Haustorium Formation Research Articles

Histological and cytological characterization of adult plant resistance to wheat stripe rust

Wheat cultivar Xingzi 9104 (XZ) possesses adult plant resistance (APR) to stripe rust caused by Puccinia striiformis f. sp. tritici (Pst). In this study, histological and cytological experiments were conducted to elucidate the mechanisms of APR in XZ. The results of leaf inoculation experiments indicated that APR was initiated at the tillering stage, gradually increased as the plant aged and highly expressed after boot stage. The histology and oxidative burst in infected leaves of plants at seedling, tillering and boot stages were examined using light microscopic and histochemical methods. Subcellular changes in the host-pathogen interactions during the seedling and boot stages were analyzed by transmission electron microscopy. The results showed that haustorium formation was retarded in the adult plants and that the differentiation of secondary intercellular hyphae was significantly inhibited, which decreased the development of microcolonies in the adult plants, especially in plants of boot stage. The expression of APR to stipe rust during wheat development was clearly associated with extensive hypersensitive cell death of host cells and localized production of reactive oxygen species, which coincided with the restriction of fungal growth in infection sites in adult plants. At the same time, cell wall-related resistance in adult plants prevented ingression of haustorial mother cells into plant cells. Haustorium encasement was coincident with malformation or death of haustoria. The results provide useful information for further determination of mechanisms of wheat APR to stripe rust. The expression of APR to stipe rust in wheat cultivar Xingzi 9104 (XZ) was clearly associated with extensive hypersensitive cell death of host cells and the localized production of reactive oxygen species.

Interspecific RNA interference of SHOOT MERISTEMLESS-like disrupts Cuscuta pentagona plant parasitism.

Infection of crop species by parasitic plants is a major agricultural hindrance resulting in substantial crop losses worldwide. Parasitic plants establish vascular connections with the host plant via structures termed haustoria, which allow acquisition of water and nutrients, often to the detriment of the infected host. Despite the agricultural impact of parasitic plants, the molecular and developmental processes by which host/parasitic interactions are established are not well understood. Here, we examine the development and subsequent establishment of haustorial connections by the parasite dodder (Cuscuta pentagona) on tobacco (Nicotiana tabacum) plants. Formation of haustoria in dodder is accompanied by upregulation of dodder KNOTTED-like homeobox transcription factors, including SHOOT MERISTEMLESS-like (STM). We demonstrate interspecific silencing of a STM gene in dodder driven by a vascular-specific promoter in transgenic host plants and find that this silencing disrupts dodder growth. The reduced efficacy of dodder infection on STM RNA interference transgenics results from defects in haustorial connection, development, and establishment. Identification of transgene-specific small RNAs in the parasite, coupled with reduced parasite fecundity and increased growth of the infected host, demonstrates the efficacy of interspecific small RNA-mediated silencing of parasite genes. This technology has the potential to be an effective method of biological control of plant parasite infection.

Characterization of non-host resistance in broad bean to the wheat stripe rust pathogen.

BackgroundNon-host resistance (NHR) confers plant species immunity against the majority of microbial pathogens and represents the most robust and durable form of plant resistance in nature. As one of the main genera of rust fungi with economic and biological importance, Puccinia infects almost all cereals but is unable to cause diseases on legumes. Little is known about the mechanism of this kind of effective defense in legumes to these non-host pathogens.ResultsIn this study, the basis of NHR in broad bean (Vicia faba L.) against the wheat stripe rust pathogen, Puccinia striiformis f. sp. tritici (Pst), was characterized. No visible symptoms were observed on broad bean leaves inoculated with Pst. Microscopic observations showed that successful location of stomata and haustoria formation were significantly reduced in Pst infection of broad bean. Attempted infection induced the formation of papillae, cell wall thickening, production of reactive oxygen species, callose deposition and accumulation of phenolic compounds in plant cell walls. The few Pst haustoria that did form in broad bean cells were encased in reactive oxygen and callose materials and those cells elicited cell death. Furthermore, a total of seven defense-related genes were identified and found to be up-regulated during the Pst infection.ConclusionsThe results indicate that NHR in broad bean against Pst results from a continuum of layered defenses, including basic incompatibility, structural and chemical strengthening of cell wall, posthaustorial hypersensitive response and induction of several defense-related genes, demonstrating the multi-layered feature of NHR. This work also provides useful information for further determination of resistance mechanisms in broad bean to rust fungi, especially the adapted important broad bean rust pathogen, Uromyces viciae-fabae, because of strong similarity and association between NHR of plants to unadapted pathogens and basal resistance of plants to adapted pathogens.

