Project description:The production of lignocellulosic-derived biofuels is a highly promising source of alternative energy, but it has been constrained by the lack of a microbial platform capable to efficiently degrade this recalcitrant material and cope with by-products that can be toxic to cells. Species that naturally grow in environments where carbon is mainly available as lignin are promising for finding new ways of removing the lignin that protects cellulose for improved conversion of lignin to fuel precursors.
Project description:Fomitiporia mediterranea (Fmed) is one of the main fungal species found in grapevine wood rot, also called “amadou”, one of the most typical symptoms of grapevine trunk disease Esca. This fungus is functionally classified as a white-rot, able to degrade all wood structure polymers, i.e., hemicelluloses, cellulose, and the most recalcitrant component, lignin. Specific enzymes are secreted by the fungus to degrade those components, namely carbohydrate active enzymes for hemicelluloses and cellulose, which can be highly specific for given polysaccharide, and peroxidases, which enable white-rot to degrade lignin, with specificities relating to lignin composition as well. Furthermore, besides polymers, a highly diverse set of metabolites often associated with antifungal activities is found in wood, this set differing among the various wood species. Wood decayers possess the ability to detoxify these specific extractives and this ability could reflect the adaptation of these fungi to their specific environment. The aim of this study is to better understand the molecular mechanisms used by Fmed to degrade wood structure, and in particular its potential adaptation to grapevine wood. To do so, Fmed was cultivated on sawdust from different origins: grapevine, beech, and spruce. Carbon mineralization rate, mass loss, wood structure polymers contents, targeted metabolites and secreted proteins were measured. We used the well-known white-rot model Trametes versicolor for comparison. Whereas no significant degradation was observed with spruce, a higher mass loss was measured on Fmed grapevine culture compared to beech culture. Moreover, on both substrates, a simultaneous degradation pattern and the degradation of wood extractives were demonstrated, and proteomic analyses identified a relative overproduction of oxidoreductases involved in lignin and extractive degradation on grapevine cultures, and only few differences in carbohydrate active enzymes. These results could explain at least partially the adaptation of Fmed to grapevine wood structural composition compared to other wood species and suggest that other biotic and abiotic factors should be considered to fully understand the potential adaptation of Fmed to its ecological niche.
Project description:Cellulose from plant biomass is the largest renewable energy resource of carbon fixed from the atmosphere, which can be converted into fermentable sugars for production into ethanol. However, the cellulose present as lignocellulosic biomass is embedded in a hemicellulose and lignin matrix from which it needs to be extracted for efficient processing. Here, we show that expression of an Arabidopsis transcription factor SHINE (SHN) in rice, a model for the grasses, causes a 34% increase in cellulose and a 45% reduction in lignin content.
Project description:White rot fungi are able to degrade woody lignin and other persistent organic compounds including artificial chemicals (e.g. chlorinated dioxin) in secondary metabolism. This ability has potential in a wide range of biotechnological applications including remediation of organopollutants and the industrial processing of paper and textiles. Ligninolytic fungi secondarily secrete extracellular oxidative enzymes thought to play an important role in these compounds decay. However, detail of metabolic pathway and initiation signals of the degradation system is unclear. To investigate genes directly and indirectly related to it, we constructed long serial analysis of gene expression (Long SAGE) library from the most studied white rot fungus, Phanerochaete chrysosporium. Keywords: transcriptome profiling To analyze the transcriptome profile during the initiation of manganese peroxidase (MnP) and lignin peroxidase (LiP) production in Phanerochaete chrysosporium, we constructed the day 3 culture (just started the enzyme production) library and the day 2 culture (the activity of enzymes is not detected) library.
Project description:Certain wood decay basidiomycetes, collectively referred to as brown-rot fungi rapidly depolymerize cellulose while leaving behind the bulk of cell wall lignin as a modified residue. The mechanism(s) employed are unclear, but considerable evidence implicates the involvement of diffusible oxidants, particularly hydroxyl radical. Toward a better understanding of this process, we have examined the transcriptome and secretome of Wolfiporia cocos when cultivated on media containing glucose, purified crystalline cellulose, aspen (Populus grandidentata) or lodgepole pine (Pinus contorta) as sole carbon source. Compared to glucose, 39, 331 and 357 genes exhibited 4-fold increases in transcript levels in cellulose, aspen and lodgepole pine, respectively. Mass spectrometry identified peptides corresponding to 64 glycoside hydrolase (GH) proteins and, of these, 17 corresponded to transcripts upregulated on one or both woody substrates. Most of these genes were broadly categorized as hemicellulases or chitinases. Consistent with an important role for ·OH in cellulose depolymerization, high transcript levels and upregulation were observed for genes involved in iron homeostasis, iron reduction and extracellular peroxide generation. These patterns of regulation differ markedly from the closely related brown rot fungus, Postia placenta, and expand the number of enzymes potentially involved in the oxidative depolymerization of cellulose.
