Pst Infection Research Articles

Modulation of chloroplast components and defense responses during programmed cell death in tobacco infected with Pseudomonas syringae

We examined how tobacco plants coordinate chloroplast components and defense responses during Pseudomonas syringae pv. tomato (Pst) infection. Tobacco leaves infiltrated with Pst induced weak necrosis at 24 h post-infiltration (hpi) and severe necrosis at 48 hpi. Membrane damage, as shown by cellular leakage and malondialdehyde, and H2O2 production began to increase at 12 hpi and continuously increased at 24–72 hpi in Pst-infiltrated leaves. Pst infection resulted in decreases in light-harvesting chlorophyll-binding proteins (Lhc), Lhcb transcripts, electron transport rate, and Fv/Fm, indicating the impairment in structure and function of photosystem II. Photochemical quenching, qP, continuously decreased in Pst-infiltrated leaves at 24–48 hpi, whereas nonphotochemical quenching, NPQ, exhibited a 2-fold increase at 24 hpi and a decrease at 48 dpi. In response to Pst infection, chlorophyll began to decrease at 48 hpi, whereas levels of protoporphyrin IX (Proto IX), Mg-Proto IX, Mg-Proto methylester, and protochlorophyllide drastically decreased or disappeared as early as 24 hpi. Pst-infiltrated leaves greatly up-regulated the expression of ROS scavenging genes, Fe-SOD, APX, and CAT1, as well as defense-related genes, PII, PR1, PR2, PALa, and CHS1. Our study suggests that the modulation of photosynthetic components during pathogen infection, particularly in relation to the fast degradation of photosensitizing porphyrin intermediates and the increase in photoprotective NPQ, may contribute to attenuating cellular damage in the early stages of programmed cell death induced by Pst.

Genome-Wide Mapping of Quantitative Trait Loci Conferring All-Stage and High-Temperature Adult-Plant Resistance to Stripe Rust in Spring Wheat Landrace PI 181410.

Stripe rust, caused by Puccinia striiformis f. sp. tritici (Pst), is one of the most destructive diseases of wheat in the world. Genetic resistance is the best strategy for control of the disease. Spring wheat landrace PI 181410 has shown high level resistance to stripe rust. The present study characterized the landrace to have both race-specific all-stage resistance and nonrace-specific high-temperature adult-plant (HTAP) resistance. To map quantitative trait loci (QTL) for the resistance in PI 181410, it was crossed with Avocet S (AvS), from which a recombinant inbred line population was developed. The F5–F8 populations were consecutively phenotyped for stripe rust response in multiple field environments under natural Pst infection, and the F7 population was phenotyped in seedlings at low temperature and in adult-plant stage with selected Pst races in the greenhouse. The F7 population was genotyped using the 90K wheat SNP chip. Three QTL, QYrPI181410.wgp-4AS, QYrPI181410.wgp-4BL, and QYrPI181410.wgp-5BL.1, from PI 181410 for all-stage resistance, were mapped on chromosome arms 4AS, 4BL, and 5BL, respectively. Four QTL, QYrPI181410.wgp-1BL, QYrPI181410.wgp-4BL, QYrPI181410.wgp-5AS, and QYrPI181410.wgp-5BL.2, were identified from PI 181410 for HTAP resistance and mapped to 1BL, 4BL, 5AS, and 5BL, respectively. Two QTL with minor effects on stripe rust response were identified from AvS and mapped to 2BS and 2BL. Four of the QTL from PI 181410 and one from AvS were potentially new. As the 4BL QTL was most effective and likely a new gene for stripe rust resistance, three kompetitive allele specific PCR (KASP) markers were developed for incorporating this gene into new wheat cultivars.

Effects of Stripe Rust Infection on the Levels of Redox Balance and Photosynthetic Capacities in Wheat.

