1 and SlCBF1 in contrast to wild type plants (WT) . PAL, C4H, C3H and COMT genes play important roles in the synthesis of ferulic acid (FA). We found that knockout of SlNPR1 reduced the expression of PAL, C4H, C3H and COMT induced by low temperature (4 °C) and inhibited the accumulation of FA content. Interestingly, FA-treated plants showed greater tolerance to chilling stress and displayed higher expression of SlICE1 and SlCBF1 , exhibited lower levels of MDA and H 2 O 2 , but higher antioxidant enzyme (APX, POD, SOD and CAT) activity than WT. These findings reveal a new regulatory pathway that SlNPR1 enhances tolerance to chilling stress in tomato plant by alleviating oxidative damage and affecting the synthesis of FA.

Mineral Nutrients, Physiological Disorders, Postharvest Water Loss, and PR Gene Expression in Bell Pepper (Capsicum annuum L.) Fruit under Shade Nets

Abstract Shade nets can be an effective technology for producing bell pepper (Capsicum annuum L.) under hot climatic conditions. However, the effects of shading on fruit quality are still unclear. The objectives were to evaluate the effects of shade level on fruit mineral nutrient content, physiological disorders, and postharvest water loss. Trials were conducted in the spring–summer of 2016, 2017, and 2018 in Tifton, Georgia, USA, following a randomized complete block design with five shade levels: 0% (open field), 30%, 47%, 63%, and 80%. Shading increased the bell pepper fruit dimensions (length, diameter, and weight) in 2016 and mineral nutrient content in 2017. Fruit sunscald incidence decreased with increasing shade level, while blossom-end rot showed inconsistent responses. Postharvest fruit water loss and transpiration rates were highest in fruits from the unshaded treatment in 2016; there were no differences in fruit water loss among the shade levels. NONEXPRESSOR OF PATHOGENESIS-RELATED 1 (NPR1) and PATHOGENESIS-RELATED 1 (PR1) genes expressed more than 1.5-fold and 10-fold, respectively, at 47% shade level compared to 80%, though not significantly. Therefore, plants grown under shading had fruit with greater size, increased mineral nutrient content, and reduced sunscald incidence compared with the unshaded control.

UBP12/UBP13-mediated deubiquitination of salicylic acid receptor NPR3 suppresses plant immunity

Salicylic acid (SA), a defense hormone produced after pathogen challenge, is critical for plant immunity. Arabidopsis NONEXPRESSER OF PR GENES 1 (NPR1) and its paralogs NPR3 and NPR4 can bind SA and mediate SA signal transduction. NPR1 functions as a transcriptional co-activator to promote defense gene expression, whereas NPR3 and NPR4 have been shown to function as negative regulators in the SA signaling pathway. Although the mechanism about NPR1 regulation has been well studied, how NPR3/NPR4 proteins are regulated in immune responses remains largely unknown. Here, we show that the stability of NPR3/NPR4 is enhanced by SA. In the absence of pathogen challenge, NPR3/NPR4 are unstable and degraded by the 26S proteasome, whereas the increase in cellular SA levels upon pathogen infection suppresses NPR3/NPR4 degradation. We found that UBP12 and UBP13, two homologous deubiquitinases from a ubiquitin-specific protease subfamily, negatively regulate plant immunity by promoting NPR3/NPR4 stability. Our genetic results further showed that UBP12/UBP13-mediated immunity suppression is partially dependent on NPR3/NPR4 functions. By interacting with NPR3 in the nucleus in an SA-dependent manner, UBP12 and UBP13 remove ubiquitin from polyubiquitinated NPR3 to protect it from being degraded. The stabilization of NPR3/NPR4 promoted by UBP12/UBP13 is essential for negative regulation of basal and SA-induced immunity.

