Biology subjects

Thomma, B.

Publications and source records attributed to Thomma, B..

35 records · Page 2Linked to original sources

bioRxiv2020

Fungal LysM effectors that comprise two LysM domains bind chitin through intermolecular dimerization

Chitin is a homopolymer of {beta}-(1,4)-linked N-acetyl-D-glucosamine (GlcNAc) and a major structural component of fungal cell walls. In plants, chitin acts as a microbe-associated molecular pattern (MAMP) that is recognized by lysin motif (LysM)-containing plant cell surface-localized pattern recognition receptors (PRRs) that activate a plethora of downstream immune responses. In order to deregulate chitin-induced plant immunity and successfully establish infection, many fungal pathogens secrete LysM domain-containing effector proteins during host colonization. It was previously shown that the LysM effector Ecp6 from the tomato leaf mould fungus Cladosporium fulvum can outcompete plant PRRs for chitin binding because two of its three LysM domains cooperate to form a composite groove with ultra-high (pM) chitin-binding affinity. However, most functionally characterized LysM effectors contain only two LysMs, including Magnaporthe oryzae MoSlp1, Verticillium dahliae Vd2LysM, and Colletotrichum higginsianum ChElp1 and ChElp2. Here, we performed modelling, structural and functional analyses to investigate whether such dual-domain LysM effectors can also form ultra-high chitin-binding affinity grooves through intramolecular LysM dimerization. However, our study suggests that intramolecular LysM dimerization does not occur. Rather, our data support the occurrence of intermolecular LysM dimerization for these effectors, associated with a significantly lower chitin binding affinity than monitored for Ecp6. Interestingly, the intermolecular LysM dimerization allows for the formation of polymeric complexes in the presence of chitin. Possibly, such polymers may precipitate at infection sites in order to eliminate chitin oligomers, and thus suppress the activation of chitin-induced plant immunity.

plant biology↗

bioRxiv2020

A plant pathogen utilizes effector proteins for microbiome manipulation

During colonization of their hosts, pathogens secrete effector proteins to promote disease development through various mechanisms. Increasing evidence shows that the host microbiome plays a crucial role in health, and that hosts actively shape their microbiomes to suppress disease. We hypothesized that pathogens evolved to manipulate host microbiomes to their advantage in turn. Here, we show that the fungal plant pathogen Verticillium dahliae utilizes effector proteins for niche colonization through selective manipulation of host microbiomes by suppressing microbes with antagonistic activities. Moreover, we show that effector proteins are similarly exploited for microbiome manipulation in the soil environment, where the fungus resides in absence of a host. In conclusion, we demonstrate that pathogens utilize effector proteins to modulate microbiome compositions and propose that their effector catalogs represent an untapped resource for novel antibiotics.

microbiology↗

bioRxiv2019

A secreted LysM effector protects fungal hyphae through chitin-dependent homodimer polymerization

Plants trigger immune responses upon recognition of fungal cell wall chitin, followed by the release of various antimicrobials, including chitinase enzymes that hydrolyze chitin. In turn, many fungal pathogens secrete LysM effectors that prevent chitin recognition by the host through scavenging of chitin oligomers. We previously showed that intrachain LysM dimerization of the Cladosporium fulvum effector Ecp6 confers an ultrahigh-affinity binding groove that competitively sequesters chitin oligomers from host immune receptors. Additionally, particular LysM effectors are found to protect fungal hyphae against chitinase hydrolysis during host colonization. However, the molecular basis for the protection of fungal cell walls against hydrolysis remained unclear. Here, we determined a crystal structure of the single LysM domain-containing effector Mg1LysM of the wheat pathogen Zymoseptoria tritici and reveal that Mg1LysM is involved in the formation of two kinds of dimers; a chitin-dependent dimer as well as a chitin-independent homodimer. In this manner, Mg1LysM gains the capacity to form a supramolecular structure by chitin-induced oligomerization of chitin-independent Mg1LysM homodimers, a property that confers protection to fungal cell walls against host chitinases.

microbiology↗

bioRxiv2019

Verticillium dahliae strains that infect the same host plant display highly divergent effector catalogs

