Cytoplasmic Lipases—A Novel Class of Fungal Defense Proteins Against Nematodes

Fungi are an attractive food source for predators such as fungivorous nematodes. Several fungal defense proteins and their protective mechanisms against nematodes have been described. Many of these proteins are lectins which are stored in the cytoplasm of the fungal cells and bind to specific glycan epitopes in the digestive tract of the nematode upon ingestion. Here, we studied two novel nematotoxic proteins with lipase domains from the model mushroom Coprinopsis cinerea. These cytoplasmically localized proteins were found to be induced in the vegetative mycelium of C. cinerea upon challenge with fungivorous nematode Aphelenchus avenae. The proteins showed nematotoxicity when heterologously expressed in E. coli and fed to several bacterivorous nematodes. Site-specific mutagenesis of predicted catalytic residues eliminated the in-vitro lipase activity of the proteins and significantly reduced their nematotoxicity, indicating the importance of the lipase activity for the nematotoxicity of these proteins. Our results suggest that cytoplasmic lipases constitute a novel class of fungal defense proteins against predatory nematodes. These findings improve our understanding of fungal defense mechanisms against predators and may find applications in the control of parasitic nematodes in agriculture and medicine.

Integrated Transcriptome Analysis Reveals Plant Hormones Jasmonic Acid and Salicylic Acid Coordinate Growth and Defense Responses upon Fungal Infection in Poplar

Plants have evolved a sophisticated system to respond to various stresses. Fungal attack or infection is one of the most important biotic stresses for most plants. During the defense response to fungal infection, the plant hormones jasmonic acid (JA) and salicylic acid (SA) play critical roles. Here, gene expression data on JA/SA treatments and Melampsora larici-populina (MLP) infection were generated. Integrated transcriptome analyses of these data were performed, and 943 genes in total were identified as common responsive genes (CRG). Gene ontology (GO) term analysis revealed that the genes from CRG are generally involved in the processes of stress responses, metabolism, and growth and development. The further cluster analysis of the CRG identified a set of core genes that are involved in the JA/SA-mediated response to fungal defense with distinct gene expression profiles upon JA/SA treatment, which highlighted the different effects of these two hormones on plant fungal defenses. The modifications of several pathways relative to metabolism, biotic stress, and plant hormone signal pathways suggest the possible roles of JA/SA on the regulation of growth and defense responses. Co-expression modules (CMs) were also constructed using the poplar expression data on JA, SA, M. larici-populina, Septoria musiva, and Marssonina brunnea treatment or infection. A total of 23 CMs were constructed, and different CMs clearly exhibited distinct biological functions, which conformably regulated the concerted processes in response to fungal defense. Furthermore, the GO term analysis of different CMs confirmed the roles of JA and SA in regulating growth and defense responses, and their expression profiles suggested that the growth ability was reduced when poplar deployed defense responses. Several transcription factors (TFs) among the CRG in the co-expression network were proposed as hub genes in regulating these processes. According to this study, our data finely uncovered the possible roles of JA/SA in regulating the balance between growth and defense responses by integrating multiple hormone signaling pathways. We were also able to provide more knowledge on how the plant hormones JA/SA are involved in the regulation of the balance between growth and plant defense.

Toxicity of Potential Fungal Defense Proteins towards the Fungivorous NematodesAphelenchus avenaeandBursaphelenchus okinawaensis

ABSTRACTResistance of fungi to predation is thought to be mediated by toxic metabolites and proteins. Many of these fungal defense effectors are highly abundant in the fruiting body and not produced in the vegetative mycelium. The defense function of fruiting body-specific proteins, however, including cytoplasmically localized lectins and antinutritional proteins such as biotin-binding proteins, is mainly based on toxicity assays using bacteria as a heterologous expression system, with bacterivorous/omnivorous model organisms as predators. Here, we present an ecologically more relevant experimental setup to assess the toxicity of potential fungal defense proteins towards the fungivorous, stylet-feeding nematodesAphelenchus avenaeandBursaphelenchus okinawaensis. As a heterologous expression host, we exploited the filamentous fungusAshbya gossypii. Using this new system, we assessed the toxicity of six previously characterized, cytoplasmically localized, potential defense proteins from fruiting bodies of different fungal phyla against the two fungivorous nematodes. We found that all of the tested proteins were toxic against both nematodes, albeit to various degrees. The toxicity of these proteins against both fungivorous and bacterivorous nematodes suggests that their targets have been conserved between the different feeding groups of nematodes and that bacterivorous nematodes are valid model organisms to assess the nematotoxicity of potential fungal defense proteins.IMPORTANCEOur results support the hypothesis that cytoplasmic proteins abundant in fungal fruiting bodies are involved in fungal resistance against predation. The toxicity of these proteins toward stylet-feeding nematodes, which are also capable of feeding on plants, and the abundance of these proteins in edible mushrooms, may open possible avenues for biological crop protection against parasitic nematodes, e.g., by expression of these proteins in crops.