Wheat stripe rust (Puccinia striiformis f. sp. tritici, Pst) is the most destructive wheat disease and a major problem for the productivity of wheat in the world. To obtain a better understanding about different effects of redox homeostasis and photosystem (PS) to Pst infection in wheat, we investigated the differences in photosynthesis and the antioxidant defense system in wheat cultivar Chuanmai42 (CM42) in response to two Chinese Pst races known as CYR32 and V26. The results showed that V26-infected wheat accumulated a higher reactive oxygen species (ROS), cell death, and energy dissipation than CYR32-infected wheat when compared with the control. Furthermore, we found that the activities of three antioxidant enzymes (APX, GR, and GPX) and four resistance-related enzymes in CYR32-infected wheat were significantly higher than that in V26-infected wheat. In addition, quantitative RT-PCR indicated that the expression levels of two genes associated with resistant stripe rust in CYR32-infected wheat were clearly higher than that in V26-infected wheat. Compared with CYR32-infected wheat, lower photochemical efficiencies were observed in V26-infected wheat at the adult stage. Meanwhile, only a marked decline in D1 protein was observed in V26-infected wheat. We therefore deduced that wheat with stripe rust resistance could maintain high resistance and photosynthetic capacity by regulating the antioxidant system, disease-resistant related enzymes and genes, and the levels of PSII reaction center proteins.

An effector protein of the wheat stripe rust fungus targets chloroplasts and suppresses chloroplast function

Chloroplasts are important for photosynthesis and for plant immunity against microbial pathogens. Here we identify a haustorium-specific protein (Pst_12806) from the wheat stripe rust fungus, Puccinia striiformis f. sp. tritici (Pst), that is translocated into chloroplasts and affects chloroplast function. Transient expression of Pst_12806 inhibits BAX-induced cell death in tobacco plants and reduces Pseudomonas-induced hypersensitive response in wheat. It suppresses plant basal immunity by reducing callose deposition and the expression of defense-related genes. Pst_12806 is upregulated during infection, and its knockdown (by host-induced gene silencing) reduces Pst growth and development, likely due to increased ROS accumulation. Pst_12806 interacts with the C-terminal Rieske domain of the wheat TaISP protein (a putative component of the cytochrome b6-f complex). Expression of Pst_12806 in plants reduces electron transport rate, photosynthesis, and production of chloroplast-derived ROS. Silencing TaISP by virus-induced gene silencing in a susceptible wheat cultivar reduces fungal growth and uredinium development, suggesting an increase in resistance against Pst infection.

TabZIP74 Acts as a Positive Regulator in Wheat Stripe Rust Resistance and Involves Root Development by mRNA Splicing.

Basic leucine zipper (bZIP) membrane-bound transcription factors (MTFs) play important roles in regulating plant growth and development, abiotic stress responses, and disease resistance. Most bZIP MTFs are key components of signaling pathways in endoplasmic reticulum (ER) stress responses. In this study, a full-length cDNA sequence encoding bZIP MTF, designated TabZIP74, was isolated from a cDNA library of wheat near-isogenic lines of Taichung29*6/Yr10 inoculated with an incompatible race CYR32 of Puccinia striiformis f. sp. tritici (Pst). Phylogenic analysis showed that TabZIP74 is highly homologous to ZmbZIP60 in maize and OsbZIP74 in rice. The mRNA of TabZIP74 was predicted to form a secondary structure with two kissing hairpin loops that could be spliced, causing an open reading frame shift immediately before the hydrophobic region to produce a new TabZIP74 protein without the transmembrane domain. Pst infection and the abiotic polyethylene glycol (PEG) and abscisic acid (ABA) treatments lead to TabZIP74 mRNA splicing in wheat seedling leaves, while both spliced and unspliced forms in roots were detected. In the confocal microscopic examination, TabZIP74 is mobilized in the nucleus from the membrane of tobacco epidermal cells in response to wounding. Knocking down TabZIP74 with barley stripe mosaic virus-induced gene silencing (BSMV-VIGS) enhanced wheat seedling susceptibility to stripe rust and decreased drought tolerance and lateral roots of silenced plants. These findings demonstrate that TabZIP74 mRNA is induced to splice when stressed by biotic and abiotic factors, acts as a critically positive regulator for wheat stripe rust resistance and drought tolerance, and is necessary for lateral root development.

Arabidopsis exhibits differential response in basal immunity and proline metabolism during defense against host and nonhost pathogen infection

