Project description:Background: Saprobic fungi are the predominant industrial sources of Carbohydrate Active enZymes (CAZymes) used for the saccharification of lignocellulose during the production of second generation biofuels. The production of more effective enzyme cocktails is a key objective for efficient biofuel production. To achieve this objective, it is crucial to understand the response of fungi to lignocellulose substrates. Our previous study used RNA-seq to identify the genes induced in Aspergillus niger in response to wheat straw, a biofuel feedstock, and showed that the range of genes induced was greater than previously seen with simple inducers [GSE33852]. Results: In this work we used RNA-seq to identify the genes induced in A. niger in response to short rotation coppice willow and compared this with the response to wheat straw from our previous study, at the same time-point. The response to willow showed a large increase in expression of genes encoding CAZymes. Genes encoding the major activities required to saccharify lignocellulose were induced on willow such as endoglucanases, cellobiohydrolases and xylanases. The transcriptome response to willow had many similarities with the response to straw with some significant differences in the expression levels of individual genes which are discussed in relation to differences in substrate composition or other factors. Differences in transcript levels include higher levels on wheat straw from genes encoding enzymes classified as members of GH62 (an arabinofuranosidase) and CE1 (a feruloyl esterase) CAZy families whereas two genes encoding endoglucanases classified as members of the GH5 family had higher transcript levels when exposed to willow. There were changes in the cocktail of enzymes secreted by A. niger when cultured with willow or straw. Assays for particular enzymes as well as saccharification assays were used to compare the enzyme activities of the cocktails. Wheat straw induced an enzyme cocktail that saccharified wheat straw to a greater extent than willow. Genes not encoding CAZymes were also induced on willow such as hydrophobins as well as genes of unknown function. Several genes were identified as promising targets for future study. Conclusions: By comparing this first study of the global transcriptional response of a fungus to willow with the response to straw, we have shown that the inducing lignocellulosic substrate has a marked effect upon the range of transcripts and enzymes expressed by A. niger. The use by industry of complex substrates such as wheat straw or willow could benefit efficient biofuel production. Six samples in total consisting of duplicate shake flask Aspergillus niger cultures from three conditions: glucose 48 h, willow 24 h, willow 24 h + glucose 5 h

Project description:Brazil is the world’s second largest producer of ethanol. Increasing demand for this biofuel has called for investment in new technologies. One example of such technologies is the second-generation (2G) ethanol production. Lignocellulose comprises mainly cellulose and hemicellulose, which consist of high-energy sugars that can be converted into ethanol. Aspergillus sp play an important role in the recycling of lignocellulosic biomass. These species have been investigated as a cell factory to produce industrial enzymes. To improve our understanding of the mechanisms used by Aspergillus sp during degradation of plant biomass and to identify the hydrolytic enzymes secreted by these fungi, we have analyzed the transcriptome and secretome of an Aspergillus species grown on sugarcane bagasse (SEB). A. fumigatus was cultivated in the presence of fructose or SEB. Its cellulolytic and xylanolytic activities depended on time. The maximum activities of endoglucanase and xylanase from the fungus grown on SEB were 0.0029 and 10.82 U mL-1, respectively. Characterization of the transcriptome by RNAseq technology helped to identify genes that participated in the degradation of the biomass demonstrating potential application in the process of enzymatic hydrolysis. The RNAseq data revealed that 2287 genes were differentially expressed in the fungus grown on SEB; 1181 and 1046 of these genes were up- and down-regulated, respectively. There were several CAZymes among the up-regulated genes. Most of these enzymes belonged to the GH family, which includes important hydrolytic and accessory enzymes involved in the degradation of lignocellulose. Similarly, proteomic studies showed that a total of 130 proteins existed in the fungus grown on SEB. These proteins were classified into several groups of secreted extracellular enzymes, and their functional classification demonstrated that 59% of the proteins were CAZymes such as GH45-endoglucanases, GH6 and GH7-cellobiohydrolases, GH3-β-glucosidases, GH10-xylanases, and AA9-LPMO. Data obtained by transcriptome and secretome analyses indicated that A. fumigatus produces a considerable number of CAZymes that participate in the hydrolysis of biomass. These CAZymes are valuable for the lignocellulosic bioenergy industry, provide information for future studies on the degradation of biomass, and improve understanding of the role genes and enzymes play in the degradation process.

Project description:Background: Saprobic fungi are the predominant industrial sources of Carbohydrate Active enZymes (CAZymes) used for the saccharification of lignocellulose during the production of second generation biofuels. The production of more effective enzyme cocktails is a key objective for efficient biofuel production. To achieve this objective, it is crucial to understand the response of fungi to lignocellulose substrates. Our previous study used RNA-seq to identify the genes induced in Aspergillus niger in response to wheat straw, a biofuel feedstock, and showed that the range of genes induced was greater than previously seen with simple inducers [GSE33852]. Results: In this work we used RNA-seq to identify the genes induced in A. niger in response to short rotation coppice willow and compared this with the response to wheat straw from our previous study, at the same time-point. The response to willow showed a large increase in expression of genes encoding CAZymes. Genes encoding the major activities required to saccharify lignocellulose were induced on willow such as endoglucanases, cellobiohydrolases and xylanases. The transcriptome response to willow had many similarities with the response to straw with some significant differences in the expression levels of individual genes which are discussed in relation to differences in substrate composition or other factors. Differences in transcript levels include higher levels on wheat straw from genes encoding enzymes classified as members of GH62 (an arabinofuranosidase) and CE1 (a feruloyl esterase) CAZy families whereas two genes encoding endoglucanases classified as members of the GH5 family had higher transcript levels when exposed to willow. There were changes in the cocktail of enzymes secreted by A. niger when cultured with willow or straw. Assays for particular enzymes as well as saccharification assays were used to compare the enzyme activities of the cocktails. Wheat straw induced an enzyme cocktail that saccharified wheat straw to a greater extent than willow. Genes not encoding CAZymes were also induced on willow such as hydrophobins as well as genes of unknown function. Several genes were identified as promising targets for future study. Conclusions: By comparing this first study of the global transcriptional response of a fungus to willow with the response to straw, we have shown that the inducing lignocellulosic substrate has a marked effect upon the range of transcripts and enzymes expressed by A. niger. The use by industry of complex substrates such as wheat straw or willow could benefit efficient biofuel production.

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	20130922_Pchrys01.raw	Raw
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	20130922_Pchrys04.raw	Raw
	20130922_Pchrys05.raw	Raw

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Interspecies and intersubstrate comparison of the biomass-degrading capacity of five filamentous fungi

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Quantitative comparison of the biomass-degrading enzyme repertoires of five filamentous fungi.

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