powdery mildew Medicago truncatula
214315853
Medicago truncatula as a model to study powdery mildew resistance
Nicolas Rispail Elena Prats Diego Rubiales
133605512
Nodulation induces systemic resistance of Medicago truncatula and Pisum sativum against Erysiphe pisi and primes for powdery mildew triggered salicylic acid accumulation.
Plants encounter beneficial and detrimental microorganisms both above and below ground and the health status of the plant depends on the composition of this pan microbiome. Beneficial microorganisms contribute to plant nutrition and/or systemically or locally protect plants against pathogens, thus facilitating adaptation to a variety of environments. Induced systemic resistance, caused by root associated microbes, manifests as above ground resistance against necrotrophic pathogens and is mediated by jasmonic acid/ethylene signaling. By contrast, systemic acquired resistance relies on salicylic acid (SA) and confers resistance against secondary infection by (hemi)biotrophic pathogens. To investigate whether symbiotic rhizobia that are ubiquitously found in natural ecosystems are able to modulate resistance against biotrophs, we tested the impact of pre established nodulation of Medicago truncatula and pea (Pisum sativum) plants against infection by the powdery mildew fungus Erysiphe pisi. We found that root symbiosis interfered with fungal penetration of M. truncatula and reduced asexual spore formation on pea leaves independently of symbiotic nitrogen fixation. Improved resistance of nodulated plants correlated with elevated levels of free SA and SA dependent marker gene expression upon powdery mildew infection. Our results suggest that nodulation primes the plants systemically for E. pisi triggered SA accumulation and defense gene expression, resulting in increased resistance.
Molecular plant microbe interactions MPMI
Lara Smigielski Eva-Maria Laubach Lina Pesch Joanna Marie Leyva Glock Frank Albrecht Alan John Slusarenko Ralph Panstruga Hannah Kuhn
209385518
Dual RNA Seq analysis of Medicago truncatula and the pea powdery mildew Erysiphe pisi uncovers distinct host transcriptional signatures during incompatible and compatible interactions and pathogen effector candidates.
Powdery mildew (PM) is a serious fungal disease of legumes. To gain novel insights into PM pathogenesis and host resistance/susceptibility, we used dual RNA Seq to simultaneously capture host and pathogen transcriptomes at 1 d post inoculation of resistant and susceptible Medicago truncatula genotypes with the PM Erysiphe pisi (Ep) Differential expression analysis indicates that R gene mediated resistance against Ep involves extensive transcriptional reprogramming. Functional enrichment of differentially expressed host genes and in silico analysis of co regulated promoters suggests that amplification of PTI, activation of the JA/ET signaling network, and regulation of growth defense balance correlate with resistance. In contrast, processes that favor biotrophy, including suppression of defense signaling and programmed cell death, and weaker cell wall defenses are important susceptibility factors. Lastly, Ep effector candidates and genes with known/putative virulence functions were identified, representing a valuable resource that can be leveraged to improve our understanding of legume PM interactions.
Genomics
Megha Gupta Gunjan Sharma Divya Saxena Roli Budhwar Madavan Vasudevan Varsha Gupta Arunima Gupta Rashi Gupta Divya Chandran
231900484
The Medicago truncatula Sugar Transport Protein 13 and its Lr67res like Variant Confer Powdery Mildew Resistance in Legumes via Defense Modulation.
Obligate biotrophic pathogens like the pea powdery mildew (PM) Erysiphe pisi establish long term feeding relationships with their host, during which they siphon sugars from host cells through haustoria. Plants in turn deploy sugar transporters to restrict carbon allocation towards pathogens, as a defense mechanism. Studies in Arabidopsis have shown that sugar transport protein 13 (STP13) a proton hexose symporter involved in apoplasmic hexose retrieval, contributes to bacterial and necrotrophic fungal resistance by limiting sugar flux towards these pathogens. By contrast, expression of Lr67res, a transport deficient wheat STP13 variant harboring two amino acid substitutions (G144R and V387L) conferred resistance against biotrophic fungi in wheat and barley, indicating its broad applicability in disease management. Here we investigated the role of STP13 and STP13G144Rin legume PM interactions. We show that Medicago truncatula STP13.1 is a proton hexose symporter involved in basal resistance against PM, and indirectly show that Lr67res mediated PM resistance, so far reported only in monocots, is transferable to legumes. Among the 30 MtSTPs, STP13.1 exhibited the highest fold induction in PM challenged leaves, and was also responsive to chitosan, ABA and sugar treatment. Functional assays in yeast showed that introduction of the G144R mutation but not V388L abolished MtSTP13.1's hexose uptake ability. Virus induced gene silencing of MtSTP13 repressed PR gene expression and enhanced PM susceptibility in M. truncatula whereas transient overexpression of MtSTP13.1 or MtSTP13.1G144R in pea induced PR and isoflavonoid pathway genes and enhanced PM resistance. We propose a model in which STP13.1 mediated sugar signaling triggers defense responses against PM in legumes.