A host-pathogen surpasses the plant defense machinery and successfully infects the plant to serve its needs. In contrast, a nonhost pathogen is restricted by plant immune responses. In this study, we deciphered the differential responses of Arabidopsis against the host-pathogen (Pseudomonas syringae pv. maculicola, Psm) and the nonhost pathogen (P. syringae pv. tabaci, Pst) infection. The Pst multiplication was restricted in Arabidopsis plant coinciding with the absence of any disease symptoms which was also associated with the increase in defense associated, pathogenesis related gene1 (PR1) expression and callose deposition. Host-pathogen infection, on the other hand, caused chlorotic symptoms with much less activation of the defense marker genes compared to the nonhost infection. Proline content was decreased in plants infected with Psm but not in case of Pst infection. Proline is a crucial plant metabolite, and osmolyte suggested to be involved in plant defense responses. However, the differential regulation of proline pathway under a susceptible or resistance response has not yet been unravelled. The expression profile of proline metabolism genes in a time course post-infection revealed drastic differences after Psm and Pst pathogen infection. The elevated expression of proline catabolic genes, AtProDH and AtP5CDH were noted under Psm infection. In contrast, plants infected with Pst showed upregulation of AtProDH but a decline in AtP5CDH transcripts along with an upregulation of the biosynthesis genes i.e. AtP5CS and AtP5CR. Our study shows that proline metabolism is tightly regulated under host and nonhost pathogen infection and impacts susceptibility and resistance of the plants, respectively.

A stripe rust effector Pst18363 targets and stabilises TaNUDX23 that promotes stripe rust disease.

Wheat stripe rust, caused by Puccinia striiformis f. sp. tritici (Pst), poses a tremendous threat to the production of wheat worldwide. The molecular mechanisms of Pst effectors that regulate wheat immunity are poorly understood. In this study, we identified an effector Pst18363 from Pst that suppresses plant cell death in Nicotiana benthamiana and in wheat. Knocking down Pst18363 expression by virus-mediated host-induced gene silencing significantly decreased the number of rust pustules, indicating that Pst18363 functions as an important pathogenicity factor in Pst. Pst18363 was proven to interact with wheat Nudix hydrolase 23 TaNUDX23. In wheat, silencing of TaNUDX23 by virus-induced gene silencing increased reactive oxygen species (ROS) accumulation induced by the avirulent Pst race CYR23, whereas overexpression of TaNUDX23 suppressed ROS accumulation induced by flg22 in Arabidopsis. In addition, TaNUDX23 suppressed Pst candidate effector Pst322-trigged cell death by decreasing ROS accumulation in N.benthamiana. Knocking down of TaNUDX23 expression attenuated Pst infection, indicating that TaNUDX23 is a negative regulator of defence. In N.benthamiana, Pst18363 stabilises TaNUDX23. Overall, our data suggest that Pst18363 stabilises TaNUDX23, which suppresses ROS accumulation to facilitate Pst infection.

ABA-Induced Sugar Transporter TaSTP6 Promotes Wheat Susceptibility to Stripe Rust.

Biotrophic pathogens, such as wheat rust fungi, survive on nutrients derived from host cells. Sugar appears to be the major carbon source transferred from host cells to various fungal pathogens; however, the molecular mechanism by which host sugar transporters are manipulated by fungal pathogens for nutrient uptake is poorly understood. TaSTP6, a sugar transporter protein in wheat (Triticum aestivum), was previously shown to exhibit enhanced expression in leaves upon infection by Puccinia striiformis f. sp. tritici (Pst), the causal agent of wheat stripe rust. In this study, we found that Pst infection caused increased accumulation of abscisic acid (ABA) and that application of exogenous ABA significantly enhanced TaSTP6 expression. Moreover, knockdown of TaSTP6 expression by barley stripe mosaic virus-induced gene silencing reduced wheat susceptibility to the Pst pathotype CYR31, suggesting that TaSTP6 expression upregulation contributes to Pst host sugar acquisition. Consistent with this, TaSTP6 overexpression in Arabidopsis (Arabidopsis thaliana) promoted plant susceptibility to powdery mildew and led to increased Glc accumulation in the leaves. Functional complementation assays in Saccharomyces cerevisiae showed that TaSTP6 has broad substrate specificity, indicating that TaSTP6 is an active sugar transporter. Subcellular localization analysis indicated that TaSTP6 localizes to the plasma membrane. Yeast two-hybrid and bimolecular fluorescence complementation experiments revealed that TaSTP6 undergoes oligomerization. Taken together, our results suggest that Pst stimulates ABA biosynthesis in host cells and thereby upregulates TaSTP6 expression, which increases sugar supply and promotes fungal infection.

Ethylene Response Factor ERF11 Activates BT4 Transcription to Regulate Immunity to Pseudomonas syringae.

