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1.
Exploiting Structural Modelling Tools to Explore Host-Translocated Effector Proteins.
Amoozadeh, S, Johnston, J, Meisrimler, CN
International journal of molecular sciences. 2021;(23)
Abstract
Oomycete and fungal interactions with plants can be neutral, symbiotic or pathogenic with different impact on plant health and fitness. Both fungi and oomycetes can generate so-called effector proteins in order to successfully colonize the host plant. These proteins modify stress pathways, developmental processes and the innate immune system to the microbes' benefit, with a very different outcome for the plant. Investigating the biological and functional roles of effectors during plant-microbe interactions are accessible through bioinformatics and experimental approaches. The next generation protein modeling software RoseTTafold and AlphaFold2 have made significant progress in defining the 3D-structure of proteins by utilizing novel machine-learning algorithms using amino acid sequences as their only input. As these two methods rely on super computers, Google Colabfold alternatives have received significant attention, making the approaches more accessible to users. Here, we focus on current structural biology, sequence motif and domain knowledge of effector proteins from filamentous microbes and discuss the broader use of novel modelling strategies, namely AlphaFold2 and RoseTTafold, in the field of effector biology. Finally, we compare the original programs and their Colab versions to assess current strengths, ease of access, limitations and future applications.
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2.
Regulators of commensal and pathogenic life-styles of an opportunistic fungus-Candida albicans.
Rai, LS, Wijlick, LV, Bougnoux, ME, Bachellier-Bassi, S, d'Enfert, C
Yeast (Chichester, England). 2021;(4):243-250
Abstract
The yeast Candida albicans is primarily a commensal of humans that colonizes the mucosal surfaces of the gastrointestinal and genital tracts. Yet, C. albicans can under certain circumstances undergo a shift from commensalism to pathogenicity. This transition is governed by fungal factors such as morphological transitions, environmental cues for instance relationships with gut microbiota and the host immune system. C. albicans utilizes distinct sets of regulatory programs to colonize or infect its host and to evade the host defense systems. Moreover, an orchestrated iron acquisition mechanism operates to adapt to specific niches with variable iron availability. Studies on regulatory networks and morphogenesis of these two distinct modes of C. albicans growth, suggest that both yeast and hyphal forms exist in both growth patterns and the regulatory circuits are inter-connected. Here, we summarize current knowledge about C. albicans commensal-to-pathogen shift, its regulatory elements and their contribution to human disease.
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3.
Investigating the cell and developmental biology of plant infection by the rice blast fungus Magnaporthe oryzae.
Eseola, AB, Ryder, LS, Osés-Ruiz, M, Findlay, K, Yan, X, Cruz-Mireles, N, Molinari, C, Garduño-Rosales, M, Talbot, NJ
Fungal genetics and biology : FG & B. 2021;:103562
Abstract
Magnaporthe oryzae is the causal agent of rice blast disease, the most widespread and serious disease of cultivated rice. Live cell imaging and quantitative 4D image analysis have provided new insight into the mechanisms by which the fungus infects host cells and spreads rapidly in plant tissue. In this video review article, we apply live cell imaging approaches to understanding the cell and developmental biology of rice blast disease. To gain entry to host plants, M. oryzae develops a specialised infection structure called an appressorium, a unicellular dome-shaped cell which generates enormous turgor, translated into mechanical force to rupture the leaf cuticle. Appressorium development is induced by perception of the hydrophobic leaf surface and nutrient deprivation. Cargo-independent autophagy in the three-celled conidium, controlled by cell cycle regulation, is essential for appressorium morphogenesis. Appressorium maturation involves turgor generation and melanin pigment deposition in the appressorial cell wall. Once a threshold of turgor has been reached, this triggers re-polarisation which requires regulated generation of reactive oxygen species, to facilitate septin GTPase-dependent cytoskeletal re-organisation and re-polarisation of the appressorium to form a narrow, rigid penetration peg. Infection of host tissue requires a further morphogenetic transition to a pseudohyphal-type of growth within colonised rice cells. At the same time the fungus secretes an arsenal of effector proteins to suppress plant immunity. Many effectors are secreted into host cells directly, which involves a specific secretory pathway and a specialised structure called the biotrophic interfacial complex. Cell-to-cell spread of the fungus then requires development of a specialised structure, the transpressorium, that is used to traverse pit field sites, allowing the fungus to maintain host cell membrane integrity as new living plant cells are invaded. Thereafter, the fungus rapidly moves through plant tissue and host cells begin to die, as the fungus switches to necrotrophic growth and disease symptoms develop. These morphogenetic transitions are reviewed in the context of live cell imaging studies.
