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Long-read annotation : Automated eukaryotic genome annotation based on long-read cDNA sequencing
Cook, David E. ; Valle-Inclan, Jose Espejo ; Pajoro, Alice ; Rovenich, Hanna ; Thomma, Bart P.H.J. ; Faino, Luigi - \ 2019
Plant Physiology 179 (2019)1. - ISSN 0032-0889 - p. 38 - 54.
Single-molecule full-length complementary DNA (cDNA) sequencing can aid genome annotation by revealing transcript structure and alternative splice forms, yet current annotation pipelines do not incorporate such information. Here we present long-read annotation (LoReAn) software, an automated annotation pipeline utilizing short-and long-read cDNA sequencing, protein evidence, and ab initio prediction to generate accurate genome annotations. Based on annotations of two fungal genomes (Verticillium dahliae and Plicaturopsis crispa) and two plant genomes (Arabidopsis [Arabidopsis thaliana] and Oryza sativa), we show that LoReAn outperforms popular annotation pipelines by integrating single-molecule cDNA-sequencing data generated from either the Pacific Biosciences or MinION sequencing platforms, correctly predicting gene structure, and capturing genes missed by other annotation pipelines.
Specific Hypersensitive Response–Associated Recognition of New Apoplastic Effectors from Cladosporium fulvum in Wild Tomato
Mesarich, Carl H. ; Ӧkmen, Bilal ; Rovenich, Hanna ; Griffiths, Scott A. ; Wang, Changchun ; Karimi Jashni, Mansoor ; Mihajlovski, Aleksandar ; Collemare, Jérôme ; Hunziker, Lukas ; Deng, Cecilia H. ; Burgt, Ate Van Der; Beenen, Henriek G. ; Templeton, Matthew D. ; Bradshaw, Rosie E. ; Wit, Pierre J.G.M. De - \ 2018
Molecular Plant-Microbe Interactions 31 (2018)1. - ISSN 0894-0282 - p. 145 - 162.
Tomato leaf mold disease is caused by the biotrophic fungus Cladosporium fulvum. During infection, C. fulvum produces extracellular small secreted protein (SSP) effectors that function to promote colonization of the leaf apoplast. Resistance to the disease is governed by Cf immune receptor genes that encode receptor-like proteins (RLPs). These RLPs recognize specific SSP effectors to initiate a hypersensitive response (HR) that renders the pathogen avirulent. C. fulvum strains capable of overcoming one or more of all cloned Cf genes have now emerged. To combat these strains, new Cf genes are required. An effectoromics approach was employed to identify wild tomato accessions carrying new Cf genes. Proteomics and transcriptome sequencing were first used to identify 70 apoplastic in planta–induced C. fulvum SSPs. Based on sequence homology, 61 of these SSPs were novel or lacked known functional domains. Seven, however, had predicted structural homology to antimicrobial proteins, suggesting a possible role in mediating antagonistic microbe-microbe interactions in planta. Wild tomato accessions were then screened for HR-associated recognition of 41 SSPs, using the Potato virus X–based transient expression system. Nine SSPs were recognized by one or more accessions, suggesting that these plants carry new Cf genes available for incorporation into cultivated tomato.
Evasion of chitin-triggered immunity by fungal plant pathogens
Rövenich, Hanna J. - \ 2017
Wageningen University. Promotor(en): B.P.H.J. Thomma; P.J.G.M. de Wit. - Wageningen : Wageningen University - ISBN 9789463436137 - 133
plant-microbe interactions - immunity - receptors - verticillium dahliae - cladosporium - plant pathogens - chitin - arabidopsis thaliana - fungi - plant-microbe interacties - immuniteit - receptoren - verticillium dahliae - cladosporium - plantenziekteverwekkers - chitine - arabidopsis thaliana - schimmels
Plants establish intricate relationships with microorganisms that range from mutualistic to pathogenic. In order to prevent colonization by potentially harmful microbes, plant hosts employ surface-localized receptor molecules that perceive ligands, which are either microbe-derived or result from microbe-mediated plant manipulation. This recognition ultimately leads to the activation of host immunity. In order to circumvent recognition or suppress immune responses, microbes secrete effector proteins that deregulate host physiological processes. While the number of identified putative effectors has rapidly increased in recent years, their functions and the mechanisms governing their recognition have largely remained unexplored. To enhance our understanding of the molecular interplay between host and microbe, the work presented here was designed to identify further components involved in the recognition of the two fungal pathogens Verticillium dahliae and Cladosporium fulvum, as well as to characterize the functions of effector proteins produced by these pathogens during tomato infection.