Pseudomonas syringae, a major hemibiotrophic bacterial pathogen, causes many devastating plant diseases. However, the transcriptional regulation of plant defense responses to P. syringae remains largely unknown. Here, we found that gain-of-function of BTB AND TAZ DOMAIN PROTEIN 4 (BT4) enhanced the resistance of Arabidopsis (Arabidopsis thaliana) to Pst DC3000 (Pseudomonas syringae pv. tomato DC3000). Disruption of BT4 also weakened the salicylic acid (SA)-induced defense response to Pst DC3000 in bt4 mutants. Further investigation indicated that, under Pst infection, transcription of BT4 is modulated by components of both the SA and ethylene (ET) signaling pathways. Intriguingly, the specific binding elements of ETHYLENE RESPONSE FACTOR (ERF) proteins, including dehydration responsive/C-repeat elements and the GCC box, were found in the putative promoter of BT4 Based on publicly available microarray data and transcriptional confirmation, we determined that ERF11 is inducible by salicylic acid and Pst DC3000 and is modulated by the SA and ET signaling pathways. Consistent with the function of BT4, loss-of-function of ERF11 weakened Arabidopsis resistance to Pst DC3000 and the SA-induced defense response. Biochemical and molecular assays revealed that ERF11 binds specifically to the GCC box of the BT4 promoter to activate its transcription. Genetic studies further revealed that the BT4-regulated Arabidopsis defense response to Pst DC3000 functions directly downstream of ERF11. Our findings indicate that transcriptional activation of BT4 by ERF11 is a key step in SA/ET-regulated plant resistance against Pst DC3000, enhancing our understanding of plant defense responses to hemibiotrophic bacterial pathogens.

Development, Validation, and Re-selection of Wheat Lines with Pyramided Genes Yr64 and Yr15 Linked on the Short Arm of Chromosome 1B for Resistance to Stripe Rust.

Stripe rust, caused by Puccinia striiformis f. sp. tritici (Pst), is one of the most destructive diseases of wheat worldwide. The disease is most preferably managed by developing and growing cultivars with high-level, durable resistance. To achieve high-level and long-lasting resistance, we developed a wheat line, RIL-Yr64/Yr15, by pyramiding Yr64 and Yr15, both on the chromosome 1BS and providing high resistance to all tested Pst races. To validate RIL-Yr64/Yr15 possessing both genes, we crossed it to Avocet S (AvS). The F4 RILs from this cross were phenotyped with Pst races under controlled greenhouse conditions and also under natural Pst infection in the field. The population was genotyped with SSR markers previously reported to be linked to the resistance gene loci and with additional SSR and SNP-KASP markers along chromosome 1B. Both phenotype and genotype data confirmed the copresence of Yr64 and Yr15 in RIL-Yr64/Yr15, and the high-resolution linkage map dissected the chromosomal regions and traced their origins. New lines possessing these genes were selected from the F5 population of cross AvS × RIL-Yr64/Yr15 by marker-assisted selection. These lines with the two highly effective genes should be more useful than individual gene lines for developing high-level, durable resistant wheat cultivars.

A novel citrate synthase isoform contributes infection and stress resistance of the stripe rust fungus.

The early development of a rust fungus is dependent on the endogenous lipids stored in the urediniospores. After it establishes a parasitic relationship with the host, sugars absorbed from the host cells by haustoria become the primary nutrients. The tricarboxylic acid (TCA) cycle is essential to oxidize these nutrients. However, few studies have addressed the role of citrate synthase (CS), a rate-limiting enzyme of the TCA cycle, during the infection process of rust fungi. In this study, a CS gene from Puccinia striiformis f. sp. tritici (Pst), PsCS1, was cloned and characterized. Transcripts of PsCS1 and the enzyme activity of the CS were increased in the early Pst infection stage. Biochemical features and subcellular localization revealed that PsCS1 encoded a mitochondrial CS. Size exclusion chromatography, yeast two-hybrid and bimolecular fluorescence complementation experiments confirmed that PsCS1 could form a functional homo-octamer. The overexpression of PsCS1 enhanced the resistance of Escherichia coli to salt stress. The knockdown of PsCS1 using a host-induced gene silencing (HIGS) system blocked Pst growth in wheat. These results indicate that PsCS1 is required for nutrient metabolism in Pst and contributes to Pst infection by regulating ATP production and the supply of carbon sources.

Induced Resistance Against Western Flower Thrips by the Pseudomonas syringae-Derived Defense Elicitors in Tomato.