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4.
Suppression or Activation of Immune Responses by Predicted Secreted Proteins of the Soybean Rust Pathogen Phakopsora pachyrhizi.
Qi, M, Grayczyk, JP, Seitz, JM, Lee, Y, Link, TI, Choi, D, Pedley, KF, Voegele, RT, Baum, TJ, Whitham, SA
Molecular plant-microbe interactions : MPMI. 2018;(1):163-174
Abstract
Rust fungi, such as the soybean rust pathogen Phakopsora pachyrhizi, are major threats to crop production. They form specialized haustoria that are hyphal structures intimately associated with host-plant cell membranes. These haustoria have roles in acquiring nutrients and secreting effector proteins that manipulate host immune systems. Functional characterization of effector proteins of rust fungi is important for understanding mechanisms that underlie their virulence and pathogenicity. Hundreds of candidate effector proteins have been predicted for rust pathogens, but it is not clear how to prioritize these effector candidates for further characterization. There is a need for high-throughput approaches for screening effector candidates to obtain experimental evidence for effector-like functions, such as the manipulation of host immune systems. We have focused on identifying effector candidates with immune-related functions in the soybean rust fungus P. pachyrhizi. To facilitate the screening of many P. pachyrhizi effector candidates (named PpECs), we used heterologous expression systems, including the bacterial type III secretion system, Agrobacterium infiltration, a plant virus, and a yeast strain, to establish an experimental pipeline for identifying PpECs with immune-related functions and establishing their subcellular localizations. Several PpECs were identified that could suppress or activate immune responses in nonhost Nicotiana benthamiana, N. tabacum, Arabidopsis, tomato, or pepper plants.
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5.
The Elicitor Protein AsES Induces a Systemic Acquired Resistance Response Accompanied by Systemic Microbursts and Micro-Hypersensitive Responses in Fragaria ananassa.
Hael-Conrad, V, Perato, SM, Arias, ME, Martínez-Zamora, MG, Di Peto, PLÁ, Martos, GG, Castagnaro, AP, Díaz-Ricci, JC, Chalfoun, NR
Molecular plant-microbe interactions : MPMI. 2018;(1):46-60
Abstract
The elicitor AsES (Acremonium strictum elicitor subtilisin) is a 34-kDa subtilisin-like protein secreted by the opportunistic fungus Acremonium strictum. AsES activates innate immunity and confers resistance against anthracnose and gray mold diseases in strawberry plants (Fragaria × ananassa Duch.) and the last disease also in Arabidopsis. In the present work, we show that, upon AsES recognition, a cascade of defense responses is activated, including: calcium influx, biphasic oxidative burst (O2⋅- and H2O2), hypersensitive cell-death response (HR), accumulation of autofluorescent compounds, cell-wall reinforcement with callose and lignin deposition, salicylic acid accumulation, and expression of defense-related genes, such as FaPR1, FaPG1, FaMYB30, FaRBOH-D, FaRBOH-F, FaCHI23, and FaFLS. All these responses occurred following a spatial and temporal program, first induced in infiltrated leaflets (local acquired resistance), spreading out to untreated lateral leaflets, and later, to distal leaves (systemic acquired resistance). After AsES treatment, macro-HR and macro-oxidative bursts were localized in infiltrated leaflets, while micro-HRs and microbursts occurred later in untreated leaves, being confined to a single cell or a cluster of a few epidermal cells that differentiated from the surrounding ones. The differentiated cells initiated a time-dependent series of physiological and anatomical changes, evolving to idioblasts accumulating H2O2 and autofluorescent compounds that blast, delivering its content into surrounding cells. This kind of systemic cell-death process in plants is described for the first time in response to a single elicitor. All data presented in this study suggest that AsES has the potential to activate a wide spectrum of biochemical and molecular defense responses in F. ananassa that may explain the induced protection toward pathogens of opposite lifestyle, like hemibiotrophic and necrotrophic fungi.