Biosynthesized silver nanoparticles inhibit Pseudomonas syringae pv. tabaci by directly destroying bacteria and inducing plant resistance in Nicotiana benthamiana

Silver (Ag)-containing agents or materials are widely used today in plant protection for their antimicrobial activity. In view of the superior inhibitory ability of biosynthesized (aldehyde-modified sodium alginate based) silver nanoparticles (AgNPs) against plant pathogenic fungi in our previous research, here we explored the antagonistic effect of biosynthesized AgNPs on plant pathogenic bacteria and the underlying mechanism. We selected Pseudomonas syringae pv. tabaci, the causal agent of tobacco wildfire disease, as the target and found that 1.2 μg/mL biosynthesized AgNPs completely inhibited the growth of P. syringae pv. tabaci in vitro and in vivo by partly destroying the cell membrane structure of the pathogen, resulting in cytoplasmic leakage. Moreover, Nicotiana benthamiana treated with 1.2 μg/mL biosynthesized AgNPs exhibited a significant upregulation of nonexpressor of pathogenesis-related genes 1 (NPR1) and pathogenesis-related gene 2 (PR2), the typical markers of the salicylic acid (SA)-mediated defense system, and an increase in peroxidase (POD) and polyphenol oxidase (PPO) activities as well as the production of reactive oxygen species (ROS). Furthermore, biosynthesized AgNPs treatment increased the chlorophyll content and dry weight of N. benthamiana. Overall, we demonstrated that biosynthesized AgNPs at a low concentration have high inhibitory effect on the pathogen causing tobacco wildfire disease by destroying bacterial cell membrane and inducing defense resistance in host plant. These results lay a theoretical foundation for further application of biosynthesized AgNPs in the control of plant bacterial diseases.

Physcomitrium patens CAD1 has distinct roles in growth and resistance to biotic stress

BackgroundPhyscomitrium patens provides an evolutionary link between green algae and vascular plants. Although the genome of P. patens includes orthologs of all the core lignin biosynthetic enzymes, the occurrence of lignin in moss is very controversial. Besides, little information is available about the lignin enzymes in moss to date. For example, cinnamyl alcohol dehydrogenase (CAD) is a crucial enzyme that catalyzes the last step of the lignin biosynthetic pathway, suggesting an ideal way to study the evolutionary process. By investigating the functions of CAD in evolution, this study will elucidate the evolutionary roles of lignin-like in the early stage of land colonization.ResultsCAD multigene family in P. patens is composed of four genes. The PpCADs contain a conserved glycine-rich domain to catalyze NADPH-dependent reduction to their corresponding alcohols, indicating that PpCADs have the potential to synthesize monolignols by bioinformatics analysis. Even though PpCAD1 could produce lignin in theory, no conventional monomer was detected in the cell wall or cytoplasm of PpCAD1_OE plants. However, the phenylpropanoids were promoted in PpCAD1_OE transformants to modify gametophore architecture and development, making the distribution of phyllids more scarcity and the moss colony more giant, possibly due to the enhanced expression of the AUX-IAA family. The transcripts of at least one gene encoding the enzyme in the lignin biosynthetic pathway were increased in PpCAD1_OE plants. In addition, the PpCAD1_OE gametophore inhibited the Botrytis cinerea assault mainly by enhanced phenylpropanoids in the cell wall instead of influencing transcripts of defense genes pathogenesis-related 10 (PR10) and nonexpresser of PR genes 1 (NPR1). Likewise, ectopic expression of PpCAD1 in Arabidopsis led to a significant increase in lignin content, exhibiting chunky roots, robust seedlings, advanced flowering, and efficient resistance against pathogens.ConclusionPpCAD occurs in more than one copy, suggesting functional divergence in the ancestral plant. PpCAD1 catalyzes monolignol biosynthesis and has homologous functions with vascular plants. Despite no detected conventional monolignol, the increased phenylpropanoids in the PpCAD1_OE gametophore, possibly intermediate metabolites in the lignin pathway, had conserved functions during the evolution of terrestrial plants. The results inferred that the lignin enzyme of the early non-vascular plant played roles in stem elongation and resistance against pathogens of P. patens during the conquest of land.