Originality and significance statementDuring host colonization, plant pathogens secrete molecules that enable host colonization, also known as effector proteins. Here, we show that strains of the fungal plant pathogen Verticillium dahliae that are able to infect the same host plant harbour highly divergent LS effector repertoires. Our study outlines the variability within LS effector gene repertoires of V. dahliae strains, which may allow the various strains to be competitive in the co-evolution with their hosts. SummaryEffectors are proteins secreted by pathogens to support colonization of host plants, often by deregulating host immunity. Effector genes are often localized within dynamic lineage-specific (LS) genomic regions, allowing rapid evolution of effector catalogues. Such localization permits pathogens to be competitive in the co-evolutionary arms races with their hosts. For a broad host-range pathogen such as Verticillium dahliae it is unclear to what extent single members of their total effector repertoires contribute to disease development on multiple hosts. Here, we determined the core and LS effector repertoires of a collection of V. dahliae strains, as well as the ability of these strains to infect a range of plant species comprising tomato, cotton, Nicotiana benthamiana, Arabidopsis, and sunflower to assess whether the presence of particular LS effectors correlates with the ability to infect particular plant species. Surprisingly, we found that V. dahliae strains that are able to infect the same host plant harbor highly divergent LS effector repertoires. Furthermore, we observed differential V. dahliae core effector gene expression between host plants. Our data suggest that different V. dahliae lineages utilise divergent effector catalogs to colonize the same host plant, suggesting considerable redundancy among the activities of effector catalogs between lineages.

plant biology↗

bioRxiv2018

The genome of the fungal pathogen Verticillium dahliae reveals extensive bacterial to fungal gene transfer

Horizontal gene transfer (HGT) involves the transmission of genetic material between distinct evolutionary lineages and can be an important source of biological innovation. Reports of inter-kingdom HGT to eukaryotic microbial pathogens have accumulated over recent years. Verticillium dahliae is a notorious plant pathogen that causes vascular wilt disease on hundreds of plant species, resulting in high economic losses every year. Previously, the effector gene Ave1 and a glucosyltransferase-encoding gene were identified as virulence factor-encoding genes that were proposed to be horizontally acquired from a plant and a bacterial donor, respectively. However, to what extent HGT contributed to the overall genome composition of V. dahliae remained elusive. Here, we systematically searched for evidence of inter-kingdom HGT events in the genome of V. dahliae and provide evidence for extensive horizontal gene acquisition from bacterial origin.

microbiology↗

bioRxiv2018

In silico prediction and characterisation of secondary metabolite clusters in the plant pathogenic fungus Verticillium dahliae

Fungi are renowned producers of natural compounds, also known as secondary metabolites (SMs) that display a wide array of biological activities. Typically, the genes that are involved in the biosynthesis of SMs are located in close proximity to each other in so-called secondary metabolite clusters (SMCs). Many plant-pathogenic fungi secrete SMs during infection in order to promote disease establishment, for instance as cytocoxic compounds. Verticillium dahliae is a notorious plant pathogen that can infect over 200 host plants worldwide. However, the SM repertoire of this vascular pathogen remains mostly uncharted. To unravel the SM potential of V. dahliae, we performed in silico predictions and in-depth analyses of its SM clusters (SMC). We identified 25 potential SMCs in the V. dahliae genome, including loci that can be implicated in DHN-melanin, ferricrocin, triacetyl fusarinine and fujikurin production.

microbiology↗

bioRxiv2018

Gene cluster conservation identifies melanin and perylenequinone biosynthesis pathways in multiple plant pathogenic fungi

Perylenequinones are a family of structurally related polyketide fungal toxins with nearly universal toxicity. These photosensitizing compounds absorb light energy which enables them to generate reactive oxygen species that damage host cells. This potent mechanism serves as an effective weapon for plant pathogens in disease establishment. The sugar beet pathogen Cercospora beticola secretes the perylenequinone cercosporin during infection. We have shown recently that the cercosporin toxin biosynthesis (CTB) gene cluster is present in several other phytopathogenic fungi, prompting the search for biosynthetic gene clusters (BGCs) of structurally similar perylenequinones in other fungi. Here, we report the identification of the elsinochrome and phleichrome BGCs of Elsino[e] fawcettii and Cladosporium phlei, respectively, based on gene cluster conservation with the CTB and hypocrellin BGCs. Furthermore, we show that previously reported BGCs for elsinochrome and phleichrome are involved in melanin production. Phylogenetic analysis of the corresponding melanin polyketide synthases (PKSs) and alignment of melanin BGCs revealed high conservation between the established and newly identified C. beticola, E. fawcettii, and C. phlei melanin BGCs. Mutagenesis of the identified perylenequinone and melanin PKSs in C. beticola and E. fawcettii coupled with mass spectrometric metabolite analyses confirmed their roles in toxin and melanin production.\n\nOriginality and significance statementGenes involved in secondary metabolite (SM) production are often clustered together to form biosynthetic pathways. These pathways frequently have highly conserved keystone enzymes which can complicate allocation of a biosynthetic gene cluster (BGC) to the cognate SM. In our study, we utilized a combination of comparative genomics, phylogenetic analyses and biochemical approaches to reliably identify BGCs for perylenequinone toxins and DHN-melanin in multiple plant pathogenic fungi. Furthermore, we show that earlier studies that aimed to identify these perylenequinone pathways were misdirected and actually reported DHN-melanin biosynthetic pathways. Our study outlines a reliable approach to successfully identify fungal SM pathways.