Inoculation with arbuscular mycorrhizal fungi suppresses initiation of haustoria in the root hemiparasite Pedicularis tricolor.

Plant parasitism and arbuscular mycorrhizal (AM) associations have many parallels and share a number of regulatory pathways. Despite a rapid increase in investigations addressing the roles of AM fungi in regulating interactions between parasitic plants and their hosts, few studies have tested the effect of AM fungi on the initiation and differentiation of haustoria, the parasite-specific structures exclusively responsible for host attachment and nutrient transfer. In this study, we tested the influence of AM fungi on haustorium formation in a root hemiparasitic plant. Using a facultative root hemiparasitic species (Pedicularis tricolor) with the potential to form AM associations, the effects of inoculation were tested with two AM fungal species, Glomus mosseae and Glomus intraradices, on haustorium initiation in P. tricolor grown alone or with Hordeum vulgare 'Fleet' (barley) as the host plant. This study consisted of two greenhouse pot experiments. Both AM fungal species dramatically suppressed intraspecific haustorium initiation in P. tricolor at a very low colonization level. The suppression over-rode inductive effects of the parasite's host plant on haustoria production and caused significant growth depression of P. tricolor. AM fungi had strong and direct suppressive effects on haustorium formation in the root hemiparasite. The significant role of AM fungi in haustorium initiation of parasitic plants was demonstrated for the first time. This study provides new clues for the regulation of haustorium formation and a route to development of new biocontrol strategies in management of parasitic weeds.

Sugarcane genes differentially expressed in response to Puccinia melanocephala infection: identification and transcript profiling

Brown rust caused by the fungus Puccinia melanocephala is a major disease of sugarcane (Saccharum spp.). A sugarcane mutant, obtained by chemical mutagenesis of the susceptible variety B4362, showed a post-haustorial hypersensitive response (HR)-mediated resistance to the pathogen and was used to identify genes differentially expressed in response to P. melanocephala via suppression subtractive hybridization (SSH). Tester cDNA was derived from the brown rust-resistant mutant after inoculation with P. melanocephala, while driver cDNAs were obtained from the non-inoculated resistant mutant and the inoculated susceptible donor variety B4362. Database comparisons of the sequences of the SSH recombinant clones revealed that, of a subset of 89 non-redundant sequences, 88% had similarity to known functional genes, while 12% were of unknown function. Thirteen genes were selected for transcript profiling in the resistant mutant and the susceptible donor variety. Genes involved in glycolysis and C4 carbon fixation were up-regulated in both interactions probably due to disturbance of sugarcane carbon metabolism by the pathogen. Genes related with the nascent polypeptide associated complex, post-translational proteome modulation and autophagy were transcribed at higher levels in the compatible interaction. Up-regulation of a putative L-isoaspartyl O-methyltransferase S-adenosylmethionine gene in the compatible interaction may point to fungal manipulation of the cytoplasmatic methionine cycle. Genes coding for a putative no apical meristem protein, S-adenosylmethionine decarboxylase, non-specific lipid transfer protein, and GDP-L-galactose phosphorylase involved in ascorbic acid biosynthesis were up-regulated in the incompatible interaction at the onset of haustorium formation, and may contribute to the HR-mediated defense response in the rust-resistant mutant.