Plant cell physiology
Megha Gupta Shubham Dubey Deepti Jain Divya Chandran
205215702
Epigenetic regulation of a powdery mildew resistance gene in Medicago truncatula.
Dear Editor, Medicago truncatula is a model legume that is congeneric with alfalfa (Medicago sativa) a forage crop of global importance. Over the last decade, tremendous genetic and genomic tools have been developed for this model system, which has greatly facilitated the study of various aspects of legume genomics and biology. From an applied perspective, genomic information gained from M. truncatula is particularly useful for genetic improvement of cultivated alfalfa, a crop that is not amenable to genetic analysis because of its allogamous and autotetraploid nature (Yang et al. 2008) For instance, M. truncatula can be used to clone the orthologs of many economically important genes in alfalfa, because the two species share many common phenotypes. Owing to the high level of sequence homology and remarkably conserved gene function between the two species, the isolated genes from M. truncatula can be directly used for alfalfa improvement by transgenic approaches. We used M. truncatula as a model system to identify genes that confer resistance to powdery mildew caused by the biotrophic fungus Erysiphe pisi. For this purpose, the resistant genotype Jemalong A17 and the susceptible genotype F83005.5 were crossed to produce an F2 mapping population. The F2 plants were inoculated with an Erysiphe pisi race (we called race 1) originally obtained from naturally occurring infection on the leaves of F83005.5. Seven to 10 d after inoculation, the resistant plants grew normally and were symptomless, while the susceptible plants displayed typical powdery mildew symptoms presented by white patches of mycelium and conidia covering the leaf surface (Figure 1A) Phenotyping of 1107 F2 plants identified 822 resistant and 285 susceptible individuals. The segregation of resistance to susceptibility fits 3:1 ratio (kh2 0.31, df 1, P 0.58) indicating that a single dominant gene controls the powdery mildew resistance in Jemalong A17. This resistance gene was designated as MtREP1 (for resistance to Erysiphe pisi race 1) We used the susceptible F2 individuals for genetic mapping of the MtREP1 locus. Initial genotyping using simple sequence repeat (SSR) markers revealed that the MtREP1 locus is located between markers MtB330 and MtB38 on the M. truncatula chromosome 5 (Figure 1A) a position coincident with one of the two QTL loci that confer powdery mildew resistance against an E. pisi isolate from alfalfa (Ameline Torregrosa et al. 2008) The availability of genomic sequence around this region allowed us to develop sequencebased CAPS (cleaved amplified polymorphic sequences) markers to finely map the locus. Through this process, we were able to delimit the MtREP1 locus between CAP52 and CAP23 that span ~194 kb, within which CAP8, CAP9, CAP20, and CAP22 were co segregated with MtREP1 (Figure 1A) The 194 kb window contains more than 30 annotated genes (Medtr5g072060 Medtr5g072480) three of which encode a typical coiled coil/nucleotide binding site/leucinerich repeat (CC NBS LRR) plant R protein (Medtr5g072140, Medtr5g072250, and Medtr5g072340) (Young et al. 2011; version Mt3.5) These three CC NBS LRR genes were considered as candidates of MtREP1. We identified a single bacterial artificial chromosome (BAC) clone mth2 76g24 that contains all three candidate genes, from which genomic sequences for each candidate gene, including their native promoters and 3' untranslated (UTR) regions, were obtained and cloned into the pCAMBIA2300 vector. Because M. truncatula is recalcitrant to efficient Agrobacterium tumefaciens mediated transformation, we chose to use alfalfa as an alternative study system. For this purpose, we first screened the alfalfa cultivar Pioneer 53V08 to identify plants that were susceptible to the E. pisi race 1. The selected susceptible plants were then subjected to A. tumefaciens mediated transformation using the genomic constructs described above. We obtained transgenic plants for each candidate gene, all of which were inoculated with the powdery mildew pathogen. Our results showed that independent transgenic plants containing Medtr5g072140 (n 23) Medtr5g072250 (n 11) or the empty vector pCAMBIA2300 (n 19) were all susceptible. In contrast, independent transformants containing the Medtr5g072340 transgene (n 15) were completely resistant to the pathogen (Figure 1B) Thus, we conclude that Medtr5g072340 is the MtREP1 gene. Alignment of genomic and cDNA sequences reveals that MtREP1 coding sequence is composed of a single exon that is predicted to encode a protein of 992 amino acids, consisting of an N terminal CC domain, a centrally located NBS domain, and numerous degenerate LRRs C terminal to the NBS domain. 