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Author: search.filters.author.Druzhinina, Irina S.

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    Genome of Pythium myriotylum Uncovers an Extensive Arsenal of Virulence-Related Genes among the Broad-Host-Range Necrotrophic Pythium Plant Pathogens
    (AMER SOC MICROBIOLOGY, 2022-01-01) Daly, Paul; Zhou, Dongmei; Shen, Danyu; Chen, Yifan; Xue, Taiqiang; Chen, Siqiao; Zhang, Qimeng; Zhang, Jinfeng; McGowan, Jamie; Cai, Feng; Pang, Guan; Wang, Nan; Sheikh, Taha Majid Mahmood; Deng, Sheng; Li, Jingjing; Soykam, Huseyin Okan; Kara, Irem; Fitzpatrick, David A.; Druzhinina, Irina S.; Akcapinar, Gunseli Bayram; Wei, Lihui
    The Pythium (Peronosporales, Oomycota) genus includes devastating plant pathogens that cause widespread diseases and severe crop losses. Here, we have uncovered a far greater arsenal of virulence factor-related genes in the necrotrophic Pythium myriotylum than in other Pythium plant pathogens. The genome of a plant-virulent P. myriotylum strain (similar to 70 Mb and 19,878 genes) isolated from a diseased rhizome of ginger (Zingiber officinale) encodes the largest repertoire of putative effectors, proteases, and plant cell wall-degrading enzymes (PCWDEs) among the studied species. P. myriotylum has twice as many predicted secreted proteins than any other Pythium plant pathogen. Arrays of tandem duplications appear to be a key factor of the enrichment of the virulence factor-related genes in P. myriotylum. The transcriptomic analysis performed on two P. myriotylum isolates infecting ginger leaves showed that proteases were a major part of the upregulated genes along with PCWDEs, Nep1-like proteins (NLPs), and elicitin-like proteins. A subset of P. myriotylum NLPs were analyzed and found to have necrosis-inducing ability from agroinfiltration of tobacco (Nicotiana benthamiana) leaves. One of the heterologously produced infection-upregulated putative cutinases found in a tandem array showed esterase activity with preferences for longer-chain-length substrates and neutral to alkaline pH levels. Our results allow the development of science-based targets for the management of P. myriotylum-caused disease, as insights from the genome and transcriptome show that gene expansion of virulence factor-related genes play a bigger role in the plant parasitism of Pythium spp. than previously thought. IMPORTANCE Pythium species are oomycetes, an evolutionarily distinct group of filamentous fungus-like stramenopiles. The Pythium genus includes several pathogens of important crop species, e.g., the spice ginger. Analysis of our genome from the plant pathogen Pythium myriotylum uncovered a far larger arsenal of virulence factor-related genes than found in other Pythium plant pathogens, and these genes contribute to the infection of the plant host. The increase in the number of virulence factor-related genes appears to have occurred through the mechanism of tandem gene duplication events. Genes from particular virulence factor-related categories that were increased in number and switched on during infection of ginger leaves had their activities tested. These genes have toxic activities toward plant cells or activities to hydrolyze polymeric components of the plant. The research suggests targets to better manage diseases caused by P. myriotylum and prompts renewed attention to the genomics of Pythium plant pathogens.
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    COMPARATIVE PHYSIOCHEMICAL ANALYSIS OF HYDROPHOBINS PRODUCED IN ESCHERICHIA COLI AND PICHIA PASTORIS
    (ELSEVIER, 2017-01-01) Przylucka, Agnes; Akcapinar, Gunseli Bayram; Bonazza, Klaus; Mello-de-Sousa, Thiago M.; Mach-Aigner, Astrid R.; Lobanov, Victor; Grothe, Hinrich; Kubicek, Christian P.; Reimhult, Erik; Druzhinina, Irina S.
    Hydrophobins (HFBs) are small surface-active proteins secreted by filamentous fungi. Being amphiphilic, they spontaneously form layers that convert surfaces from hydrophilic to hydrophobic and vice versa. We have compared properties of the class II HFB4 and HFB7 from Trichoderma virens as produced in Escherichia coli and Pichiapastoris. Since the production in E. coli required denaturation/renaturation steps because of inclusion bodies, this treatment was also applied to HFBs produced and secreted in yeast. The protein yields for both systems were similar. Both HFBs produced by E. coli proved less active on PET compared to HFBs produced in P. pastoris. HFBs produced in E. coli decreased the hydrophilicity of glass the most, which correlated with the adsorption of a more dense protein layer on glass compared to HFBs produced in P. pastoris. The hydrophobins produced in P. pastoris formed highly structured monolayers. Layers of hydrophobins produced in E. coli were less prone to self-organization. Our data suggests that irrespective of the production host, the HFBs could be used in various applications that are based on their surface activity. However, the production host and the subsequent purification procedure will influence the stability of HFB layers. In the area of high-value biomedical devices and nanomaterials, where the formation of highly ordered protein monolayers is essential, our results point to P. pastoris as the preferred production host. Furthermore, the choice of an appropriate hydrophobin for a given application appears to be equally important. (C) 2017 The Authors. Published by Elsevier B.V.
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    The pleiotropic functions of intracellular hydrophobins in aerial hyphae and fungal spores
    (PUBLIC LIBRARY SCIENCE, 2021-01-01) Cai, Feng; Zhao, Zheng; Gao, Renwei; Chen, Peijie; Ding, Mingyue; Jiang, Siqi; Fu, Zhifei; Xu, Pingyong; Chenthamara, Komal; Shen, Qirong; Akcapinar, Gunseli Bayram; Druzhinina, Irina S.