|The two-speed genome of Verticillium dahliae mediates emergence of potent virulence factors
Thomma, B.P.H.J. ; Faino, L. ; Li, J. ; Shi-Kunne, X. ; Depotter, J.R.L. ; Kramer, H.M. ; Berg-Velthuis, G.C.M. van den; Cook III, David ; Rövenich, H.J. ; Seidl, M.F. - \ 2017
In: Book of Abstracts 29th Fungal Genetics Conference Asilomar 17, Pacific Grove, CA, USA 14-19 March 2017. - - p. 4 - 4.
Genomic plasticity enables adaptation to changing environments, which is especially relevant for pathogens that engage in “arms races” with their hosts. In many pathogens, virulence genes reside in highly variable, transposon-rich, physically distinct genomic compartments. However, understanding of the evolution of such compartments, and the role of transposons therein, remains limited. We show that transposons are the major driving force for adaptive genome evolution in the fungal plant pathogen Verticillium dahliae, and that highly variable lineage-specific (LS) regions evolved by genomic rearrangements that are mediated by erroneous double-strand repair, often utilizing transposons. Remarkably, LS regions are enriched in active transposons, which may contribute to local genome plasticity. Thus, we provide evidence for genome shaping by transposons, both in an active and passive manner, which impacts the evolution of V. dahliae virulence. Based on this knowledge, we are now able to identify crucial virulence factors of V. dahliae, which also allows investigating causal relationships between particular effectors and pathotypes.
The receptor kinase FER is a RALF-regulated scaffold controlling plant immune signaling
Stegmann, Martin ; Monaghan, Jacqueline ; Smakowska-Luzan, Elwira ; Rovenich, Hanna ; Lehner, Anita ; Holton, Nicholas ; Belkhadir, Youssef ; Zipfel, Cyril - \ 2017
Science 355 (2017)6322. - ISSN 0036-8075 - p. 287 - 289.
In plants, perception of invading pathogens involves cell-surface immune receptor kinases. Here, we report that the Arabidopsis SITE-1 PROTEASE (S1P) cleaves endogenous RAPID ALKALINIZATION FACTOR (RALF) propeptides to inhibit plant immunity. This inhibition is mediated by the malectin-like receptor kinase FERONIA (FER), which otherwise facilitates the ligand-induced complex formation of the immune receptor kinases EF-TU RECEPTOR (EFR) and FLAGELLIN-SENSING 2 (FLS2) with their co-receptor BRASSINOSTEROID INSENSITIVE 1–ASSOCIATED KINASE 1 (BAK1) to initiate immune signaling. We show that FER acts as a RALF-regulated scaffold that modulates receptor kinase complex assembly. A similar scaffolding mechanism may underlie FER function in other signaling pathways.
Verticillium dahliae LysM effectors differentially contribute to virulence on plant hosts
Kombrink, Anja ; Rovenich, Hanna ; Shi, Xiaoqian ; Rojas-Padilla, Eduardo ; Berg-Velthuis, Grardy van den; Domazakis, Emmanouil ; Jonge, Ronnie De; Valkenburg, Dirk-Jan ; Sánchez-Vallet, Andrea ; Seidl, Michael F. ; Thomma, Bart P.H.J. - \ 2017
Molecular Plant Pathology 18 (2017)4. - ISSN 1464-6722 - p. 596 - 608.
Chitin-binding LysM effectors contribute to virulence of various plant pathogenic fungi that are causal agents of foliar diseases. Here, we report on LysM effectors of the soil-borne fungal vascular wilt pathogen Verticillium dahliae. Comparative genomics revealed three core LysM effectors that are conserved in a collection of V. dahliae strains. Remarkably, and in contrast to the previously studied LysM effectors of other plant pathogens, no expression of core LysM effectors was monitored in planta in a taxonomically diverse panel of host plants. Moreover, targeted deletion of the individual LysM effector genes in V. dahliae strain JR2 did not compromise virulence in infections on Arabidopsis, tomato or Nicotiana benthamiana. Interestingly, an additional lineage-specific LysM effector is encoded in the genome of V. dahliae strain VdLs17 but not in any other V. dahliae strain sequenced to date. Remarkably, this lineage-specific effector is expressed in planta and contributes to virulence of V. dahliae strain VdLs17 on tomato, but not on Arabidopsis or on N. benthamiana. Functional analysis revealed that this LysM effector binds chitin, is able to suppress chitin-induced immune responses, and protects fungal hyphae against hydrolysis by plant hydrolytic enzymes. Thus, in contrast to the core LysM effectors of V. dahliae, this lineage-specific LysM effector of strain VdLs17 contributes to virulence in planta.