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6.
Amyloid-Like β-Aggregates as Force-Sensitive Switches in Fungal Biofilms and Infections.
Lipke, PN, Klotz, SA, Dufrene, YF, Jackson, DN, Garcia-Sherman, MC
Microbiology and molecular biology reviews : MMBR. 2018;(1)
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Abstract
Cellular aggregation is an essential step in the formation of biofilms, which promote fungal survival and persistence in hosts. In many of the known yeast cell adhesion proteins, there are amino acid sequences predicted to form amyloid-like β-aggregates. These sequences mediate amyloid formation in vitro. In vivo, these sequences mediate a phase transition from a disordered state to a partially ordered state to create patches of adhesins on the cell surface. These β-aggregated protein patches are called adhesin nanodomains, and their presence greatly increases and strengthens cell-cell interactions in fungal cell aggregation. Nanodomain formation is slow (with molecular response in minutes and the consequences being evident for hours), and strong interactions lead to enhanced biofilm formation. Unique among functional amyloids, fungal adhesin β-aggregation can be triggered by the application of physical shear force, leading to cellular responses to flow-induced stress and the formation of robust biofilms that persist under flow. Bioinformatics analysis suggests that this phenomenon may be widespread. Analysis of fungal abscesses shows the presence of surface amyloids in situ, a finding which supports the idea that phase changes to an amyloid-like state occur in vivo. The amyloid-coated fungi bind the damage-associated molecular pattern receptor serum amyloid P component, and there may be a consequential modulation of innate immune responses to the fungi. Structural data now suggest mechanisms for the force-mediated induction of the phase change. We summarize and discuss evidence that the sequences function as triggers for protein aggregation and subsequent cellular aggregation, both in vitro and in vivo.
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7.
Dynamic Fungal Cell Wall Architecture in Stress Adaptation and Immune Evasion.
Hopke, A, Brown, AJP, Hall, RA, Wheeler, RT
Trends in microbiology. 2018;(4):284-295
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Abstract
Deadly infections from opportunistic fungi have risen in frequency, largely because of the at-risk immunocompromised population created by advances in modern medicine and the HIV/AIDS pandemic. This review focuses on dynamics of the fungal polysaccharide cell wall, which plays an outsized role in fungal pathogenesis and therapy because it acts as both an environmental barrier and as the major interface with the host immune system. Human fungal pathogens use architectural strategies to mask epitopes from the host and prevent immune surveillance, and recent work elucidates how biotic and abiotic stresses present during infection can either block or enhance masking. The signaling components implicated in regulating fungal immune recognition can teach us how cell wall dynamics are controlled, and represent potential targets for interventions designed to boost or dampen immunity.
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8.
Leptosphaeria maculans effector AvrLm4-7 affects salicylic acid (SA) and ethylene (ET) signalling and hydrogen peroxide (H2 O2 ) accumulation in Brassica napus.
