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

ABSTRACTPlants 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.

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Mitochondrial Control of Fungal Cell Walls: Models and Relevance in Fungal Pathogens

Current Topics in Microbiology and Immunology - The Fungal Cell Wall ◽

10.1007/82_2019_183 ◽

2019 ◽

pp. 277-296 ◽

Cited By ~ 1

Author(s):

Barbara Koch ◽

Ana Traven

Keyword(s):

Cell Walls ◽

Fungal Pathogens ◽

Fungal Cell ◽

Fungal Cell Walls

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Bdelloid rotifers use hundreds of horizontally acquired genes against fungal pathogens

10.1101/2021.09.04.458992 ◽

2021 ◽

Author(s):

Reuben W Nowell ◽

Timothy G Barraclough ◽

Christopher G Wilson

Keyword(s):

Fungal Pathogen ◽

Cell Walls ◽

Fungal Pathogens ◽

Fungal Cell ◽

Bdelloid Rotifers ◽

Fungal Toxins ◽

Foreign Genes ◽

Fungal Cell Walls

Obligately asexual lineages are typically rare and short-lived. According to one hypothesis, they adapt too slowly to withstand relentlessly coevolving pathogens. Bdelloid rotifers seem to have avoided this fate, by enduring millions of years without males or sex. We investigated whether bdelloids' unusual capacity to acquire non-metazoan genes horizontally has enhanced their resistance to pathogens. We found that horizontally transferred genes are three times more likely than native genes to be upregulated in response to a natural fungal pathogen. This enrichment was twofold stronger than that elicited by a physical stressor (desiccation), and the genes showed little overlap. Among hundreds of upregulated non-metazoan genes were RNA ligases putatively involved in resisting fungal toxins and glucanases predicted to bind to fungal cell walls, acquired from bacteria. Our results provide evidence that bdelloids mitigate a predicted challenge of long-term asexuality in part through their ability to acquire and deploy so many foreign genes.

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The Chitin-Binding Cladosporium fulvum Effector Protein Avr4 Is a Virulence Factor

Molecular Plant-Microbe Interactions ◽

10.1094/mpmi-20-9-1092 ◽

2007 ◽

Vol 20 (9) ◽

pp. 1092-1101 ◽

Cited By ~ 147

Author(s):

H. Peter van Esse ◽

Melvin D. Bolton ◽

Ioannis Stergiopoulos ◽

Pierre J. G. M. de Wit ◽

Bart P. H. J. Thomma

Keyword(s):

Cell Walls ◽

Fungal Pathogens ◽

Cladosporium Fulvum ◽

Fungal Cell ◽

Pathogen Virulence ◽

Passalora Fulva ◽

Tomato Leaf Mold ◽

Defensive Activity ◽

Plant Chitinases ◽

Intrinsic Function

The biotrophic fungal pathogen Cladosporium fulvum (syn. Passalora fulva) is the causal agent of tomato leaf mold. The Avr4 protein belongs to a set of effectors that is secreted by C. fulvum during infection and is thought to play a role in pathogen virulence. Previous studies have shown that Avr4 binds to chitin present in fungal cell walls and that, through this binding, Avr4 can protect these cell walls against hydrolysis by plant chitinases. In this study, we demonstrate that Avr4 expression in Arabidopsis results in increased virulence of several fungal pathogens with exposed chitin in their cell walls, whereas the virulence of a bacterium and an oomycete remained unaltered. Heterologous expression of Avr4 in tomato increased the virulence of Fusarium oxysporum f. sp. lycopersici. Through tomato GeneChip analyses, we demonstrate that Avr4 expression in tomato results in the induced expression of only a few genes. Finally, we demonstrate that silencing of the Avr4 gene in C. fulvum decreases its virulence on tomato. This is the first report on the intrinsic function of a fungal avirulence protein that has a counter-defensive activity required for full virulence of the pathogen.

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Mannan detecting C-type lectin receptor probes recognise immune epitopes with diverse chemical, spatial and phylogenetic heterogeneity in fungal cell walls

10.1101/677104 ◽

2019 ◽

Author(s):

Ingrida Vendele ◽

Janet A. Willment ◽

Lisete M. Silva ◽

Angelina S. Palma ◽

Wengang Chai ◽

...

Keyword(s):

