RNA-sequencing reveals the complexities of the transcriptional response to lignocellulosic biofuel substrates in Aspergillus niger.

Steven T Pullan,Matthew Kokolski,Gregory A Tucker,David B Archer,Almar Neiteler,Stéphane Delmas,Roger Ibbett,Jolanda M Van Munster,Paul Daly,Raymond Wilson,Martin J Blythe,Sanyasi Gaddipati

doi:10.1186/s40694-014-0003-x

Abstract

BackgroundSaprobic 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.ResultsIn 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.ConclusionsBy 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.Electronic supplementary materialThe online version of this article (doi:10.1186/s40694-014-0003-x) contains supplementary material, which is available to authorized users.

Highlights

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
To re-introduce a simple carbon source in the presence of lignocellulose, glucose was added to the willow cultures after 24 h and incubated for a further 5 h
The inducing substrate used for the production of enzymes has marked effects upon the enzyme cocktail achieved highlighting the limitations of the use of simple inducing substrates for the generation of complex enzyme mixtures

Summary

Introduction

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. The enzyme cocktails produced mainly by saprobic fungi are usually deficient in activities required for complete saccharification of a particular substrate. One cause of this may be the industrial use of simple, cost effective inducers, such as lactose or cellulose which fail to induce the full range of hydrolytic and accessory activities encoded within an organism’s genome. The pairing of inducer and lignocellulosic substrate is a key consideration in biofuel production

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Results

Conclusion