Secretome analysis of biomass degrading filamentous fungi
Ontology highlight
ABSTRACT:
ORGANISM(S): Aspergillus Brunneoviolaceus
SUBMITTER: 
Liwei Gao
PROVIDER: PXD048232 | iProX | Wed Jan 03 00:00:00 GMT 2024
REPOSITORIES: iProX
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2017-02-27 | PXD005532 | Pride
Project description:Wood-degrading fungi play a critical role in global carbon cycling, and their varied mechanisms for deconstruction offer pathways for industrial bioconversion. In this study, we used comparative genomics to isolate upregulation patterns among fungi with brown rot (carbohydrate-selective) or white rot (lignin-degrading) nutritional modes. Specifically, we used whole-transcriptome profiling to compare early, middle, and late decay stages on wood wafers, matching differentially-expressed gene (DEG) patterns with fungal growth and enzyme activities. This approach highlighted 34 genes uniquely upregulated in early brown rot stages, with notable candidates involved in generating reactive oxygen species (ROS) as a pretreatment mechanism during brown rot. This approach further isolated 18 genes in late brown rot stages that may be adapted to handle oxidatively-reacted lignocellulose components. By summing gene expression levels in functional classes, we also identified a broad and reliable distinction in glycoside hydrolase (GH) versus lignocellulose oxidative (LOX) transcript counts that may reflect the energy investment burden of lignin-degrading machinery among white rot fungi.
2018-01-05 | GSE108189 | GEO
Project description:Fermentation of sugars derived from plant biomass feedstock is crucial in achieving sustainability. Hence, utilizing customized enzymatic cocktails to obtain oligosaccharides instead of monomers is an alternative fermentation strategy to produce prebiotics, cosmetics, and biofuels. This study developed an engineered strain of Aspergillus niger producing a tailored cellulolytic cocktail capable of partially degrading sugarcane straw to yield cellooligosaccharides. The A. niger prtT strain created resulted in a reduced extracellular protease production. The prtT background was then used to create strains by deleting exoenzymes encoding genes involved in mono- or disaccharide formation. Consequently, we successfully generated a tailored prtTbglA strain by eliminating a beta-glucosidase (bglA) gene and subsequently deleted two cellobiohydrolases and one beta-xylosidase encoding genes using a multiplex strategy, resulting in the Quintuple strain (prtT; bglA; cbhA; cbhB; xlnD). When applied for sugarcane biomass degradation, the tailored secretomes produced by A. niger resulted in a higher ratio of cellobiose and cellotriose compared with glucose relative to the reference strain. Mass spectrometry revealed that the Quintuple strain secreted alternative cellobiohydrolases and beta-glucosidases to compensate for the absence of major cellulases. Enzymes targeting minor polysaccharides in plant biomass were also upregulated in this tailored strain. Tailored secretome use increased COS/glucose ratio during sugarcane biomass degradation showing that deleting some enzymatic components is an effective approach for producing customized enzymatic cocktails. Our findings highlight the plasticity of fungal genomes as enzymes that target minor components of plant cell walls, and alternative cellulases were produced by the mutant strain. Despite deletion of important secretome components, fungal growth was maintained in plant biomass.
2024-07-12 | MSV000095310 | MassIVE
Project description:Microarrays have become established tools for describing microbial systems, however the assessment of expression profiles for environmental microbial communities still presents unique challenges. Notably, the concentration of particular transcripts are likely very dilute relative to the pool of total RNA, and PCR-based amplification strategies are vulnerable to amplification biases and the appropriate primer selection. Thus, we apply a signal amplification approach, rather than template amplification, to analyze the expression of selected lignin-degrading enzymes in soil. Controls in the form of known amplicons and cDNA from Phanerochaete chrysosporium were included and mixed with the soil cDNA both before and after the signal amplification in order to assess the dynamic range of the microarray. We demonstrate that restored prairie soil expresses a diverse range of lignin-degrading enzymes following incubation with lignin substrate, while farmed agricultural soil does not. The mixed additions of control cDNA with soil cDNA indicate that the mixed biomass in the soil does interfere with low abundance transcript changes, nevertheless our microarray approach consistently reports the most robust signals. Keywords: comparative analysis, microbial ecology, soil microbial communities We used lignin degradation as a model process to demonstrate the use of an oligonucleotide microarray for directly detecting gene expression in soil communities using signal amplification instead of template amplification to avoid the introduction of PCR bias. In the current study, we analyzed mRNA isolated from two distinct soil microbial communities and demonstrate our ability to detect the expression of a small subset of lignin degrading genes following exposure to a lignitic substrate. We also included purified control amplicons and mixed target experiments with pure P. chrysosporium genomic cDNA to determine the level of interference from soil biomass on target hybridization.