Novel effectors identified in the apoplast of Cladosporium fulvum-infected tomato
Mesarich, C. ; Ökmen, B. ; Rövenich, H.J. ; Karimi Jashni, M. ; Wang, C. ; Griffiths, S.A. ; Collemare, J.A.R. ; Deng, C. ; Wit, P.J.G.M. de - \ 2016
Tomato leaf mold disease is caused by the biotrophic fungal pathogen Cladosporium fulvum. To colonize the leaf apoplast, C. fulvum secretes a collection of effector proteins that modulate host immune responses, as well as other proteins (e.g., carbohydrate-active enzymes or CAZys) that facilitate nutrient acquisition. In the presence of cognate Cf immune receptors, however, many of these proteins trigger immune responses that render the pathogen avirulent. Characterization of the C. fulvum apoplastic secretome is required to further understand the abovementioned processes, and to identify novel sources of resistance against this pathogen. We have used liquid chromatography–tandem mass spectrometry (LC–MS/MS) to identify 141 secreted and surface-associated fungal proteins present in apoplastic fluid harvested from compatible C. fulvum–tomato interactions. In addition to the known effectors identified in previous studies, this collection contains >70 new C. fulvum candidate effector (CfCE) proteins. Using a Potato virus X (PVX)-based expression system, we show that nine of these CfCEs, including Ecp11-1, which has homology to AvrLm3 and AvrLmJ1 of Leptosphaeria maculans, trigger cell death in particular wild accessions of tomato. Thus, our study has likely uncovered novel avirulence effectors of C. fulvum, as well as Cf immune receptors in wild tomato with new specificities against this pathogen. An overview of the C. fulvum apoplastic secretome will be presented.
Is the Verticillium effector Ave1 a dual function protein?
Rövenich, H.J. ; Boshoven, J.C. ; Grandellis, C. ; Seidl, M.F. ; Ottado, J. ; Thomma, B.P.H.J. - \ 2016
Verticillium dahliae is a soil-borne fungal pathogen with a broad host range. Comparative genomics and phylogenetic analyses identified the virulence gene Ave1, which has many homologs in plants, several other fungi and the bacterial pathogen Xanthomonas axonopodis pv. citri (Xac). The homology is determined by the presence of a highly conserved sequence coding for a so-called plant natriuretic peptide (PNP). PNPs constitute a class of systemically mobile molecules that function in plant homeostasis and growth. The microbial homolog characterized to date is XacPNP. Similar to Arabidopsis PNP-A, XacPNP alters physiological responses such as stomatal opening in plants. During infection, XacPNP is required for the suppression of host cell death to establish favourable conditions for the survival of the bacterial biotroph. In vitro stomatal aperture assays in Arabidopsis epidermis tissue demonstrated that also Ave1 from V. dahliae retains an active PNP site. Interestingly, however, complementation of ΔAve1 in V. dahliae with homologous sequences from tomato or Xac did not result in the recovery of V. dahliae virulence. In addition, structural modelling suggests that, compared to its homologs, V. dahliae Ave1 has a highly variable surface opposite the conserved PNP site. These findings indicate that Ave1 may have functionally diverged from its microbial and plant homologs and obtained an additional function that contributes to V. dahliae virulence.
Convergent evolution of filamentous microbes towards evasion of glycan-triggered immunity
Rovenich, Hanna ; Zuccaro, Alga ; Thomma, Bart P.H.J. - \ 2016
New Phytologist 212 (2016)4. - ISSN 0028-646X - p. 896 - 901.