Western flower thrips (WFT) Frankliniella occidentalis (Pergande) is a key agricultural pest of cultivated tomatoes. Induced host plant resistance by activating jasmonic acid (JA) signaling pathway constitutes a promising method for WFT control. The phytotoxin coronatine (COR), produced by Pseudomonas syringae pv. tomato DC3000 (Pst), mimics the plant hormone JA-Isoleucine and can promote resistance against herbivorous arthropods. Here we determined the effect of Pst and COR on tomato resistance against WFT, induction of JA and salicylic acid (SA) associated defenses, and plant chemistry. Additionally, we investigated the presence of other components in Pst-derived and filtered culture medium, and their interactive effect with COR on tomato resistance to WFT. Our results showed that infiltration of COR or Pst reduced WFT feeding damage in tomato plants. COR and Pst induced the expression of JA-associated gene and protein marker. COR also induced expression of a SA-related responsive gene, although at much less magnitude. Activation of JA defenses in COR and Pst infiltrated plants did not affect density of type VI leaf trichomes, which are defenses reported to be induced by JA. An untargeted metabolomic analysis showed that both treatments induced strong changes in infiltrated leaves, but leaf responses to COR or Pst slightly differed. Application of the Pst-derived and filtered culture medium, containing COR but not viable Pst, also increased tomato resistance against WFT confirming that the induction of tomato defenses does not require a living Pst population to be present in the plant. Infiltration of tomato plants with low concentrations of COR in diluted Pst-derived and filtered culture medium reduced WFT feeding damage in a greater magnitude than infiltration with an equivalent amount of pure COR indicating that other elicitors are present in the medium. This was confirmed by the fact that the medium from a COR-mutant of Pst also strongly reduced silver damage. In conclusion, our results indicate that induction of JA defenses by COR, Pst infection, the medium of Pst and the medium of a Pst COR- mutant increased resistance against WFT. This was not mediated by the reinforcement of leaf trichome densities, but rather the induction of chemical defenses.

Root exudates drive the soil-borne legacy of aboveground pathogen infection

BackgroundPlants are capable of building up beneficial rhizosphere communities as is evidenced by disease-suppressive soils. However, it is not known how and why soil bacterial communities are impacted by plant exposure to foliar pathogens and if such responses might improve plant performance in the presence of the pathogen. Here, we conditioned soil by growing multiple generations (five) of Arabidopsis thaliana inoculated aboveground with Pseudomonas syringae pv tomato (Pst) in the same soil. We then examined rhizosphere communities and plant performance in a subsequent generation (sixth) grown in pathogen-conditioned versus control-conditioned soil. Moreover, we assessed the role of altered root exudation profiles in shaping the root microbiome of infected plants.ResultsPlants grown in conditioned soil showed increased levels of jasmonic acid and improved disease resistance. Illumina Miseq 16S rRNA gene tag sequencing revealed that both rhizosphere and bulk soil bacterial communities were altered by Pst infection. Infected plants exhibited significantly higher exudation of amino acids, nucleotides, and long-chain organic acids (LCOAs) (C > 6) and lower exudation levels for sugars, alcohols, and short-chain organic acids (SCOAs) (C ≤ 6). Interestingly, addition of exogenous amino acids and LCOA also elicited a disease-suppressive response.ConclusionCollectively, our data suggest that plants can recruit beneficial rhizosphere communities via modification of plant exudation patterns in response to exposure to aboveground pathogens to the benefit of subsequent plant generations.

The calcium sensor TaCBL4 and its interacting protein TaCIPK5 are required for wheat resistance to stripe rust fungus.

Calcineurin B-like proteins (CBLs) act as Ca2+ sensors to activate specific protein kinases, namely CBL-interacting protein kinases (CIPKs). Recent research has demonstrated that the CBL-CIPK complex is not only required for abiotic stress signaling, but is also probably involved in biotic stress perception. However, the role of this complex in immune signaling, including pathogen perception, is unknown. In this study, we isolated one signaling component of the TaCBL-TaCIPK complex (TaCBL4-TaCIPK5) and characterized its role in the interaction between wheat (Triticum aestivum) and Puccinia striiformis f. sp. tritici (Pst, stripe rust fungus). Among all TaCBLs in wheat, TaCBL4 mRNA accumulation markedly increased after infection by Pst. Silencing of TaCBL4 resulted in enhanced susceptibility to avirulent Pst infection. In addition, screening determined that TaCIPK5 physically interacted with TaCBL4 in planta and positively contributed to wheat resistance to Pst. Moreover, the disease resistance phenotype of TaCBL4 and TaCIPK5 co-silenced plants was consistent with that of single-knockdown plants. The accumulation of reactive oxygen species (ROS) was significantly altered in all silenced plants during Pst infection. Together these findings demonstrate that the TaCBL4-TaCIPK5 complex positively modulates wheat resistance in a ROS-dependent manner, and provide new insights into the roles of CBL-CIPK in wheat.