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Project description:The biodegradation of lignite (brown coal) by microorganisms has the potential for bioremediation of contaminated mining sites and to generate alternative ways to valorize lignite, such as by producing humic acids or building block chemicals. Previously, a lignite-degrading strain of Trichoderma was isolated, but the genomic and transcriptomic basis of its lignite-degrading ability remained unknown. Here we report that the sequenced genome of the T. cf. simile WF8 strain encoded for enzymes with roles in the degradation of lignite, and potentially tolerance to lignite-breakdown products. There was only a small number of annotated unique genes in the T. cf. simile WF8 genome compared to other fungi, and likely the expression of gene families shared with other fungi is a key factor in lignite biosolubilization by T. cf. simile. The transcriptomes were analyzed of T. cf. simile cultured at two time-points with the lignite-breakdown model compounds 4-phenoxybenzoic acid (which was growth inhibitory), and phenetole and 9-10-dibutoxyanthracene (neither of which inhibited growth), and showed ~20% of genes up-regulated by one or more of these compounds. The analysis highlights candidates for characterization and engineering enzyme over-expressing T. cf. simile strains with potentially improved degradation capacity, e.g., laccases and peroxidases, or tolerance and catabolism of breakdown products, e.g., cytochrome P450s, and ring cleavage dioxygenases. Published in International Biodeterioration & Biodegradation (https://doi.org/10.1016/j.ibiod.2025.105997)
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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.
2020-11-25 | PXD011166 | Pride
Project description:The regulation of plant biomass degradation by fungi is critical to the carbon cycle, and applications in bioproducts and biocontrol. Trichoderma harzianum is an important plant biomass degrader, enzyme producer, and biocontrol agent, but few putative major transcriptional regulators have been deleted in this species. The T. harzianum ortholog of the transcriptional activator XYR1/XlnR/XLR-1 was deleted, and the mutant strains were analyzed by growth profiling, enzymatic activities, and transcriptomics on cellulose. From plate cultures, the Δxyr1 mutant had reduced growth on D-xylose, xylan, and cellulose, and from shake-flask cultures with cellulose, the Δxyr1 mutant had ~ 90% lower β-glucosidase activity, and no detectable β-xylosidase or cellulase activity. The comparison of the transcriptomes from 18 h shake-flask cultures on D-fructose, without a carbon source, and cellulose, showed major effects of XYR1 deletion whereby the Δxyr1 mutant on cellulose was transcriptionally most similar to the cultures without a carbon source. The cellulose induced 43 plant biomass-degrading CAZymes including xylanases as well as cellulases, and most of these had massively lower expression in the Δxyr1 mutant. Expression of a sub-set of carbon catabolic enzymes, other transcription factors, and sugar transporters was also lower in the Δxyr1 mutant on cellulose. In summary, T. harzianum XYR1 is the master regulator of cellulases and xylanases, as well as regulating carbon catabolic enzymes.
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Project description:Wood-degrading fungi vary in their strategies for deconstructing wood, and their competitive successes shape the rate and fate of carbon released from wood, Earth’s largest pool of aboveground terrestrial carbon. In this study, one-on-one interspecific interactions between two model brown rot (carbohydrate-selective) fungi, Gloeophyllum trabeum and Rhodonia (Postia) placenta, were studied on wood wafers where a clearly resolved interaction zone (IZ) could be generated, reproducibly. Comparative RNAseq and proteomics between the IZ and non-interacting hyphae of each species identified combative strategies for each fungus. Glycoside hydrolases were a relatively smaller portion of the interaction secretome compared to non-interacting hyphae. The interaction zone showed higher pectinase specific activity than all other sampling locations, and higher laminarinase specific activity (branched β‐glucan proxy) was seen in the IZ secretome relative to equivalent hyphae in single‐species cultures. Our efforts also identified two distinct competitive strategies in these two fungi with a shared nutritional mode (brown rot) but polyphyletic ancestral lineages. Gloeophyllum trabeum (Gloeophyllum clade) employed secondary metabolite (SM) synthesis in response to a competitor, as shown by the upregulation of several SM-synthesizing genes in the interaction. R. placenta (Antrodia clade) instead upregulated a larger variety of uncharacterized oxidoreductases in interacting hyphae, suggesting that an oxidative burst may be a response to competitors in this fungus. Both species produced several hypothetical proteins exclusively in the interaction zone, leaving abundant unknowns on the battlefield. This work supports the existence of multiple interaction strategies among brown rot fungi and highlights the functional diversity among wood decay fungi.
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2010-11-29 | E-GEOD-23606 | biostudies-arrayexpress
Project description:Microarrays have become established tools for describing microbial systems, however the assessment of expression profiles for environmental microbial communities still presents unique challenges. Notably, the concentration of particular transcripts are likely very dilute relative to the pool of total RNA, and PCR-based amplification strategies are vulnerable to amplification biases and the appropriate primer selection. Thus, we apply a signal amplification approach, rather than template amplification, to analyze the expression of selected lignin-degrading enzymes in soil. Controls in the form of known amplicons and cDNA from Phanerochaete chrysosporium were included and mixed with the soil cDNA both before and after the signal amplification in order to assess the dynamic range of the microarray. We demonstrate that restored prairie soil expresses a diverse range of lignin-degrading enzymes following incubation with lignin substrate, while farmed agricultural soil does not. The mixed additions of control cDNA with soil cDNA indicate that the mixed biomass in the soil does interfere with low abundance transcript changes, nevertheless our microarray approach consistently reports the most robust signals. Keywords: comparative analysis, microbial ecology, soil microbial communities
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