Chitin - Effector - Glycan recognition - Microbial cell wall - β-glucan
I. II. III. IV. V. VI. References Summary: All filamentous microbes produce and release a wide range of glycans, which are essential determinants of microbe-microbe and microbe-host interactions. Major cell wall constituents, such as chitin and β-glucans, are elicitors of host immune responses. The widespread capacity for glycan perception in plants has driven the evolution of various strategies that help filamentous microbes to evade detection. Common strategies include structural and chemical modifications of cell wall components as well as the secretion of effector proteins that suppress chitin- and β-glucan-triggered immune responses. Thus, the necessity to avoid glycan-triggered immunity represents a driving force in the convergent evolution of filamentous microbes towards its suppression.
|Towards the determination of exact interaction mechanisms between the tomato immune receptor Ve1 and the fungal effector Ave1
Rövenich, H.J. ; Zhang, Z. ; Song, Y. ; Thomma, B. - \ 2015
In: Book of Abstracts 28th Fungal Genetics Conference. - - p. 213 - 213.
Recognition of pathogen effectors, small secreted molecules that facilitate host colonization, represents the basis for plant resistance against well-adapted pathogens. In tomato, the Ve1 gene confers resistance against race 1 strains of the vascular wilt pathogen V. dahliae. Ve1 encodes a surface-localized leucine-rich repeat (LRR) receptor-like protein (RLP), which recognizes the recently identified secreted V. dahliae effector Ave1. Immunopurification of affinity-tagged Ave1 transiently co-expressed with HA-tagged Ve1 in tobacco showed that the receptor protein co-purifies with Ave1, indicating an interaction between the pathogen effector and the immune receptor. A mutational screen of extracellular solvent-exposed residues across the LRR domains within the C1 region of Ve1 showed that stable Ve1 mutant alleles of the two consecutive LRR regions LRR3-LRR8 and LRR20-LRR23 were compromised in their HR-inducing activity when co-expressed with Ave1. Accordingly, when challenged with race1 V. dahliae, transgenic Arabidopsis plants carrying the non-functional Ve1 alleles displayed symptoms similar to inoculated non-transgenic controls. These results suggest that Ve1 functionality is determined by two distinct clusters of LRR domains that may be involved Ave1 perception. Despite the fact that further mutational analyses in combination with biochemical methodologies can be employed to further elucidate the interaction between the receptor and the effector proteins, structural information will greatly enhance our ability to improve those strategies.
|Evolution of virulence in the vascular wilt pathogen Verticillium dahliae
Faino, L. ; Seidl, M.F. ; Cook III, D.E. ; Shi-Kunne, Xiaoqian ; Boshoven, J.C. ; Rövenich, H.J. ; Damme, M.M.A. van; Li, J. ; Rojas Padilla, J.E. ; Song, Y. ; Valkenburg, D.J. ; Berg, G.C.M. van den; Thomma, B.P.H.J. - \ 2015
In: Book of Abstracts 28th Fungal Genetics Conference. - - p. 4 - 4.
Fungi cause severe crop losses and threaten food security worldwide. The soil-borne fungal pathogen Verticillium dahliae causes vascular wilt disease on hundreds of plant species, and disease control is challenging because resistance in plants is relatively rare. Moreover, V. dahliae has a flexible genome allowing it to escape host immunity and maintain aggressiveness. So far, knowledge on mechanisms governing this genomic flexibility remains limited. Through comparative population genomics we have started to unravel mechanisms to establish the genomic diversity that is essential for adaptive genome co-evolution during the continued arms race with host plants. To this end, two V. dahliae genomes were assembled from telomere-to-telomere using long-read sequencing technology and optical mapping, and compared these to the genomes of other Verticillium spp., revealing a pre-speciation genome duplication event. Comparative genomics using the two finished V. dahliae genomes furthermore revealed recent segmental duplications that established lineage-specific regions. Interestingly, these regions are enriched for in planta-expressed effector genes encoding secreted proteins that enable host colonization, and thus contribute to the evolution of virulence. Our evidence suggests that error-prone homology-dependent DNA repair has caused genomic rearrangements, leading to extensive structural variations. Re-sequencing of additional strains showed that independent losses of genetic material favored the escape of host recognition and, likely, host specificity. We propose that evolution of V. dahliae is linked to segmental genome duplications mediated by improperly repaired DNA breaks.
The calcium-dependent protein kinase CPK28 buffers plant immunity and regulates BIK1 turnover
Monaghan, Jacqueline ; Matschi, Susanne ; Shorinola, Oluwaseyi ; Rovenich, Hanna ; Matei, Alexandra ; Segonzac, Cécile ; Malinovsky, Frederikke Gro ; Rathjen, John P. ; Maclean, Dan ; Romeis, Tina ; Zipfel, Cyril - \ 2014
Cell Host & Microbe 16 (2014)5. - ISSN 1931-3128 - p. 605 - 615.