Bacteria Exploit Autophagy for Proteasome Degradation and Enhanced Virulence in Plants.

Autophagy and the ubiquitin-proteasome system (UPS) are two major protein degradation pathways implicated in the response to microbial infections in eukaryotes. In animals, the contribution of autophagy and the UPS to antibacterial immunity is well documented and several bacteria have evolved measures to target and exploit these systems to the benefit of infection. In plants, the UPS has been established as a hub for immune responses and is targeted by bacteria to enhance virulence. However, the role of autophagy during plant-bacterial interactions is less understood. Here, we have identified both pro- and antibacterial functions of autophagy mechanisms upon infection of Arabidopsis thaliana with virulent Pseudomonas syringae pv tomato DC3000 (Pst). We show that Pst activates autophagy in a type III effector (T3E)-dependent manner and stimulates the autophagic removal of proteasomes (proteaphagy) to support bacterial proliferation. We further identify the T3E Hrp outer protein M1 (HopM1) as a principle mediator of autophagy-inducing activities during infection. In contrast to the probacterial effects of Pst-induced proteaphagy, NEIGHBOR OF BRCA1-dependent selective autophagy counteracts disease progression and limits the formation of HopM1-mediated water-soaked lesions. Together, we demonstrate that distinct autophagy pathways contribute to host immunity and bacterial pathogenesis during Pst infection and provide evidence for an intimate crosstalk between proteasome and autophagy in plant-bacterial interactions.

Host-Induced Gene Silencing of the MAPKK Gene PsFUZ7 Confers Stable Resistance to Wheat Stripe Rust.

RNA interference (RNAi) is a powerful genetic tool to accelerate research in plant biotechnology and control biotic stresses by manipulating target gene expression. However, the potential of RNAi in wheat to efficiently and durably control the devastating stripe rust fungus Puccinia striiformis f. sp. tritici (Pst) remained largely under explored so far. To address this issue, we generated transgenic wheat (Triticum aestivum) lines expressing dsRNA targeting PsFUZ7 transcripts of Pst We analyzed expression of PsFUZ7 and related genes, and resistance traits of the transgenic wheat lines. We show that PsFUZ7 is an important pathogenicity factor that regulates infection and development of Pst A PsFUZ7 RNAi construct stably expressed in two independent transgenic wheat lines confers strong resistance to PstPst hyphal development is strongly restricted, and necrosis of plant cells in resistance responses was significantly induced. We conclude that trafficking of RNA molecules from wheat plants to Pst may lead to a complex molecular dialogue between wheat and the rust pathogen. Moreover, we confirm the RNAi-based crop protection approaches can be used, to our knowledge, as a novel control strategy against rust pathogens in wheat.

Different Arabidopsis thaliana photosynthetic and defense responses to hemibiotrophic pathogen induced by local or distal inoculation of Burkholderia phytofirmans.

Pathogen infection of plant results in modification of photosynthesis and defense mechanisms. Beneficial microorganisms are known to improve plant tolerance to stresses. Burkholderia phytofirmans PsJN (Bp), a beneficial endophytic bacterium, promotes growth of a wide range of plants and induces plant resistance against abiotic and biotic stresses such as coldness and infection by a necrotrophic pathogen. However, mechanisms underlying its role in plant tolerance towards (hemi)biotrophic invaders is still lacking. We thus decipher photosynthetic and defense responses during the interaction between Arabidopsis, Bp and the hemibiotrophic bacterium Pseudomonas syringae pv. tomato DC3000 (Pst). Different Bp inoculations allowed analyzes at both systemic and local levels. Despite no direct antibacterial action, our results showed that only local presence of Bp alleviates Pst growth in planta during the early stage of infection. Molecular investigations showed that seed inoculation of Bp, leading to a restricted presence in the root system, transiently primed PR1 expression after challenge with Pst but continuously primed PDF1.2 expression. Bacterization with Bp reduced Y(ND) but had no impact on PSII activity or RuBisCO accumulation. Pst infection caused an increase of Y(NA) and a decrease in ΦPSI, ETRI and in PSII activity, showed by a decrease in Fv/Fm, Y(NPQ), ΦPSII, and ETRII values. Inoculation with both bacteria did not display any variation in photosynthetic activity compared to plants inoculated with only Pst. Our findings indicated that the role of Bp here is not multifaceted, and relies only on priming of defense mechanisms but not on improving photosynthetic activity.