    Higher fungi can rapidly produce large numbers of spores suitable for aerial dispersal. The efficiency of the dispersal and spore resilience to abiotic stresses correlate with their hydrophobicity provided by the unique amphiphilic and superior surface-active proteins-hydrophobins (HFBs)- that self-assemble at hydrophobic/hydrophilic interfaces and thus modulate surface properties. Using the HFB-enriched mold Trichoderma (Hypocreales, Ascomycota) and the HFB-free yeast Pichia pastoris (Saccharomycetales, Ascomycota), we revealed that the rapid release of HFBs by aerial hyphae shortly prior to conidiation is associated with their intracellular accumulation in vacuoles and/or lipid-enriched organelles. The occasional internalization of the latter organelles in vacuoles can provide the hydrophobic/hydrophilic interface for the assembly of HFB layers and thus result in the formation of HFB-enriched vesicles and vacuolar multicisternal structures (VMSs) putatively lined up by HFBs. These HFB-enriched vesicles and VMSs can become fused in large tonoplast-like organelles or move to the periplasm for secretion. The tonoplast-like structures can contribute to the maintenance of turgor pressure in aerial hyphae supporting the erection of sporogenic structures (e.g., conidiophores) and provide intracellular force to squeeze out HFB-enriched vesicles and VMSs from the periplasm through the cell wall. We also show that the secretion of HFBs occurs prior to the conidiation and reveal that the even spore coating of HFBs deposited in the extracellular matrix requires microscopic water droplets that can be either guttated by the hyphae or obtained from the environment. Furthermore, we demonstrate that at least one HFB, HFB4 in T. guizhouense, is produced and secreted by wetted spores. We show that this protein possibly controls spore dormancy and contributes to the water sensing mechanism required for the detection of germination conditions. Thus, intracellular HFBs have a range of pleiotropic functions in aerial hyphae and spores and are essential for fungal development and fitness.
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    Evolutionary compromises in fungal fitness: hydrophobins can hinder the adverse dispersal of conidiospores and challenge their survival
    (SPRINGERNATURE, 2020-01-01) Cai, Feng; Gao, Renwei; Zhao, Zheng; Ding, Mingyue; Jiang, Siqi; Yagtu, Civan; Zhu, Hong; Zhang, Jian; Ebner, Thomas; Mayrhofer-Reinhartshuber, Michael; Kainz, Philipp; Chenthamara, Komal; Akcapinar, Guenseli Bayram; Shen, Qirong; Druzhinina, Irina S.
    Fungal evolutionary biology is impeded by the scarcity of fossils, irregular life cycles, immortality, and frequent asexual reproduction. Simple and diminutive bodies of fungi develop inside a substrate and have exceptional metabolic and ecological plasticity, which hinders species delimitation. However, the unique fungal traits can shed light on evolutionary forces that shape the environmental adaptations of these taxa. Higher filamentous fungi that disperse through aerial spores produce amphiphilic and highly surface-active proteins called hydrophobins (HFBs), which coat spores and mediate environmental interactions. We exploited a library of HFB-deficient mutants for two cryptic species of mycoparasitic and saprotrophic fungi from the genusTrichoderma(Hypocreales) and estimated fungal development, reproductive potential, and stress resistance. HFB4 and HFB10 were found to be relevant forTrichodermafitness because they could impact the spore-mediated dispersal processes and control other fitness traits. An analysis in silico revealed purifying selection for all cases except for HFB4 fromT. harzianum, which evolved under strong positive selection pressure. Interestingly, the deletion of thehfb4gene inT. harzianumconsiderably increased its fitness-related traits. Conversely, the deletion ofhfb4inT. guizhouenseled to the characteristic phenotypes associated with relatively low fitness. The net contribution of thehfb4gene to fitness was found to result from evolutionary tradeoffs between individual traits. Our analysis of HFB-dependent fitness traits has provided an evolutionary snapshot of the selective pressures and speciation process in closely related fungal species.
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    At least three families of hyphosphere small secreted cysteine-rich proteins can optimize surface properties to a moderately hydrophilic state suitable for fungal attachment
    (2022-01-01) Zhao, Zheng; Cai, Feng; Gao, Renwei; Ding, Mingyue; Jiang, Siqi; Chen, Peijie; Pang, Guan; Chenthamara, Komal; Shen, Qirong; Bayram Akcapinar, Gunseli; Druzhinina, Irina S.
    The secretomes of filamentous fungi contain a diversity of small secreted cysteine-rich proteins (SSCPs) that have a variety of properties ranging from toxicity to surface activity. Some SSCPs are recognized by other organisms as indicators of fungal presence, but their function in fungi is not fully understood. We detected a new family of fungal surface-active SSCPs (saSSCPs), here named hyphosphere proteins (HFSs). An evolutionary analysis of the HFSs in Pezizomycotina revealed a unique pattern of eight single cysteine residues (C-CXXXC-C-C-C-C-C) and a long evolutionary history of multiple gene duplications and ancient interfungal lateral gene transfers, suggesting their functional significance for fungi with different lifestyles. Interestingly, recombinantly produced saSSCPs from three families (HFSs, hydrophobins and cerato-platanins) showed convergent surface-modulating activity on glass and on poly(ethylene-terephthalate), transforming their surfaces to a moderately hydrophilic state, which significantly favoured subsequent hyphal attachment. The addition of purified saSSCPs to the tomato rhizosphere had mixed effects on hyphal attachment to roots, while all tested saSSCPs had an adverse effect on plant growth in vitro. We propose that the exceptionally high diversity of saSSCPs in Trichoderma and other fungi evolved to efficiently condition various surfaces in the hyphosphere to a fungal-beneficial state.