Project description:Filamentous fungi are widely used in the production of biomass degrading enzymes, e.g. cellulases and pectinases. In order to study the secretome of biomass degrading fungi, proteomics studies were carried out on the extracellular proteins of fungal strains.
Project description:The rapid transport of ribosomal proteins (RPs) into the nucleus and their efficient assembly into rRNA are prerequisites for ribosome biogenesis. Proteins that act as dedicated chaperones for RPs to maintain their stability and facilitate their assembly have not been identified in filamentous fungi. PlCYP5 is a nuclear cyclophilin in the nematophagous fungus Purpureocillium lilacinum, and up-regulated expression in response to abiotic stress and nematode egg-parasitism. Here, we found that PlCYP5 interacted with the unassembled small ribosomal subunit protein, PlRPS15, of the uS19 family. PlRPS15 contained a eukaryote-specific N-terminal extension that mediated the interaction. The phenotypes of the PlCYP5 loss-of-function mutant were similar to those of the PlRPS15 knockdown mutant (e.g., growth and ribosome biogenesis defects). PlCYP5 maintained the solubility of PlRPS15 independent of its catalytic peptide-prolyl isomerase function and supported the integration of PlRPS15 into pre-ribosomes. PlCYP5 homologs in Arabidopsis thaliana, Homo sapiens, Schizosaccharomyces pombe, Sclerotinia sclerotiorum, Botytis cinerea, and Metarhizium anisopliae were identified. Notably, the interaction of their homologs corresponding to the PlCYP5-PlRPS15 pattern existed in three filamentous fungi, while lacked in other species. In summary, our data disclosed a special RP dedicated chaperone system in filamentous fungi, in which cyclophilin was enlisted to perform the chaperone funtion.
Project description:The efficiency of microorganisms to degrade lignified plants is of great importance in Earth’s carbon cycle, but also in industrial biorefinery processes, such as for biofuel production. Here, we present a large-scale proteomics approach to investigate and compare the enzymatic response of five filamentous fungi when grown on five very different substrates: bagasse, birch, spruce, cellulose and glucose. The five fungi included the ascomycetes Aspergillus terreus, Hypocrea jecorina, Myceliophthora thermophila, Neurospora crassa and the white-rot basidiomycete Phanerochaete chrysosporium, all expressing a diverse repertoire of enzymes. Many studies have previously been performed with these fungi under various growth conditions, but this study is unique as it presents comparable quantitative protein abundance values across five filamentous fungi and five diverse substrates that allows for direct comparison of fungal response to the different substrates; this approach gives indications to substrate specificity of individual carbohydrate-active enzymes (CAZymes) and identifies several novel co-expressed non-CAZymes. Specifically, we present a quantitative comparison of 34 lytic polysaccharide monooxidenases (LPMOs), which are crucial enzymes in biomass deconstruction.
Project description:We describe a simple, plate-based method to analyze the secretomes of microorganisms growing on insoluble substrates that allows harsh sample preparation methods promoting desorption, and subsequent identification, of substrate-bound proteins, while minimizing contamination with non-secreted proteins from leaking or lysed cells. The method described here enables rapid large-scale comparative studies of the secretomes of filamentous fungi and other microorganisms growing on a variety of solid substrates, which eventually may have implications for our understanding of enzymatic lignocellulose degradation.
Project description:A variety of small RNAs, including the Dicer-dependent miRNAs and the Dicer-independent Piwi-interacting RNAs, associate with Argonaute family proteins to regulate gene expression in diverse cellular processes. These two species of small RNA have not been found in fungi. Here, by analyzing small RNA associated with the Neurospora Argonaute protein QDE-2, we show that diverse pathways generate miRNA-like small RNAs (milRNAs) and Dicer-independent small interfering RNAs (disiRNAs) in this filamentous fungus. Surprisingly, milRNAs are produced by at least four different mechanisms that use a distinct combination of factors, including Dicers, QDE-2, the exonuclease QIP and an RNAse III domain-containing protein MRPL3. In contrast, disiRNAs originate from loci producing overlapping sense and antisense transcripts, and do not require the known RNAi components for their production. Taken together, these results uncover several pathways for small RNA production in filamentous fungi, shedding light on the diversity and evolutionary origins of eukaryotic small RNAs.