microbiology↗

bioRxiv2018

Homogenization of sub-genome secretome gene expression patterns in the allodiploid fungus Verticillium longisporum

Hybridization is an important evolutionary mechanism that can enable organisms to adapt to environmental challenges. It has previously been shown that the fungal allodiploid species Verticillium longisporum, causal agent of Verticillium stem striping in rape seed, has originated from at least three independent hybridization events between two haploid Verticillium species. To reveal the impact of genome duplication as a consequence of the hybridization, we studied the genome and transcriptome dynamics upon two independent V. longisporum hybridization events, represented by the hybrid lineages "A1/D1" and "A1/D3". We show that the V. longisporum genomes are characterized by extensive chromosomal rearrangements, including between parental chromosomal sets. V. longisporum hybrids display signs of evolutionary dynamics that are typically associated with the aftermath of allodiploidization, such as haploidization and a more relaxed gene evolution. Expression patterns of the two sub-genomes within the two hybrid lineages are more similar than those of the shared A1 parent between the two lineages, showing that expression patterns of the parental genomes homogenized within a lineage. However, as genes that display differential parental expression in planta do not typically display the same pattern in vitro, we conclude that sub-genome-specific responses occur in both lineages. Overall, our study uncovers the genomic and transcriptomic plasticity during evolution of the filamentous fungal hybrid V. longisporum and illustrate its adaptive potential. ImportanceVerticillium is a genus of plant-associated fungi that include a handful of plant pathogens that collectively affect a wide range of hosts. On several occasions, haploid Verticillium species hybridized into the stable allodiploid species Verticillium longisporum, which is, in contrast to haploid Verticillium species, a Brassicaceae specialist. Here, we studied the evolutionary genome and transcriptome dynamics of V. longisporum and the impact of the hybridization. V. longisporum genomes display a mosaic structure due do genomic rearrangements between the parental chromosome sets. Similar to other allopolyploid hybrids, V. longisporum displays an ongoing loss of heterozygosity and a more relaxed gene evolution. Also, differential parental gene expression is observed, with an enrichment for genes that encode secreted proteins. Intriguingly, the majority of these genes displays sub-genome-specific responses under differential growth conditions. In conclusion, hybridization has incited the genomic and transcriptomic plasticity that enables adaptation to environmental changes in a parental allele-specific fashion.

microbiology↗

bioRxiv2018

Dynamic virulence-related regions of the fungal plant pathogen Verticillium dahliae display remarkably enhanced sequence conservation

Selection pressure impacts genomes unevenly, as different genes adapt with differential speed to establish an organisms optimal fitness. Plant pathogens co-evolve with their hosts, which implies continuously adaption to evade host immunity. Effectors are secreted proteins that mediate immunity evasion, but may also typically become recognized by host immune receptors. To facilitate effector repertoire alterations, in many pathogens, effector genes reside in dynamic genomic regions that are thought to display accelerated evolution, a phenomenon that is captured by the two-speed genome hypothesis. The genome of the vascular wilt pathogen Verticillium dahliae has been proposed to obey to a similar two-speed regime with dynamic, lineage-specific regions that are characterized by genomic rearrangements, increased transposable element activity and enrichment in in planta-induced effector genes. However, little is known of the origin of, and sequence diversification within, these lineage-specific regions. Based on comparative genomics among Verticillium spp. we now show differential sequence divergence between core and lineage-specific genomic regions of V. dahliae. Surprisingly, we observed that lineage-specific regions display markedly increased sequence conservation. Since single nucleotide diversity is reduced in these regions, host adaptation seems to be merely achieved through presence/absence polymorphisms. Increased sequence conservation of genomic regions important for pathogenicity is an unprecedented finding for filamentous plant pathogens and signifies the diversity of genomic dynamics in host-pathogen co-evolution.

