Biology subjects

Hassing, B.

Publications and source records attributed to Hassing, B..

4 recordsLinked to original sources

bioRxiv2022

The Venturia inaequalis effector repertoire is expressed in waves, and is dominated by expanded families with predicted structural similarity to avirulence proteins from other fungi

BackgroundScab, caused by the biotrophic fungus Venturia inaequalis, is the most economically important disease of apples worldwide. During infection, V. inaequalis occupies the subcuticular environment, where it secretes virulence factors, termed effectors, to promote host colonization. Consistent with other plant-pathogenic fungi, many of these effectors are expected to be non-enzymatic proteins, some of which can be recognized by corresponding host resistance proteins to activate plant defences, thus acting as avirulence determinants. To develop durable control strategies against scab, a better understanding of the roles that these effector proteins play in promoting subcuticular growth by V. inaequalis, as well as in activating, suppressing or circumventing resistance protein-mediated defences in apple, is required. ResultsWe generated the first comprehensive RNA-seq transcriptome of V. inaequalis during colonization of apple. Analysis of this transcriptome revealed five temporal waves of gene expression that peaked during early, mid or mid-late infection. While the number of genes encoding secreted, non-enzymatic proteinaceous effector candidates (ECs) varied in each wave, most belonged to waves that peaked in expression during mid-late infection. Spectral clustering based on sequence similarity determined that the majority of ECs belonged to expanded protein families. To gain insights into function, the tertiary structures of ECs were predicted using AlphaFold2. Strikingly, despite an absence of sequence similarity, many ECs were predicted to have structural similarity to avirulence proteins from other plant-pathogenic fungi, including members of the MAX, LARS, ToxA and FOLD effector families. In addition, several other ECs, including an EC family with sequence similarity to the AvrLm6 avirulence effector from Leptosphaeria maculans, were predicted to adopt a KP6-like fold. Thus, proteins with a KP6-like fold represent another structural family of effectors shared among plant-pathogenic fungi. ConclusionsOur study reveals the transcriptomic profile underpinning subcuticular growth by V. inaequalis and provides an enriched list of ECs that can be investigated for roles in virulence and avirulence. Furthermore, our study supports the idea that numerous sequence-unrelated effectors across plant-pathogenic fungi share common structural folds. In doing so, our study gives weight to the hypothesis that many fungal effectors evolved from ancestral genes through duplication, followed by sequence diversification, to produce sequence-unrelated but structurally similar proteins.

microbiology↗

bioRxiv2020

A homologue of the mammalian tumour suppressor protein PTEN is a functional lipid phosphatase and required for chemotaxis in filamentous fungi

Phosphoinositides (PI) are essential components of eukaryotic membranes and function in a large number of signalling processes. While lipid second messengers are well studied in mammals and yeast, their role in filamentous fungi is poorly understood. We used fluorescent PI-binding molecular probes to localise the phosphorylated phosphatidylinositol species PI[3]P, PI[3,5]P2, PI[4]P and PI[4,5]P2 in hyphae of the endophyte Epichloe festucae in axenic culture and during interaction with its grass host Lolium perenne. We also analysed the roles of the phosphatidylinositol-4-phosphate 5-kinase MssD and the predicted phosphatidylinositol-3,4,5-triphosphate 3-phosphatase TepA, a homologue of the mammalian tumour suppressor protein PTEN. Deletion of tepA in E. festucae and in the root-infecting tomato pathogen Fusarium oxysporum had no impact on growth in culture or the host interaction phenotype. However, this mutation did uncover the presence of PI[3,4,5]P3 in septa of E. festucae and showed that TepA is required for chemotropism in F. oxysporum. The identification of PI[3,4,5]P3 in septa of{Delta} tepA strains suggests that filamentous fungi are able to generate PI[3,4,5]P3 using an alternative biosynthetic pathway and that fungal PTEN homologues are functional lipid phosphatases. The F. oxysporum chemotropism defect demonstrates a conserved role of PTEN homologues in chemotaxis across protists, fungi and mammals.

cell biology↗

bioRxiv2019

Phosphatidic acid produced by phospholipase D is required for hyphal cell-cell fusion and fungal-plant symbiosis

Although lipid signaling has been shown to serve crucial roles in mammals and plants, little is known about this process in filamentous fungi. Here we analyse the contribution of phospholipase D (PLD) and its product phosphatidic acid (PA) in hyphal morphogenesis and growth of Epichloe festucae and Neurospora crassa, and in the establishment of a symbiotic interaction between E. festucae and Lolium perenne. Growth of E. festucae and N. crassa PLD deletion strains in axenic culture, and for E. festucae in association with L. perenne, were analysed by light-, confocal- and electron microscopy. Changes in PA distribution were analysed in E. festucae using a PA biosensor and the impact of these changes on endocytic recycling and superoxide production investigated. We found that E. festucae PldB and the N. crassa ortholog, PLA-7, are required for polarized growth, cell fusion and ascospore development, whereas PldA/PLA-8 are dispensable for these functions. Exogenous addition of PA rescues the cell-fusion phenotype in E. festucae. PldB is also crucial for E. festucae to establish a symbiotic association with L. perenne. This study identifies a new component of the cell-cell communication and cell fusion signaling network that controls hyphal morphogenesis and growth in filamentous fungi.

genetics↗

bioRxiv2018

Analysis of Epichloë festucae small secreted proteins in the interaction with Lolium perenne

Epichloe festucae is an endophyte of the agriculturally important perennial ryegrass. This species systemically colonises the aerial tissues of this host where its growth is tightly regulated thereby maintaining a mutualistic symbiotic interaction. Recent studies have suggested that small secreted proteins, termed effectors, play a vital role in the suppression of host defence responses. To date only a few effectors with important roles in mutualistic interactions have been described. Here we make use of the fully assembled E. festucae genome and EffectorP to generate a suite of 141 effector candidates. These were analysed with respect to their genome location and expression profiles in planta and in several symbiosis-defective mutants. We found an association between effector candidates and a class of transposable elements known as MITEs, but no correlation with other dynamic features of the E. festucae genome, such as transposable element-rich regions. Three effector candidates and a small GPI-anchored protein were chosen for functional analysis based on their high expression in planta compared to in culture and their differential regulation in symbiosis defective E. festucae mutants. All three candidate effector proteins were shown to possess a functional signal peptide and two could be detected in the extracellular medium by western blotting. Localization of the effector candidates in planta suggests that they are not translocated into the plant cell, but rather, are localized in the apoplastic space or are attached to the cell wall. Deletion and overexpression of the effector candidates, as well as the putative GPI-anchored protein, did not affect the plant growth phenotype or restrict growth of E. festucae mutants in planta. These results indicate that these proteins are either not required for the interaction at the observed life stages or that there is redundancy between effectors expressed by E. festucae.

genetics↗