3' RACE (rapid amplification of cDNA ends) analysis identified 1486 bp transcribed sequence after the stop codon. Sequence alignment of the 1486 bp cDNA sequence with genomic DNA
Molecular plant
Shengming Yang Fang Tang Eveline Teixeira Caixeta Hongyan Zhu
3389429
Mining the Cicer arietinum genome for the mildew locus O (Mlo) gene family and comparative evolutionary analysis of the Mlo genes from Medicago truncatula and some other plant species
The mildew locus O (Mlo) gene family is ubiquitous in land plants. Some members of this gene family are involved in negative regulation of powdery mildew resistance, while others are involved in several other biological functions. Mlo proteins have characteristic seven transmembrane domains and a calmodulin binding domain at their C termini, and are associated with plasma membrane. The Mlo gene family has been studied in several economically important cereals, but little information is available on this gene family in the important legumes, Medicago truncatula Gaertn. and Cicer arietinum L. We carried out a comprehensive and comparative investigation of the Mlo gene family in these two species using the genome sequences available at the M. truncatula genome database (Mt v4.0) and NCBI (C. arietinum) A genome wide homology based search using Arabidopsis Mlo proteins as query identified 16 MtMlo (M. truncatula Mlo) and 14 CarMlo (C. arietinum Mlo) genes. The MtMlo and CarMlo genes had comparable gene structure, protein sequence and topology. Their chromosomal locations indicated the occurrence of extensive reorganization in the genomes of the two species after their divergence from the common ancestor. A multiple sequence alignment of 53 Mlo proteins from these two and several other species showed a highly conserved sequence block of seven amino acids, viz. L ETPTW, towards their N termini. The evolutionary phylogenetic analysis grouped the MtMlo and CarMlo members into four clusters, and most of the MtMlo and CarMlo members formed one to one ortholog pairs. The ka/ks analyses indicated that the MtMlo and CarMlo genes are subjected to intense purifying selection.
Journal of Plant Research
Reena Deshmukh V K Singh Brahma Deo Singh
15234038
Genetic dissection of resistance to anthracnose and powdery mildew in Medicago truncatula.
Medicago truncatula was used to characterize resistance to anthracnose and powdery mildew caused by Colletotrichum trifolii and Erysiphe pisi, respectively. Two isolates of E. pisi (Ep p from pea and Ep a from alfalfa) and two races of C. trifolii (races 1 and 2) were used in this study. The A17 genotype was resistant and displayed a hypersensitive response after inoculation with either pathogen, while lines F83005.5 and DZA315.16 were susceptible to anthracnose and powdery mildew, respectively. To identify the genetic determinants underlying resistance in A17, two F7 recombinant inbred line (RIL) populations, LR4 (A17 x DZA315.16) and LR5 (A17 x F83005.5) were phenotyped with E. pisi isolates and C. trifolii races, respectively. Genetic analyses showed that i) resistance to anthracnose is governed mainly by a single major locus to both races, named Ct1 and located on the upper part of chromosome 4; and ii) resistance to powdery mildew involves three distinct loci, Epp1 on chromosome 4 and Epa1 and Epa2 on chromosome 5. The use of a consensus genetic map for the two RIL populations revealed that Ct1 and Epp1, although located in the same genome region, were clearly distinct. In silico analysis in this region identified the presence of several clusters of nucleotide binding site leucine rich repeat genes. Many of these genes have atypical resistance gene analog structures and display differential expression patterns in distinct stress related cDNA libraries.