Concerted action of two avirulent spore effectors activates Reaction to Puccinia graminis 1 ( Rpg1 ) - mediated cereal stem rust resistance

The barley stem rust resistance gene Reaction to Puccinia graminis 1 (Rpg1), encoding a receptor-like kinase, confers durable resistance to the stem rust pathogen Puccinia graminis f. sp. tritici. The fungal urediniospores form adhesion structures with the leaf epidermal cells within 1 h of inoculation, followed by hyphae and haustorium formation. The RPG1 protein is constitutively expressed and not phosphorylated. On inoculation with avirulent urediniospores, it is phosphorylated in vivo within 5 min and subsequently degraded. Application of arginine-glycine-aspartic acid peptide loops prevented the formation of adhesion structures for spore attachment, the phosphorylation of RPG1, and germination of the viable spores. Arginine-glycine-aspartic acid affinity chromatography of proteins from the ungerminated avirulent rust spores led to the purification and identification of a protein with fibronectin type III and breast cancer type 1 susceptibility protein domains and a vacuolar protein sorting-associated protein 9 with a coupling of ubiquitin to endoplasmic reticulum degradation domain. Both proteins are required to induce in vivo phosphorylation and degradation of RPG1. Combined application of both proteins caused hypersensitive reaction on the stem rust-resistant cultivar Morex but not on the susceptible cultivar Steptoe. Expression studies indicated that mRNA of both genes are present in ungerminated urediniospores and are constitutively transcribed in sporelings, infected leaves, and haustoria in the investigated avirulent races. Evidence is presented that RPG1, in yeast, interacts with the two protein effectors from the urediniospores that activate cooperatively the stem rust resistance protein RPG1 long before haustoria formation.

Molecular characterization of a Fus3/Kss1 type MAPK from Puccinia striiformis f. sp. tritici, PsMAPK1.

Puccinia striiformis f. sp. tritici (Pst) is an obligate biotrophic fungus that causes the destructive wheat stripe rust disease worldwide. Due to the lack of reliable transformation and gene disruption method, knowledge about the function of Pst genes involved in pathogenesis is limited. Mitogen-activated protein kinase (MAPK) genes have been shown in a number of plant pathogenic fungi to play critical roles in regulating various infection processes. In the present study, we identified and characterized the first MAPK gene PsMAPK1 in Pst. Phylogenetic analysis indicated that PsMAPK1 is a YERK1 MAP kinase belonging to the Fus3/Kss1 class. Single nucleotide polymerphisms (SNPs) and insertion/deletion were detected in the coding region of PsMAPK1 among six Pst isolates. Real-time RT-PCR analyses revealed that PsMAPK1 expression was induced at early infection stages and peaked during haustorium formation. When expressed in Fusarium graminearum, PsMAPK1 partially rescued the map1 mutant in vegetative growth and pathogenicity. It also partially complemented the defects of the Magnaporthe oryzae pmk1 mutant in appressorium formation and plant infection. These results suggest that F. graminearum and M. oryzae can be used as surrogate systems for functional analysis of well-conserved Pst genes and PsMAPK1 may play a role in the regulation of plant penetration and infectious growth in Pst.

Why Biotrophs Can't Live Alone

Why Biotrophs Can't Live Alone

Suppression of Salicylic Acid-Mediated Plant Defense Responses During Initial Infection of Dyer's Woad by <i>Puccinia thlaspeos</i>