Cell Wall ◽

Immune Responses ◽

Cell Walls ◽

Fungal Pathogens ◽

Fungal Species ◽

Fungal Cell Wall ◽

Immune Recognition ◽

Cell Surfaces ◽

Fungal Cell ◽

Wall Components

AbstractDuring the course of fungal infection, pathogen recognition by the innate immune system is critical to initiate efficient protective immune responses. The primary event that triggers immune responses is the binding of Pattern Recognition Receptors (PRRs), which are expressed at the surface of host immune cells, to Pathogen-Associated Molecular Patterns (PAMPs) located predominantly in the fungal cell wall. Most fungi have mannosylated PAMPs in their cell walls and these are recognized by a range of C-type lectin receptors (CTLs). However, the precise spatial distribution of the ligands that induce immune responses within the cell walls of fungi are not well defined. We used recombinant IgG Fc-CTLs fusions of three murine mannan detecting CTLs, including dectin-2, the mannose receptor (MR) carbohydrate recognition domains (CRDs) 4-7 (CRD4-7), and human DC-SIGN (hDC-SIGN) and the β-1,3 glucan-binding lectin dectin-1 to map PRR ligands in the fungal cell wall. We show that epitopes of mannan-specific CTL receptors can be clustered or diffuse, superficial or buried in the inner cell wall. We demonstrate that PRR ligands do not correlate well with phylogenetic relationships between fungi, and that Fc-lectin binding discriminated between mannosides expressed on different cell morphologies of the same fungus. We also demonstrate CTL epitope differentiation during different phases of the growth cycle ofCandida albicansand that MR and DC-SIGN labelled outer chainN-mannans whilst dectin-2 labelled coreN-mannans displayed deeper in the cell wall. These immune receptor maps of fungal walls therefore reveal remarkable spatial, temporal and chemical diversity, indicating that the triggering of immune recognition events originates from multiple physical origins at the fungal cell surface.Author SummaryInvasive fungal infections remain an important health problem in immunocompromised patients. Immune recognition of fungal pathogens involves binding of specific cell wall components by pathogen recognition receptors (PRRs) and subsequent activation of immune defences. Some cell wall components are conserved among fungal species while other components are species-specific and phenotypically diverse. The fungal cell wall is dynamic and capable of changing its composition and organization when adapting to different growth niches and environmental stresses. Differences in the composition of the cell wall lead to differential immune recognition by the host. Understanding how changes in the cell wall composition affect recognition by PRRs is likely to be of major diagnostic and clinical relevance. Here we address this fundamental question using four soluble immune receptor-probes which recognize mannans and β-glucan in the cell wall. We use this novel methodology to demonstrate that mannan epitopes are differentially distributed in the inner and outer layers of fungal cell wall in a clustered or diffuse manner. Immune reactivity of fungal cell surfaces did not correlate with relatedness of different fungal species, and mannan-detecting receptor-probes discriminated between cell surface mannans generated by the same fungus growing under different conditions. These studies demonstrate that mannan-epitopes on fungal cell surfaces are differentially distributed within and between the cell walls of fungal pathogens.

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Dectin-1-Targeted Antifungal Liposomes Exhibit Enhanced Efficacy

mSphere ◽

10.1128/msphere.00025-19 ◽

2019 ◽

Vol 4 (1) ◽

Cited By ~ 1

Author(s):

Suresh Ambati ◽

Aileen R. Ferarro ◽

S. Earl Kang ◽

Jianfeng Lin ◽

Xiaorong Lin ◽

...

Keyword(s):

Invasive Aspergillosis ◽

Cell Walls ◽

Fungal Pathogens ◽

Antifungal Drugs ◽

Beta Glucan ◽

Fungal Cell ◽

Content Type ◽

Current Therapies ◽

Mammalian Protein ◽

Fungal Cell Walls

The fungus Aspergillus fumigatus causes pulmonary invasive aspergillosis resulting in nearly 100,000 deaths each year. Patients are often treated with antifungal drugs such as amphotericin B (AmB) loaded into liposomes (AmB-LLs), but all antifungal drugs, including AmB-LLs, have serious limitations due to human toxicity and insufficient fungal cell killing. Even with the best current therapies, 1-year survival among patients with invasive aspergillosis is only 25 to 60%. Hence, there is a critical need for improved antifungal therapeutics. Dectin-1 is a mammalian protein that binds to beta-glucan polysaccharides found in nearly all fungal cell walls. We coated AmB-LLs with Dectin-1 to make DEC-AmB-LLs. DEC-AmB-LLs bound strongly to fungal cells, while AmB-LLs had little affinity. DEC-AmB-LLs killed or inhibited A. fumigatus 10 times more efficiently than untargeted liposomes, decreasing the effective dose of AmB. Dectin-1-coated drug-loaded liposomes targeting fungal pathogens have the potential to greatly enhance antifungal therapeutics.

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Targeted antifungal liposomes

10.1101/518043 ◽

2019 ◽

Cited By ~ 1

Author(s):

Suresh Ambati ◽

Aileen R. Ferarro ◽

S. Earl Khang ◽

Xiaorong Lin ◽

Michelle Momany ◽

...