A unique invertase is important for sugar absorption of an obligate biotrophic pathogen during infection.

An increased invertase activity in infected plant tissue has been observed in many plant-pathogen interactions. However, the origin of this increased invertase activity (plant and/or pathogen) is still under debate. In addition, the role of pathogen invertases in the infection process is also unclear. We identified and cloned a gene with homology to invertases from Puccinia striiformis f. sp. tritici (Pst). Transcript levels of PsINV were analyzed by quantitative reverse transcription PCR in both compatible and incompatible Pst-wheat interactions . Function of the gene product was confirmed by heterologous expression, and its function in Pst infection was analyzed by host-induced gene silencing (HIGS). Pst abundantly secretes invertase during its invasion attempts whether in a compatible or incompatible interaction with wheat. Further research into the different domains of this protein indicated that the rust-specific sequence contributes to a higher efficiency of sucrose hydrolysis. With PsINV silenced by HIGS during the infection process, growth of Pst is inhibited and conidial fructification incomplete. Finally, pathogenicity of Pst is impaired and spore yield significantly reduced. Our results clearly demonstrate that this Pst invertase plays a pivotal role in this plant-pathogen interaction probably by boosting sucrose hydrolysis to secure the pathogen's sugar absorption.

Bacterial and plant natriuretic peptides improve plant defence responses against pathogens.

Plant natriuretic peptides (PNPs) have been implicated in the regulation of ions and water homeostasis, and their participation in the plant immune response has also been proposed. Xanthomonas citri ssp. citri contains a gene encoding a PNP-like protein (XacPNP) which has no homologues in other bacteria. XacPNP mimics its Arabidopsis thaliana homologue AtPNP-A by modifying host responses to create favourable conditions for pathogen survival. However, the ability of XacPNP to induce plant defence responses has not been investigated. In order to study further the role of XacPNP in vivo, A. thaliana lines over-expressing XacPNP, lines over-expressing AtPNP-A and AtPNP-A-deficient plants were generated. Plants over-expressing XacPNP or AtPNP-A showed larger stomatal aperture and were more resistant to saline or oxidative stress than were PNP-deficient lines. In order to study further the role of PNP in biotic stress responses, A. thaliana leaves were infiltrated with pure recombinant XacPNP, and showed enhanced expression of genes related to the defence response and a higher resistance to pathogen infections. Moreover, AtPNP-A expression increased in A. thaliana on Pseudomonas syringae pv. tomato (Pst) infection. This evidence led us to analyse the responses of the transgenic plants to pathogens. Plants over-expressing XacPNP or AtPNP-A were more resistant to Pst infection than control plants, whereas PNP-deficient plants were more susceptible and showed a stronger hypersensitive response when challenged with non-host bacteria. Therefore, XacPNP, acquired by horizontal gene transfer, is able to mimic PNP functions, even with an increase in plant defence responses.

Characterization of a Ran gene from Puccinia striiformis f. sp. tritici involved in fungal growth and anti-cell death.

Ran, an important family of small GTP-binding proteins, has been shown to regulate a variety of important cellular processes in many eukaryotes. However, little is known about Ran function in pathogenic fungi. In this study, we report the identification and functional analysis of a Ran gene (designated PsRan) from Puccinia striiformis f. sp. tritici (Pst), an important fungal pathogen affecting wheat production worldwide. The PsRan protein contains all conserved domains of Ran GTPases and shares more than 70% identity with Ran proteins from other organisms, indicating that Ran proteins are conserved in different organisms. PsRan shows a low level of intra-species polymorphism and is localized to the nucleus. qRT-PCR analysis showed that transcript level of PsRan was induced in planta during Pst infection. Silencing of PsRan did not alter Pst virulence phenotype but impeded fungal growth of Pst. In addition, heterologous overexpression of PsRan in plant failed to induce cell death but suppressed cell death triggered by a mouse BAX gene or a Pst Ras gene. Our results suggest that PsRan is involved in the regulation of fungal growth and anti-cell death, which provides significant insight into Ran function in pathogenic fungi.