Plant perception of pathogen-associated molecular patterns (PAMPs) triggers a phosphorylation relay leading to PAMP-triggered immunity (PTI). Despite increasing knowledge of PTI signaling, how immune homeostasis is maintained remains largely unknown. Here we describe a forward-genetic screen to identify loci involved in PTI and characterize the Arabidopsis calcium-dependent protein kinase CPK28 as a negative regulator of immune signaling. Genetic analyses demonstrate that CPK28 attenuates PAMP-triggered immune responses and antibacterial immunity. CPK28 interacts with and phosphorylates the plasma-membrane-associated cytoplasmic kinase BIK1, an important convergent substrate of multiple pattern recognition receptor (PRR) complexes. We find that BIK1 is rate limiting in PTI signaling and that it is continuously turned over to maintain cellular homeostasis. We further show that CPK28 contributes to BIK1 turnover. Our results suggest a negative regulatory mechanism that continually buffers immune signaling by controlling the turnover of this key signaling kinase.
|Evolution and biological function of effectors of the vascular wilt pathogen Verticillium dahliae
Berg, G.C.M. van den; Boshoven, J.C. ; Cook III, D.E. ; Damme, M.M.A. van; Ebert, M.K. ; Faino, L. ; Kombrink, A. ; Li, J. ; Rojas Padilla, J.E. ; Rövenich, H.J. ; Sánchez-Vallet, A. ; Seidl, M.F. ; Shi, X. ; Song, Y. ; Valkenburg, D.J. ; Thomma, B.P.H.J. - \ 2014
In: Book of Abstracts COST 2014 - 2nd Annual Conference of the SUSTAIN Action. - - p. 22 - 22.
Verticillium dahliae, causal agent of vascular wilt disease, is one of the most notorious plant pathogens of tomato. By comparative population genomics, we previously identified the race 1 specific effector that activates tomato immunity mediated by the cell surface receptor Ve1. Further comparative genomics revealed extensive genomic rearrangements between individual V. dahliae isolates that could be implicated in the occurrence of lineage-specific regions involved in niche adaptation and virulence. Remarkably, genes that reside in lineage-specific regions are over-represented during in planta expression. Moreover, in contrast to candidate effector genes that reside in the core genome, targeted disruption of several lineage-specific effector genes resulted in compromised virulence. One of the lineage-specific effectors that could be implicated in virulence on tomato is a LysM effector, a homolog of the Ecp6 effector of the foliar tomato pathogen Cladosporium fulvum that functions in suppression of chitin-triggered host immunity. Structural analysis revealed a novel mechanism for chitin binding by Ecp6 through intrachain LysM dimerization, leading to a composite binding site that binds chitin with ultra-high affinity. Considering the importance of lineage-specific regions for V. dahliae aggressiveness, detailed genomic information on the recombination sites is required to understand how lineage-specific regions arise. To this end, we re-sequenced a V. dahliae genome with PacBio technology, leading to a gapless assembly of eight complete chromosomes. Our efforts to identify genomic signatures at the recombination sites and the molecular mechanism(s) that establish chromosomal rearrangements will be discussed.
|Evolution and biological function of effectors of the vascular wilt pathogen Verticillium dahliae
Berg, G.C.M. van den; Boshoven, J.C. ; Cook III, D.E. ; Damme, M.M.A. van; Ebert, M.K. ; Faino, L. ; Kombrink, A. ; Li, J. ; Rojas Padilla, J.E. ; Rövenich, H. ; Sánchez-Vallet, A. ; Seidl, M.F. ; Shi, X. ; Song, Y. ; Valkenburg, D.J. ; Thomma, B.P.H.J. - \ 2014
In: Book of Abstracts XVI International Congress on Molecular Plant-Microbe Interactions. - - p. 17 - 17.