Problem statement: Puccinia thlaspeos is a microcyclic rust pathogen that is being investigated as a potential biocontrol agent of the noxious weed, dyer’s woad (Isatis tinctoria). Although, the initial events in the colonization of dyer’s woad by the rust pathogen has been elucidated using scanning electron microscopy and PCR, little is known regarding the susceptibility response of this plant to its rust pathogen. Approach: The induction kinetics and amplitude of the Salicylic Acid (SA)-responsive Pathogenesis-Related (PR) genes, PR-1, β-1, 3-glucanase and ChiA in the compatible interaction between the rust pathogen Puccinia thlaspeos and dyer’s woad were examined during the first 72 h of the infection process. Furthermore SA, an inducer of plant defense response was applied to infected plants in order to reprogram the host defense response at periods that coincided with key events of the infection process. Results: PR genes were upregulated following host penetration by the pathogen. A subsequent pathogen-mediated suppression of PR genes was seen that corresponded with haustorium formation. This was followed by a second up-regulation of these genes that was, in turn, followed by a second long-term pathogen-induced suppression of the defense response that appears to allow successful infections in dyer’s woad. Exogenous application of SA to uninoculated plants led to activation of defense responses by 8 h after treatment. In treatments where inoculated plants were treated with SA, responses differed depending on the timing of SA application. Application of SA at times corresponding to the pre-haustorial and posthaustorial phases of infection triggered an up-regulation of defense genes and increased protection against the pathogen. However, the application of SA during haustorium formation could not override the pathogen-mediated suppression of defense responses and consequently, did not offer the host increased protection. Conclusion: Although pathogen-induced PR expression was observed during rust infection of dyer’s woad, it appears to be insufficient to inhibit fungal growth during the compatible interaction between dyer’s woad and the rust pathogen. Suppression of pathogeninduced host defense responses during and after haustorium formation is postulated to be vital in the establishment of biotrophy in this system.

Transgressive segregation for very low and high levels of basal resistance to powdery mildew in barley

Basal resistance of barley to powdery mildew is a quantitatively inherited trait that limits the growth and sporulation of barley powdery mildew pathogen by a non-hypersensitive mechanism of defense. Two experimental barley lines were developed with a very high (ErBgh) and low (EsBgh) level of basal resistance to powdery mildew by cycles of convergent crossing and phenotypic selection between the most resistant and between the most susceptible lines, respectively, from four mapping populations of barley. Phenotypic selection in convergent crossing was highly effective in producing contrasting phenotypes for basal resistance and susceptibility. In ErBgh, almost 90% of infection units failed to form a primary haustorium in the epidermal cells in association with papilla formation, but in EsBgh only 33% of infection units failed to form a primary haustorium. The contrast between ErBgh and EsBgh for successful formation of secondary and subsequent haustoria was much less obvious (69% versus 79% successful secondary haustorium formation). In an earlier investigation, we determined seven QTLs for basal resistance in the four mapping populations. Checking the peak markers of these QTLs indicated that only four out of seven QTLs were confirmed to be present in the selected resistant lines and only four QTLs for susceptibility were confirmed to be present in the selected susceptible lines. Surprisingly, none of the expected QTLs could be detected in the resistant line ErBgh. We discuss some reasons why marker aided selection might be less efficient in raising levels of basal resistance than phenotypic selection. The very resistant and susceptible lines developed here are valuable material to be used in further experiments to characterize the molecular basis of basal resistance to powdery mildew.

Histological responses of host and non-host plants to Hyaloperonospora parasitica

Differences in Hyaloperonospora parasitica development and plant tissue responses were compared for 10 cruciferous hosts (including both resistant and susceptible genotypes), 3 leguminous and 1 graminaceous non-host species. Cotyledons, or true leaves in the case of Triticum aestivum and Pisum sativum, were studied at 2, 8, 24 h and 3, 5, 7 days post inoculation (dpi). The high levels of zoosporangial germination observed on all species tested, as well as on glass slides, suggested that inhibition of germination did not play a significant role in distinguishing host versus non-host resistance. During the early stages of infection, at spore germination and host penetration, there was no evidence of a clear-cut difference between Brassica host species which displayed a hypersensitive, partially resistant or susceptible reaction compared with non-host species. Haustoria formation was the key infection phase for the establishment of biotrophy. Across all tested species, haustoria were initiated inside the epidermal cells. However, there were significant differences in the frequency and timing of haustorial formation and the final size of haustoria among the tested species at early infection stage. Fully developed haustoria were never observed in Raphanus raphanistrum, Triticum aestivum, Lupinus angustifolius nor Trifolium subterraneum. Instead, the haustorium development appears to abort in the penetrated epidermal cells of these species. Although haustoria were formed in the epidermal and mesophyll cells of Sinapsis alba and Pisum sativum, subsequent hyphal growth and/or continued haustoria formation were rare or few, respectively. Hypersensitive reaction was the key resistance response observed among the host and non-host resistant species tested. It is noteworthy that, in the initial stages of pathogenesis, there was no differentiating point that separated the non-host species from those that were hosts.