microbiology↗

bioRxiv2018

Nuclear and mitochondrial genomes of the hybrid fungal plant pathogen Verticillium longisporum display a mosaic structure

Allopolyploidization, genome duplication through interspecific hybridization, is an important evolutionary mechanism that can enable organisms to adapt to environmental changes or stresses. This increased adaptive potential of allopolyploids can be particularly relevant for plant pathogens in their quest for host immune response evasion. Allodiploidization likely caused the shift in host range of the fungal pathogen plant Verticillium longisporum, as V. longisporum mainly infects Brassicaceae plants in contrast to haploid Verticillium spp. In this study, we investigated the allodiploid genome structure of V. longisporum and its evolution in the hybridization aftermath. The nuclear genome of V. longisporum displays a mosaic structure, as numerous contigs consists of sections of both parental origins. V. longisporum encountered extensive genome rearrangements, whereas the contribution of gene conversion is negligible. Thus, the mosaic genome structure mainly resulted from genomic rearrangements between parental chromosome sets. Furthermore, a mosaic structure was also found in the mitochondrial genome, demonstrating its bi-parental inheritance. In conclusion, the nuclear and mitochondrial genomes of V. longisporum parents interacted dynamically in the hybridization aftermath. Conceivably, novel combinations of DNA sequence of different parental origin facilitated genome stability after hybridization and consecutive niche adaptation of V. longisporum.

genomics↗

bioRxiv2017

Long Read Annotation (LoReAn): automated eukaryotic genome annotation based on long-read cDNA sequencing

Single-molecule full-length 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 LoReAn (Long Read Annotation) 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 and two plant genomes, we show that LoReAn outperforms popular annotation pipelines by integrating single-molecule cDNA sequencing data generated from either the PacBio or MinION sequencing platforms, and correctly predicting gene structure and capturing genes missed by other annotation pipelines.

bioinformatics↗

bioRxiv2017

Variable impact of Verticillium longisporum on oilseed rape yield in field trials in the United Kingdom

The Verticillium genus comprises economically important plant pathogens that collectively affect a broad range of annual and perennial crops. Verticillium longisporum mainly infects brassicaceous hosts, including oilseed rape. The most conspicuous symptom of V. longisporum infection on oilseed rape is black stem striping that appears towards the end of the cropping season. Thus far, the impact of V. longisporum infection on oilseed yield remains unclear. Verticillium stem striping recently emerged as a new disease in UK and currently displays a widespread occurrence in England. In this study, we assessed the impact of Verticillium stem striping on British oilseed rape production. To this end, four cultivars (Incentive, Vision, Harper and Quartz) were grown in field plots with different levels of V. longisporum disease pressure at different locations over two consecutive years. Whereas Incentive and Vision developed relatively few stem striping symptoms, Harper and especially Quartz showed severe symptoms during these field experiments. Furthermore, higher inoculum levels induced more severe symptoms in these cultivars. Intriguingly, significant yield reductions upon V. longisporum infection only occurred in a single field trial on all tested oilseed rape cultivars. Thus we conclude that Verticillium stem striping does not consistently impact oilseed rape yield, despite the occurrence of abundant disease symptoms.

plant biology↗

bioRxiv2017

Evolution within the fungal genus Verticillium is characterized by chromosomal rearrangement and gene loss

The fungal genus Verticillium contains ten species, some of which are notorious plant pathogens causing vascular wilt diseases in host plants, while others are known as saprophytes and opportunistic plant pathogens. Whereas the genome of V. dahliae, the most notorious plan pathogen of the genus, has been well characterized, evolution and speciation of other members of the genus received little attention thus far. Here, we sequenced the genomes of the nine haploid Verticillium spp. to study evolutionary trajectories of their divergence from a last common ancestor. Frequent occurrence of chromosomal rearrangement and gene family loss was identified. In addition to ~11,000 core genes that are shared among all species, only 200-600 species-specific genes occur. Intriguingly, these species-specific genes show different features than core genes.

microbiology↗

bioRxiv2017

The emerging British Verticillium longisporum population consists of aggressive Brassica pathogens