PL-1.2 - Verticillium dahliae, causal agent of vascular wilt disease, is one of the most notorious plant pathogens of tomato. By comparative population genomics, we previously identified the race 1 specific effector that activates tomato immunity mediated by the cell surface receptor Ve1. Further comparative genomics revealed extensive genomic rearrangements between individual V. dahliae isolates that could be implicated in the occurrence of lineage-specific regions involved in niche adaptation and virulence. Remarkably, genes that reside in lineage-specific regions are over-represented during in planta expression. Moreover, in contrast to candidate effector genes that reside in the core genome, targeted disruptionof several lineage-specific effector genes resulted in compromised virulence. One of the lineage-specific effectors that could be implicated in virulence on tomato is a LysM effector, a homolog of the Ecp6 effector of the foliar tomato pathogen Cladosporium fulvum that functions in suppression of chitin-triggered host immunity. Structural analysis revealed a novel mechanism for chitin binding by Ecp6 through intrachain LysM dimerization, leading to a composite binding site that binds chitin with ultra-high affinity. Considering the importance of lineage-specific regions for V. dahliae aggressiveness, detailed genomic information on the recombination sites is required to understand how lineage-specific regions arise. To this end, we re-sequenced a V. dahliae genome with PacBio technology, leading to a gapless assembly of eight complete chromosomes. Our efforts to identify genomic signatures at the recombination sites and the molecular mechanism(s) that establish chromosomal rearrangements will be discussed.
|Transcriptome sequencing: a powerful tool for the identification of avirulence effector genes from the tomato leaf mould pathogen Cladosporium fulvum
Mesarich, C.H. ; Ökmen, B. ; Burgt, I.A. van der; Griffiths, S.A. ; Wang, C. ; Rövenich, H. ; Beenen, H.G. ; Etalo, D.W. ; Joosten, M.H.A.J. ; Wit, P.J.G.M. de - \ 2014
In: Book of Abstracts XVI International Congress on Molecular Plant-Microbe Interactions. - - p. 74 - 74.
P278 - Leaf mould disease of tomato (Solanum lycopersicum) is caused by the biotrophic fungal pathogen Cladosporium fulvum. Upon leaf entry, C. fulvum secretes a plethora of effector proteins to modulate host defence responses and promote successful infection. However, these effectors also trigger defence in tomato cultivars carrying cognate Cf disease resistance genes (avirulence effectors). Consequently, effector identification and functional characterization is required to further understand how C. fulvum facilitates infection, as well as the mechanisms involved in Cf-mediated effector perception and defence activation. Here, a combined bioinformatic and transcriptome sequencing approach was employed to identify avirulence effect or genes of C. fulvum. Firstly, we sought to clone the Avr5 avirulence effector gene, corresponding to the previously cloned Cf-5 resistance gene of tomato. RNA-Seq was performed on the sequenced race 0 strain (0WU; carries the Avr5 gene), as well as a race 5 strain (IPO 1979; lacks a functional Avr5 gene), during infection of susceptible tomato. Forty-four in planta-induced C. fulvum Candidate Effector (CfCE) genes of 0WU were identified that putatively encode a secreted, small cysteine-rich protein. An expressed transcript sequence comparison between strains revealed two polymorphic CfCE genes in IPO 1979. One of these conferred avirulence to IPO 1979 on Cf-5 tomato following complementation with the corresponding 0WU allele, confirming identification of Avr5. The remaining CfCE proteins were then tested for their ability to elicit a hypersensitive response upon transient expression in wild tomato species containing unknown resistance traits. Using this approach, several potentially novel avirulence determinants of C. fulvum were identified.
Filamentous pathogen effector functions: of pathogens, hosts and microbiomes
Rövenich, H. ; Boshoven, J.C. ; Thomma, B. - \ 2014
Current Opinion in Plant Biology 20 (2014). - ISSN 1369-5266 - p. 96 - 103.
chitin-triggered immunity - secreted fungal effector - potato famine pathogen - cladosporium-fulvum - protease inhibitor - magnaporthe-oryzae - plant-pathogens - genome evolution - tomato - virulence
Microorganisms play essential roles in almost every environment on earth. For instance, microbes decompose organic material, or establish symbiotic relationships that range from pathogenic to mutualistic. Symbiotic relationships have been particularly well studied for microbial plant pathogens and have emphasized the role of effectors; secreted molecules that support host colonization. Most effectors characterized thus far play roles in deregulation of host immunity. Arguably, however, pathogens not only deal with immune responses during host colonization, but also encounter other microbes including competitors, (myco)parasites and even potential co-operators. Thus, part of the effector catalog may target microbiome co-inhabitants rather than host physiology.