Benzothiadiazole and BABA improve resistance to Uromyces pisi (Pers.) Wint. in Pisum sativum L. with an enhancement of enzymatic activities and total phenolic content

Benzothiadiazole (BTH) and DL-β-aminobutyric acid (BABA) induced systemic resistance was investigated in susceptible and resistant pea genotypes against Uromyces pisi. Resistance was characterized by reduced infection frequency mainly due to decreases in appressorium formation, stomatal penetration, growth of infection hyphae and haustorium formation. Changes in β-1,3-glucanase, chitinase, phenylalanine ammonia-lyase and peroxidase activities and in total phenolics content, demonstrate that U. pisi resistance is induced by BTH and BABA treatments at early and late stages of the fungal infection process, but that the chemicals operate via different mechanisms. In fact, our study showed that BTH treatment primed the activity of pathogenesis related-proteins such as β-1,3-glucanase, chitinase and peroxidase in both susceptible and resistant genotypes. On the other hand, BABA treatment did not increase the enzymatic activities in the studied genotypes, but significantly increased their total phenolic contents.

The immediate activation of defense responses in Arabidopsis roots is not sufficient to prevent Phytophthora parasitica infection

*The outcome of plant-microbe interactions is determined by a fine-tuned molecular interplay between the two partners. Little is currently known about the molecular dialogue between plant roots and filamentous pathogens. We describe here a new pathosystem for the analysis of molecular mechanisms occurring during the establishment of a compatible interaction between Arabidopsis thaliana roots and a root-infecting oomycete. *We performed cytological and genetic analyses of root infection during the compatible interaction between A. thaliana and Phytophthora parasitica. *Phytophthora parasitica uses appressoria to penetrate A. thaliana roots. Initial biotrophic growth is accompanied by the formation of haustoria, and is followed by a necrotrophic lifestyle. Arabidopsis thaliana mutants with impaired salicylic acid (SA), jasmonic acid (JA) or ethylene (ET) signaling pathways are more susceptible than the wild-type to infection. The salicylate- and jasmonate-dependent signaling pathways are concertedly activated when P. parasitica penetrates the roots, but are downregulated during invasive growth, when ethylene-mediated signaling predominates. *Thus, defense responses in A. thaliana roots are triggered immediately on contact with P. parasitica. Our findings suggest that the pattern of early defense mechanism activation differs between roots and leaves.

Parasitic plant infection is partially controlled through symbiotic pathways

Fernández‐Aparicio M, Rispail N, Prats E, Morandi D, García‐Garrido JM, Dumas‐Gaudot E, Duc G & Rubiales D (2009). Parasitic plant infection is partially controlled through symbiotic pathways. Weed Research.50, 76–82.SummaryLegumes are unique in interacting with Rhizobium, arbuscular mycorrhizal (AM) fungi, and parasitic plants. To dissect common parts of these three plant–organism interactions, infection by Orobanche crenata was studied in mutants with altered symbiotic phenotypes of Medicago truncatula and Pisum sativum. Orobanche crenata inoculation of mutant lines carrying defective mutation in the genes dmi2/sym19 and dmi3 resulted in an increase in O. crenata establishment. Similarly, inoculation of mutants carrying mutation in the gene sunn/sym29 that controls the autoregulation mechanism of the symbiosis, also lead to a significant increase in haustoria formation. Altogether, our results suggest that parasitic plant infection is partly controlled by both the conserved symbiotic pathway that mediates symbiont recognition and establishment and the autoregulation mechanism that regulates the extent of colonisation by Rhizobium and AM fungi.