An activated form of NB-ARC protein RLS1 functions with cysteine-rich receptor-like protein RMC to trigger cell death in rice

A key event that follows pathogen recognition by a resistance (R) protein containing an NB-ARC (nucleotide-binding adaptor shared by Apaf-1, R proteins, and Ced-4) domain is hypersensitive response (HR)-type cell death accompanied by accumulation of reactive oxygen species and nitric oxide. However, the integral mechanisms that underlie this process remain relatively opaque. Here, we show that a gain-of-function mutation in the NB-ARC protein RLS1 (Rapid Leaf Senescence 1) triggers high-light-dependent HR-like cell death in rice. The RLS1-mediated defense response is largely independent of salicylic acid accumulation, NPR1 (Nonexpressor of Pathogenesis-Related Gene 1) activity, and RAR1 (Required for Mla12 Resistance 1) function. A screen for suppressors of RLS1 activation identified RMC (Root Meander Curling) as essential for the RLS1-activated defense response. RMC encodes a cysteine-rich receptor-like secreted protein (CRRSP) and functions as an RLS1-binding partner. Intriguingly, their co-expression resulted in a change in the pattern of subcellular localization and was sufficient to trigger cell death accompanied by a decrease in the activity of the antioxidant enzyme APX1. Collectively, our findings reveal an NB-ARC–CRRSP signaling module that modulates oxidative state, the cell death process, and associated immunity responses in rice.

Ammonium and nitric oxide condition the establishment of Arabidopsis Ler/Kas-2 immune-related hybrid incompatibility

Main ConclusionHigh ammonium suppresses hybrid incompatibility between Ler and Kas-2 accessions through lowering nitric oxide levels and nitrate reductase activity required for autoimmunity.The immune-related hybrid incompatibility (HI) between Landsberg erecta (Ler) and Kashmir-2 (Kas-2) accessions is due to a deleterious genetic interaction between the RPP1 (RECOGNITION OF PERONOSPORA PARASITICA1)-like Ler locus and Kas-2 alleles of the receptor-like kinase SRF3 (STRUBBELIG RECEPTOR FAMILY 3). The genetic incompatibility is temperature-dependent and leads to constitutive activation of the salicylic acid (SA) pathway, dwarfism and cell death at 14–16 °C. Here we investigated the effect of nutrition on the occurrence of Ler/Kas-2 HI and found that high ammonium suppresses Ler/Kas-2 incompatible phenotypes independently of the ammonium/nitrate ratio. Ammonium feeding leads to compromised disease resistance to Pseudomonas syringae pv. tomato DC3000, lower total SA, nitric oxide and nitrate reductase activity in Ler/Kas-2 incompatible hybrids. In addition, we find that Ler/Kas-2 incompatibility is dependent on NPR1 (NONEXPRESSER OF PR GENES 1) and nitric oxide production. Overall, this work highlights the effect of nutrition on the expression of incompatible phenotypes independently of temperature.

Salicylic acid-activated BIN2 phosphorylation of TGA3 promotes ArabidopsisPR gene expression and disease resistance.

The plant defense hormone, salicylic acid (SA), plays essential roles in immunity and systemic acquired resistance. Salicylic acid induced by the pathogen is perceived by the receptor nonexpressor of pathogenesis-related genes 1 (NPR1), which is recruited by TGA transcription factors to induce the expression of pathogenesis-related (PR) genes. However, the mechanism by which post-translational modifications affect TGA's transcriptional activity by salicylic acid signaling/pathogen infection is not well-established. Here, we report that the loss-of-function mutant of brassinosteroid insensitive2 (BIN2) and its homologs, bin2-3 bil1 bil2, causes impaired pathogen resistance and insensitivity to SA-induced PR gene expression, whereas the gain-of-function mutant, bin2-1, exhibited enhanced SA signaling and immunity against the pathogen. Our results demonstrate that salicylic acid activates BIN2 kinase, which in turn phosphorylates TGA3 at Ser33 to enhance TGA3 DNA binding ability and NPR1-TGA3 complex formation, leading to the activation of PR gene expression. These findings implicate BIN2 as a new component of salicylic acid signaling, functioning as a key node in balancing brassinosteroid-mediated plant growth and SA-induced immunity.