Molecular plant microbe interactions MPMI
Carine Ameline-Torregrosa Marc Cazaux Dariush Danesh Fabien Chardon Steven B Cannon Marie-Therese Esquerre-Tugaye Bernard Dumas Nevin D Young Deborah A Samac Thierry Huguet Christophe Jacquet
17284797
Transcriptional profiling of Medicago truncatula during Erysiphe pisi infection
Resistance to powdery mildew has been studied in a number of plant species, yet the molecular mechanisms remain largely unknown. Transcription factors (TFs) play a critical role in the plant defense response by regulating the transcriptional machinery which coordinates the expression of a large group of genes involved in plant defense. Using high throughput quantitative real time PCR (qPCR) technology more than 1000 Medicago truncatula TFs were screened in a pair of susceptible and resistant genotypes of M. truncatula after 4 h of Erysiphe pisi infection. Seventy nine TF genes, belonging to 33 families showed a significant transcriptional change in response to E. pisi infection. Forty eight TF genes were differentially expressed in the resistant genotypes compared to the susceptible one in response to E. pisi infection, including pathogenesis related transcriptional factors, AP2/EREBP (APETALA2/ETHYLENE RESPONSIVE ELEMENT BINDING FACTORS) WRKY (highly conserved WRKYGQK amino acid sequence) MYB (Myeloblastoma) homeodomain (HD) and zinc finger C2C2 (CYS2 CYS2) C2H2, (CYS2 HIS2) LIM (Lin 11, Isl 1, Mec 3) gene families, which are involved in known defense responses. Our results suggest that these TF genes are among the E. pisi responsive genes in resistant M. truncatula that may constitute a regulatory network which controls the transcriptional changes in defense genes involved in resistance to E. pisi.
Front. Plant Sci.
Miguel Curto Franziska Krajinski Armin Schlereth Diego Rubiales
18194993
Plant Defense Responses in Medicago truncatula Unveiled by Microarray Analysis
Plant defense responses are made up of broad plant defense mechanisms that involve an integrated signaling pathway. Powdery mildew caused by Erysiphe pisi is one of the most important diseases of pea. However, the mechanisms and pathways involved in the resistance against E. pisi are yet undiscovered. We studied the transcriptome of two Medicago truncatula genotypes, the powdery mildew susceptible commercial variety Parabinga and the resistant accession SA1306, at 4 and 12 h after E. pisi infection, using Mt16kOLI1 microarrays. Four hundred and forty six probes were differentially expressed between the two M. truncatula genotypes along the time points studied. RNA accumulation patterns suggest that the most prominent responses to pathogen infection occur at early infection stages. Most of the regulated genes are related to cell wall reinforcement, flavonoid, and phenylpropanoid biosynthesis. In addition, pathogenesis related proteins and signaling pathways controlled by jasmonic acid and salicylic acid were found to be regulated during pathogen infection. This study provides the first comprehensive view of the genes and pathways activated in the E. pisi/M. truncatula pathosystem, allowing the identification of targets against this important disease.
Plant Molecular Biology Reporter
Miguel Curto Franziska Krajinski Helge Kuster Diego Rubiales
31251512
Durable broad spectrum powdery mildew resistance in pea er1 plants is conferred by natural loss of function mutations in PsMLO1.
Loss of function alleles of plant specific MLO (Mildew Resistance Locus O) genes confer broad spectrum powdery mildew resistance in monocot (barley) and dicot (Arabidopsis thaliana, tomato) plants. Recessively inherited powdery mildew resistance in pea (Pisum sativum) er1 plants is, in many aspects, reminiscent of mlo conditioned powdery mildew immunity, yet the underlying gene has remained elusive to date. We used a polymerase chain reaction (PCR) based approach to amplify a candidate MLO cDNA from wild type (Er1) pea. Sequence analysis of the PsMLO1 candidate gene in two natural er1 accessions from Asia and two er1 containing pea cultivars with a New World origin revealed, in each case, detrimental nucleotide polymorphisms in PsMLO1, suggesting that PsMLO1 is Er1. We corroborated this hypothesis by restoration of susceptibility on transient expression of PsMLO1 in the leaves of two resistant er1 accessions. Orthologous legume MLO genes from Medicago truncatula and Lotus japonicus likewise complemented the er1 phenotype. All tested er1 genotypes showed unaltered colonization with the arbuscular mycorrhizal fungus, Glomus intraradices, and with nitrogen fixing rhizobial bacteria. Our data demonstrate that PsMLO1 is Er1 and that the loss of PsMLO1 function conditions durable broad spectrum powdery mildew resistance in pea.
Molecular plant pathology
Matt Humphry Anja Reinstadler Sergey Ivanov Ton Bisseling Ralph Panstruga