Characterization of resistance response of pea (Pisum spp.) against rust (Uromyces pisi)

AbstractComponents of resistance to the pea rust (Uromyces pisi) were studied at the histological level in seedlings of seven pea (Pisum spp.) accessions showing partial resistance under field conditions. Resistance was characterized macroscopically by an increased latent period and a decreased infection frequency in spite of a compatible interaction (high infection type). Histological investigations revealed little differences among accessions in spore germination and appressoria formation, differences being more evident once the stomata were penetrated by the infection structures. Resistance was mainly due to a restriction of haustorium formation with significant levels of early aborted colonies in accessions IFPI3260, PI347321 and PI347347. Furthermore, a reduction in the number of haustoria per colony, hyphal tips per colony and smaller colony size were found in all genotypes studied. Resistance was not associated with early‐ or late‐acting hypersensitivity in any of the genotypes studied.

Specificity and levels of nonhost resistance to nonadapted Blumeria graminis forms in barley

The genetic basis of nonhost resistance of barley to nonadapted formae speciales of Blumeria graminis is not known, as there is no barley line that is susceptible to these nonadapted formae speciales, such as the wheat powdery mildew pathogen, Blumeria graminis f.sp. tritici (Bgt). Barley accessions with rudimentary susceptibility to an isolate of the nonadapted Bgt were identified. Those accessions were intercrossed in two cycles and two lines, called SusBgt(SC) and SusBgt(DC), with substantial susceptibility to Bgt at the seedling stage were selected. The quantitative variation among barley accessions and in the progenies after convergent crossing suggests a polygenic basis for this nonhost resistance. Both lines allowed an unusually high level of haustorium formation and colony development by Bgt. The SusBgt lines and their ancestor lines also allowed haustorium formation and conidiation by four out of seven isolates of other nonadapted B. graminis forms. Analysis of the infection process suggested that nonhost resistance factors are specific to the form and developmental stage of B. graminis. Resistances to establishment (first haustorium), colonization (subsequent haustoria) and conidiation are not associated. The lines developed will be of use in elucidating the genetic basis of nonhost resistance to Bgt in barley, and in gene expression and complementation studies on nonhost resistance.

Identification and characterization of partial resistance to rust in a germplasm collection of Lathyrus sativus L.

Abstract Lathyrus sativus has high potential both as food and fodder crop but can be severely infected by rust. This study assessed the resistance level of a collection of L. sativus accessions against three rust species (Uromyces pisi, U. ciceris‐arietini and U. viciae‐fabae) and characterized, for the first time, macro and microscopically the resistance to U. pisi. All L. sativus accessions were highly resistant to U. ciceris‐arietini and to U. viciae‐fabae but showed a compatible interaction with U. pisi. However, in spite of this compatible interaction, high levels of rate reducing resistance were identified with significant disease severity reduction, fitting the Partial Resistance definition. Detailed histological observations showed that resistance was mainly because of haustorium formation restriction with high‐level of colony early abortion, and reduction in the haustoria number per colony. This was occasionally associated with reduction on the infection hyphae intercellular growth, in a clear example of prehaustorial resistance. Differences in germination, orientated germ tube growth and appressoria differentiation were observed but were, generally, of marginal importance to explain the observed resistance levels.