Nováková, M, Šašek, V, Trdá, L, Krutinová, H, Mongin, T, Valentová, O, Balesdent, MH, Rouxel, T, Burketová, L
Molecular plant pathology. 2016;(6):818-31
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Abstract
To achieve host colonization, successful pathogens need to overcome plant basal defences. For this, (hemi)biotrophic pathogens secrete effectors that interfere with a range of physiological processes of the host plant. AvrLm4-7 is one of the cloned effectors from the hemibiotrophic fungus Leptosphaeria maculans 'brassicaceae' infecting mainly oilseed rape (Brassica napus). Although its mode of action is still unknown, AvrLm4-7 is strongly involved in L. maculans virulence. Here, we investigated the effect of AvrLm4-7 on plant defence responses in a susceptible cultivar of B. napus. Using two isogenic L. maculans isolates differing in the presence of a functional AvrLm4-7 allele [absence ('a4a7') and presence ('A4A7') of the allele], the plant hormone concentrations, defence-related gene transcription and reactive oxygen species (ROS) accumulation were analysed in infected B. napus cotyledons. Various components of the plant immune system were affected. Infection with the 'A4A7' isolate caused suppression of salicylic acid- and ethylene-dependent signalling, the pathways regulating an effective defence against L. maculans infection. Furthermore, ROS accumulation was decreased in cotyledons infected with the 'A4A7' isolate. Treatment with an antioxidant agent, ascorbic acid, increased the aggressiveness of the 'a4a7' L. maculans isolate, but not that of the 'A4A7' isolate. Together, our results suggest that the increased aggressiveness of the 'A4A7' L. maculans isolate could be caused by defects in ROS-dependent defence and/or linked to suppressed SA and ET signalling. This is the first study to provide insights into the manipulation of B. napus defence responses by an effector of L. maculans.
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The defence elicitor AsES causes a rapid and transient membrane depolarization, a triphasic oxidative burst and the accumulation of nitric oxide.
Martos, GG, Terán, Mdel M, Díaz Ricci, JC
Plant physiology and biochemistry : PPB. 2015;:443-50
Abstract
The newly characterized elicitor AsES obtained from Acremonium strictum induces a strong defence response in strawberry plants and confers plants resistance against the fungal pathogen Colletotricum acutatum the casual agent of anthracnose disease. Previous studies showed that AsES causes the accumulation of reactive oxygen species (ROS) that peaked 4 h post treatment (hpt), but due to the experimental approach used it was not clear whether the accumulation of ROS observed was intracellular or extracellular or took place as a single peak. By using a different experimental setup, a more complex early events associated to the activation of the innate immunity were observed. In this paper we report that strawberry plant cells treated with AsES exhibits a triphasic production of H2O2 and a rapid intracellular accumulation of NO. The first phase consists in a progressive extracellular accumulation of H2O2 that starts immediately after the treatment with AsES and is preceded by a rapid and transient cell membrane depolarization. During this phase takes place also a rapid intracellular accumulation of NO. Microscopic observations of mesophyll cells treated with AsES reveals that NO accumulates at the chloroplast. After the first extracellular H2O2 production phase, two intracellular H2O2 accumulation events occur, the first 2 hpt, and the second 7 hpt. Cells treated with AsES also show a transient increase of ion leakage, and a progressive alkalinization of the extracellular medium.
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10.
Functional annotation of putative hypothetical proteins from Candida dubliniensis.
Kumar, K, Prakash, A, Tasleem, M, Islam, A, Ahmad, F, Hassan, MI
Gene. 2014;(1):93-100
Abstract
An extensive analysis of C. dubliniensis proteomics data showed that ~22% protein are conserved hypothetical proteins (HPs) whose function is still not determined precisely. Analysis of gene sequence of HPs provides a platform to establish sequence-function relationships to a more profound understanding of the molecular machinery of organisms at systems level. Here we have combined the latest versions of bioinformatics tools including, protein family, motifs, intrinsic features from the amino acid sequence, sequence-function relationship, pathway analysis, etc. to assign a precise function to HPs for which no any experimental information is available. Our results show that 27 HPs have well defined functions and we categorized them as enzyme, nucleic acid binding, transport protein, etc. Five HPs showed adhesin character that is likely to be essential for the survival of yeast and pathogenesis. We also addressed issues related to the sub-cellular localization and signal peptide identification which provides an idea about its colocalization and function. The outcome of the present study may facilitate better understanding of mechanism of virulence, drug resistance, pathogenesis, adaptability to host, tolerance for host immune response, and drug discovery for treatment of C. dubliniensis infections.