Functional analysis of the tomato immune receptor Ve1 through domain swaps with Its non-functional homolog Ve2
Fradin, E.F. ; Zhang, Z. ; Rövenich, H. ; Song, Y. ; Liebrand, T.W.H. ; Masini, L. ; Berg, G.C.M. van den; Joosten, M.H.A.J. ; Thomma, B.P.H.J. - \ 2014
PLoS ONE 9 (2014)2. - ISSN 1932-6203 - 14 p.
leucine-rich repeat - disease resistance protein - cladosporium-fulvum - arabidopsis-thaliana - endoplasmic-reticulum - hypersensitive response - verticillium resistance - plasma-membrane - innate immunity - kinase bri1
Resistance in tomato against race 1 strains of the fungal vascular wilt pathogens Verticillium dahliae and V. albo-atrum is mediated by the Ve locus. This locus comprises two closely linked inversely oriented genes, Ve1 and Ve2, which encode cell surface receptors of the extracellular leucine-rich repeat receptor-like protein (eLRR-RLP) type. While Ve1 mediates Verticillium resistance through monitoring the presence of the recently identified V. dahliae Ave1 effector, no functionality for Ve2 has been demonstrated in tomato. Ve1 and Ve2 contain 37 eLRRs and share 84% amino acid identity, facilitating investigation of Ve protein functionality through domain swapping. In this study it is shown that Ve chimeras in which the first thirty eLRRs of Ve1 were replaced by those of Ve2 remain able to induce HR and activate Verticillium resistance, and that deletion of these thirty eLRRs from Ve1 resulted in loss of functionality. Also the region between eLRR30 and eLRR35 is required for Ve1-mediated resistance, and cannot be replaced by the region between eLRR30 and eLRR35 of Ve2. We furthermore show that the cytoplasmic tail of Ve1 is required for functionality, as truncation of this tail results in loss of functionality. Moreover, the C-terminus of Ve2 fails to activate immune signaling as chimeras containing the C-terminus of Ve2 do not provide Verticillium resistance. Furthermore, Ve1 was found to interact through its C-terminus with the eLRR-containing receptor-like kinase (eLRR-RLK) interactor SOBIR1 that was recently identified as an interactor of eLRR-RLP (immune) receptors. Intriguingly, also Ve2 was found to interact with SOBIR1.
|The tomato receptor-like protein Ve1 directly interacts with the fungal effector Ave1 to initiate immune signaling
Rövenich, H.J. ; Zhang, Z. ; Fradin, E.F. ; Thomma, B.P.H.J. - \ 2013
In: Book of Abstracts Keystone Symposia on Molecular and Cellular Biology, Big Sky, Montana, USA, 7-12 April 2013. - - p. 58 - 58.
|Comparative pathogenomics of Verticillium dahliae
Thomma, B.P.H.J. ; Berg, G.C.M. van den; Boshoven, J.C. ; Damme, M. van; Esse, H.P. van; Faino, L. ; Jonge, R. de; Keykha Saber, M. ; Kombrink, A. ; Yanes Paz, E. ; Rövenich, H.J. ; Sanchez Vallet, A. ; Santhanam, P. ; Song, Y. ; Valkenburg, D.J. ; Zhang, Z. - \ 2013
In: Book of Abstracts 11th International Verticillium Symposium, Göttingen, Germany, 5-8 May 2013. - - p. 38 - 38.
The tomato immune receptor Ve1 governs resistance to race 1 strains of Verticillium, while race 2 strains are not recognized. Until recently, the Verticillium effector that is detected by Ve1 remained unknown. By high-throughput population genome sequencing, the gene that encodes the Ave1 effector (for Avirulence on Ve1 tomato) was identified. Interestingly, Ave1 homologs were also found in the fungal pathogens Cercospora beticola, Colletotrichum higginsianum and Fusarium oxysporum f. sp. lycopersici, some of which are recognized by Ve1. Based on the differential recognition of the Ave1 homologs, the epitope of the Ave1 protein has been identified. The identification of Ave1 facilitates functional analysis of the Ve1 immune receptor. Strictly asexual microorganisms, such as V. dahliae, are often considered as evolutionary dead ends as it remains unknown how they can generate the genetic variation that is required for coevolution with their hosts. Based on comparative population genomics we show that extensive chromosomal rearrangements establish highly dynamic ‘plastic’ genomic regions that act as a source for genetic variation to mediate aggressiveness and that are enriched for in planta-expressed effector genes. We propose that V. dahliae evolves by prompting chromosomal rearrangements, enabling rapid development of novel effector genes.