Infection and disease development of Quambalaria spp. on Corymbia and Eucalyptus species

Quambalaria spp. are eucalypt leaf and shoot pathogens of growing global importance, yet virtually nothing is known regarding the manner in which they infect and colonize their hosts. A study of the infection process of Q. pitereka and Q. eucalypti on Corymbia and Eucalyptus species was thus undertaken using light, scanning and transmission electron microscopy after artificial inoculation. Conidial germination was triggered when relative humidity levels exceeded 90% and commenced within 2 h in the presence of free water. Light reduced germination but did not prevent germination from occurring. Conidial germination and hyphal growth occurred on the upper and lower leaf surfaces with penetration occurring via the stomata or wounds on the leaf surface or juvenile stems. There was no evidence of direct penetration of the host. Following penetration through the stomata, Q. pitereka and Q. eucalypti hyphae grew only intercellularly without the formation of haustoria or interaction apparatus, which is characteristic of the order Microstromatales. Instead, the presence of an interaction zone is demonstrated in this paper. Conidiophores arose through stomatal openings producing conidia 7 days after infection.

Analysis of isolation and expression analysis of ItCYP79B2 during rust infection of dyer's woad (Isatis tinctoria) by Puccinia thlaspeos

Dyer's woad (Isatis tinctoria) has been shown to have unusually high quantities of indole glucosinolates. It is known that production of glucosinolates and their hydrolysis products is part of a plant's defense response to pathogens and insects. In Arabidopsis thaliana, which, like dyer's woad, is in the family Brassicaceae, the gene sequences encoding CYP79B2 and CYP79B3 catalyze the conversion of tryptophan to indole-3-acetaldoxime during the biosynthesis of indole glucosinolates. In this study, a sequence encoding CYP79B2 was isolated from dyer's woad. ItCYP79B2 from I. tinctoria shows 97% sequence identity with CYP79B2 and 89% sequence identity with CYP79B3 from A. thaliana. Thus, it can be inferred that the I. tinctoria sequence, like that of A. thaliana, is involved in the biosynthesis of indole glucosinolates. The kinetics and amplitude of ItCYP79B2 expression during the first 72 h after infection by the pathogenic rust fungus Puccinia thlaspeos were also studied. There was a significant down-regulation of ItCYP79B2 during the first 8 h after infection of the host that coincides with fungal penetration. This was followed by induction of ItCYP79B2, coinciding with the formation of haustoria. However, after haustoria formation, ItCYP9B2 was again suppressed during the time period coinciding with successful asymptomatic systemic colonization of the host. The results of this study indicates that suppression of a gene involved in indole glucosinolate production during the penetration and post-haustorial phases of infection plays a role in pathogen establishment. They also imply that the pathogen possesses mechanisms for circumventing the elevated expression of this gene during the haustorial phase

Functional Contribution of Chorismate Synthase, Anthranilate Synthase, and Chorismate Mutase to Penetration Resistance in Barley–Powdery Mildew Interactions

Plant processes resulting from primary or secondary metabolism have been hypothesized to contribute to defense against microbial attack. Barley chorismate synthase (HvCS), anthranilate synthase alpha subunit 2 (HvASa2), and chorismate mutase 1 (HvCM1) occupy pivotal branch points downstream of the shikimate pathway leading to the synthesis of aromatic amino acids. Here, we provide functional evidence that these genes contribute to penetration resistance to Blumeria graminis f. sp. hordei, the causal agent of powdery mildew disease. Single-cell transient-induced gene silencing of HvCS and HvCM1 in mildew resistance locus a (Mla) compromised cells resulted in increased susceptibility. Correspondingly, overexpression of HvCS, HvASa2, and HvCM1 in lines carrying mildew resistance locus o (Mlo), a negative regulator of penetration resistance, significantly decreased susceptibility. Barley stripe mosaic virus-induced gene silencing of HvCS, HvASa2, and HvCM1 significantly increased B. graminis f. sp. hordei penetration into epidermal cells, followed by formation of haustoria and secondary hyphae. However, sporulation of B. graminis f. sp. hordei was not detected on the silenced host plants up to 3 weeks after inoculation. Taken together, these results establish a previously unrecognized role for the influence of HvCS, HvASa2, and HvCM1 on penetration resistance and on the rate of B. graminis f. sp. hordei development in Mla-mediated, barley-powdery mildew interactions.