The impact of diseases depends on the dynamic interplay between host, pathogen and the environment. Newly emerging diseases may be the consequence of novel pathogen introductions that are typically associated with unpredictable outcomes, as their interaction with the host in a novel environment is unprecedented. Alternatively, new diseases may emerge from latent, previously established, pathogen populations that are triggered by changes in environmental factors like weather, agricultural practices and ecosystem management. Verticillium stem striping recently emerged in British oilseed rape (Brassica napus) production from a latent Verticillium longisporum population. V. longisporum is a hybrid fungal pathogen consisting of three lineages, each representing a separate hybridization event. Despite its prevalence, little is known of the pathogenicity of the British V. longisporum population. In this study, the pathogenicity of British isolates was tested on four different cultivars of three different Brassica crop species as well as on the model plant Arabidopsis thaliana and compared with previously characterized V. longisporum strains from other regions of the world, including representatives of all three hybrid lineages. Intriguingly, the British isolates appeared to be amongst the most pathogenic strains on Brassica crops. In conclusion, Verticillium stem striping poses a genuine threat to oilseed rape production as the British V. longisporum population consists of aggressive pathogens that have the potential to significantly impact Brassica crops.

plant biology↗

bioRxiv2017

Tomato immune receptor Ve1 recognizes surface-exposed co-localized N- and C-termini of Verticillium dahliae effector Ave1

Effectors are secreted by plant pathogens to facilitate infection, often through deregulation of host immune responses. During host colonization, race 1 strains of the soil-borne vascular wilt fungus Verticillium dahliae secrete the effector protein Ave1 that triggers immunity in tomato genotypes that encode the Ve1 immune receptor. Homologs of V. dahliae Ave1 (VdAve1) are found in plants and in few plant pathogenic microbes, and are differentially recognized by Ve1. However, how VdAve1 is recognized by Ve1 remained unknown. Interestingly, C-terminally affinity-tagged versions of VdAve1 failed to activate Ve1-mediated immunity, suggesting that exposure of the C-terminus of VdAve1 is required for Ve1-mediated recognition. This was confirmed by subsequent analysis of C-terminal deletion mutants, and by domain swap experiments. Although required, only the C-terminus of VdAve1 is not sufficient to activate Ve1-mediated immunity. Intriguingly, a three-dimensional structural model of VdAve1 revealed that the N- and C-termini co-localize on a surface-exposed patch of the VdAve1 protein. Indeed, subsequent analyses of N-terminal deletion mutants confirmed that also the N-terminus of VdAve1 is required to activate Ve1-mediated immunity. Thus, we conclude that a surface-exposed patch of the VdAve1 protein that is composed by co-localized N- and C-termini is recognized by the tomato immune receptor Ve1.

plant biology↗

bioRxiv2017

A previously established divergent lineage of the hybrid fungal pathogen Verticillium longisporum emerges as stem striping pathogen in British oilseed rape

Population genetic structures illustrate evolutionary trajectories of organisms adapting to differential environmental conditions. Pathogen populations are typically shaped by co-evolution with their hosts through genetic co-structuring. Verticillium stem striping was mainly observed in continental Europe, but has recently emerged in other countries including the United Kingdom. The disease is caused by the hybrid fungal species Verticilliumlongisporum that originates from at least three separate hybridization events, yet strains from the hybridization event between Verticillium progenitor species A1 and D1 are mainly responsible for Verticillium stem striping. By using multi-locus genotype analysis, we reveal a hitherto un-described dichotomy within V. longisporum lineage A1/D1 that correlates with the geographic distribution of the isolates with an \"A1/D1 West\" and an \"A1/D1 East\" cluster according to their relative location in Europe. Genome comparison between representatives of the A1/D1 West and East clusters confirmed mutual common origin, excluding distinctiveness through separate hybridization events. The A1/D1 West population is responsible for the sudden emergence of Verticillium stem striping in the UK. Remarkably, this emergence is caused by a British V. longisporum population that is genetically more diverse than the entire A1/D1 East cluster. Conceivably, V. longisporum has previously established in the UK, but remained latent or undiagnosed as an oilseed rape pathogen until recently. This finding illustrates that a recent introduction is not a prerequisite for a pathogen to emerge, as environmental factors and cultural practices can also play a pivotal role in outbreaks of novel diseases.

microbiology↗

bioRxiv2016

Verticillium dahliae LysM effectors differentially contribute to virulence on plant hosts

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.

plant biology↗