Keyword(s):

Invasive Aspergillosis ◽

Cell Walls ◽

Fungal Pathogens ◽

Antifungal Drugs ◽

Human Toxicity ◽

Beta Glucan ◽

Fungal Cell ◽

Order Of Magnitude ◽

One Year ◽

Fungal Cell Walls

AbstractAspergillus species cause pulmonary invasive aspergillosis resulting in nearly a hundred thousand deaths each year. Patients at the greatest risk of developing life-threatening aspergillosis have weakened immune systems and/or various lung disorders. Patients are treated with antifungals such as amphotericin B (AmB), casofungin acetate, or triazoles (itraconazole, voriconazole etc.), but these antifungal agents have serious limitations due to lack of sufficient fungicidal effect and human toxicity. Liposomes with AmB intercalated into the lipid membrane (AmBisomes, AmB-LLs), have several-fold reduced toxicity compared to detergent solubilized drug. However, even with the current antifungal therapies, one-year survival among patients is only 25 to 60%. Hence, there is a critical need for improved antifungal therapeutics.Dectin-1 is a mammalian innate immune receptor in the membrane of some leukocytes that binds as a dimer to beta-glucans found in fungal cell walls, signaling fungal infection. Using a novel protocol, we coated AmB-LLs with Dectin-1’s beta-glucan binding domain to make DEC-AmB-LLs. DEC-AmB-LLs bound rapidly, efficiently, and with great strength Aspergillus fumigatus and to Candida albicans and Cryptococcus neoformans, highly divergent fungal pathogens of global importance. By contrast, un-targeted AmB-LLs and BSA-coated BSA-AmB-LLs showed 200-fold lower affinity for fungal cells. DEC-AmB-LLs reduced the growth and viability of A. fumigatus an order of magnitude more efficiently than untargeted control liposomes delivering the same concentrations of AmB, in essence increasing the effective dose of AmB. Future efforts will focus on examining pan-antifungal targeted liposomal drugs in animal models of disease.TweetWe coated anti-fungal drug loaded liposomes to fungal cell walls with a beta-glucan binding protein and thereby increased drug effectiveness by an order of magnitude.ImportanceThe fungus Aspergillus fumigatus causes pulmonary invasive aspergillosis resulting in nearly a hundred thousand deaths each year. Patients are often treated with antifungal drugs such as amphotericin B loaded into liposomes, AmB-LLs, but all antifungal drugs including AmB-LLs have serious limitations due to human toxicity and insufficient fungal cell killing. Even with the best current therapies, one-year survival among patients with invasive aspergillosis is only 25 to 60%. Hence, there is a critical need for improved antifungal therapeutics.Dectin-1 is a mammalian protein that binds to beta-glucan polysaccharides found in nearly all fungal cell walls. We coated AmB-LLs with Dectin-1 to make DEC-AmB-LLs. DEC-AmB-LLs bond strongly to fungal cells, while AmB-LLs had little affinity. DEC-AmB-LLs killed or inhibited A. fumigatus ten times more efficiently than untargeted lipsomes, increasing the effective dose of AmB. Dectin-1 coated liposomes targeting fungal pathogens have the potential to greatly enhance antifungal therapeutics.

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Trimming Surface Sugars Protects Histoplasma from Immune Attack

mBio ◽

10.1128/mbio.00553-16 ◽

2016 ◽

Vol 7 (2) ◽

Cited By ~ 3

Author(s):

Gordon D. Brown

Keyword(s):

Cell Walls ◽

Fungal Pathogens ◽

A Fungus-Inducible Pepper Carboxylesterase Exhibits Antifungal Activity by Decomposing the Outer Layer of Fungal Cell Walls

Molecular Plant-Microbe Interactions ◽

10.1094/mpmi-11-17-0266-r ◽

2018 ◽

Vol 31 (5) ◽

pp. 505-515 ◽

Cited By ~ 3

Author(s):

Hyo-Hyoun Seo ◽

Ae Ran Park ◽

Hyun-Hwa Lee ◽

Sangkyu Park ◽

Yun-Jeong Han ◽

...

Keyword(s):

Antifungal Activity ◽

Cell Walls ◽

Fungal Pathogens ◽

Mass Spectrometry Analysis ◽

Gas Chromatography Mass Spectrometry ◽

Protective Effects ◽

Fungal Cell ◽

Spectrometry Analysis ◽

Hydrolysis Of ◽

Fungal Cell Walls

Colletotrichum species are major fungal pathogens that cause devastating anthracnose diseases in many economically important crops. In this study, we observed the hydrolyzing activity of a fungus-inducible pepper carboxylesterase (PepEST) on cell walls of C. gloeosporioides, causing growth retardation of the fungus by blocking appressorium formation. To determine the cellular basis for the growth inhibition, we observed the localization of PepEST on the fungus and found the attachment of the protein on surfaces of conidia and germination tubes. Moreover, we examined the decomposition of cell-wall materials from the fungal surface after reaction with PepEST, which led to the identification of 1,2-dithiane-4,5-diol (DTD) by gas chromatography mass spectrometry analysis. Exogenous DTD treatment did not elicit expression of defense-related genes in the host plant but did trigger the necrosis of C. gloeosporioides. Furthermore, the DTD compound displayed protective effects on pepper fruits and plants against C. gloeosporioides and C. coccodes, respectively. In addition, DTD was also effective in preventing other diseases, such as rice blast, tomato late blight, and wheat leaf rust. Therefore, our results provide evidence that PepEST is involved in hydrolysis of the outmost layer of the fungal cell walls and that DTD has antifungal activity, suggesting an alternative strategy to control agronomically important phytopathogens.

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