Project description:Certain wood decay basidiomycetes, collectively referred to as brown-rot fungi rapidly depolymerize cellulose while leaving behind the bulk of cell wall lignin as a modified residue. The mechanism(s) employed are unclear, but considerable evidence implicates the involvement of diffusible oxidants, particularly hydroxyl radical. Toward a better understanding of this process, we have examined the transcriptome and secretome of Wolfiporia cocos when cultivated on media containing glucose, purified crystalline cellulose, aspen (Populus grandidentata) or lodgepole pine (Pinus contorta) as sole carbon source. Compared to glucose, 39, 331 and 357 genes exhibited 4-fold increases in transcript levels in cellulose, aspen and lodgepole pine, respectively. Mass spectrometry identified peptides corresponding to 64 glycoside hydrolase (GH) proteins and, of these, 17 corresponded to transcripts upregulated on one or both woody substrates. Most of these genes were broadly categorized as hemicellulases or chitinases. Consistent with an important role for ·OH in cellulose depolymerization, high transcript levels and upregulation were observed for genes involved in iron homeostasis, iron reduction and extracellular peroxide generation. These patterns of regulation differ markedly from the closely related brown rot fungus, Postia placenta, and expand the number of enzymes potentially involved in the oxidative depolymerization of cellulose. Medium containing glucose, microcrystalline cellulose, ground aspen or ground lodgepole pine was inoculated with W. cocos. RNA was purified from cultures. Single read 100 bp Illumina runs were performed.
Project description:The Trametes versicolor genome is predicted to encode many enzymes that can effectively degrade lignin, making it a has potentially useful application intool for biopulping and biobleaching. Poplar is an important and widely cultivated species of tree species, which isand extensively applied used in the pulping industry. However, the wood degradation mechanism of T. versicolor from transcriptomic level is not clear. To reveal identify the enzymes that contributeing to lignocellulose degraredauction and its degradation mechanisms, we evaluated transcriptomic how study theof T. versicolor transcriptome was changes during evaluated growthing on the poplar wood relative to growth on glucose medium. 853 genes were differentially expressed;, 360 genes were up-regulated on poplar wood, and 493 genes were down-regulated on poplar wood. Notably, most genes relative involved into lignin degradation were up-regulated, including eight lignin peroxidase (LiP) genes, and two manganese peroxidase (MnP) genes etc. Genes encoding cellulose and hemicelluloses degrading-enzymesation were mostly down-regulated, including six endo-β-1, 4-glucanase genes, three cellobiohydrolase I genes, and one cellobiohydrolase II gene, etc. MeanwhileAdditionally, expression of more significant expansion of P450s in T. versicolor genome, along with differences in carbohydrate- and lignin-degrading enzymes, could bewere correlated withto poplar wood degradation. Our results revealed transcriptomic characterizeation transcriptomic changes related toof lignocellulose degradation. Therefore, our results cwould be benuseful for the development ofefit T. versicolor as a tool to improve the efficiency of lignin degradation, and provide a theoretical foundation for a new paper pulp manufacturing processe 1,T.versicolor groewn on PDA medium. 2, T. versicolor growing on the a glucose carbon medium of glucose. 3, T. versicolor growing on poplar medium
Project description:The fungus Polyporus brumalis is a wood decay fungus previously evidenced as efficient lignin degrader with high potential for plant biomass pre-treatment before conversion into bio-energy. Here we used an RNASeq approach that highlighted the active transcription of an unparalleled number of lignin active peroxidases and H2O2 generating enzymes during growth on wheat straw. These enzymes, together with metabolic processes related to detoxification appear as key determinants of the fungal adaption to lignin degradation.
Project description:The ability to obtain carbon and energy is a major requirement to exist in any environment. For several ascomycete fungi (post-)genomic analyses have shown that species that occupy a large variety of habitats possess a diverse enzymatic machinery, while species with a specific habitat have a more focused enzyme repertoire that is well-adapted to the prevailing substrate. White-rot basidiomycete fungi also live in a specific habitat, as they are found exclusively in wood. In this study we evaluated how well the white-rot fungus Dichomitus squalens has adapted to degrade its natural wood substrate. The transcriptome and exoproteome of D. squalens were analysed after cultivation on two natural substrates, aspen and spruce wood, and two non-woody substrates, wheat bran and cotton seed hulls. D. squalens produced ligninolytic enzymes mainly at the early time point of the wood cultures, indicating the need to degrade lignin to get access to wood polysaccharides. Surprisingly, the response of the fungus to the non-woody polysaccharides was nearly as good match to the substrate composition as observed for the wood polysaccharides. This indicates that D. squalens has preserved its ability to efficiently degrade plant polysaccharides not present in its natural habitat.
Project description:Lignocellulosic biomass is composed of three major biopolymers: cellulose, hemicellulose and lignin. Although lignin has long been considered a waste product in biomass conversion efforts, its utilization has since been identified as critical to the economic viability of second-generation biofuel production. There is thus increasing interest in finding enzymes and enzyme cocktails which can efficiently deconstruct both the cellulose/hemicellulose and lignin components of lignocellulosic biomass. Analytical tools capable of quickly detecting both glycan and lignin deconstruction could are needed to support the development and characterization of efficient enzymes/enzyme cocktails.