Abstract
Background: The thermophilic, anaerobic bacterium, Clostridium thermocellum is a model organism for consolidated processing due to its efficient fermentation of cellulose. Constituents of dilute acid pretreatment hydrolysate are known to inhibit C. thermocellum and other microorganisms. To evaluate the biological impact of this type of hydrolysate, a transcriptomic analysis of growth in hydrolysate-containing medium was conducted on 17.5% v/v Populus hydrolysate-tolerant mutant (PM) and wild type (WT) strains of C. thermocellum.
Results: In two levels of Populus hydrolysate medium (0% and 10% v/v), the PM showed both gene specific increases and decreases of gene expression compared to the wild-type strain. The PM had increased expression of genes in energy production and conversion, and amino acid transport and metabolism in both standard and 10% v/v Populus hydrolysate media. In particular, expression of the histidine metabolism increased up to 100 fold. In contrast, the PM decreased gene expression in cell division and sporulation (standard medium only), cell defense mechanisms, cell envelope, cell motility, and cellulosome in both media. The PM downregulated inorganic ion transport and metabolism in standard medium but upregulated it in the hydrolysate media when compared to the WT. The WT differentially expressed 1072 genes in response to the hydrolysate medium which included increased transcription of cell defense mechanisms, cell motility, and cellulosome, and decreased expression in cell envelope, amino acid transport and metabolism, inorganic ion transport and metabolism, and lipid metabolism, while the PM only differentially expressed 92 genes. The PM tolerates up to 17.5% v/v Populus hydrolysate and growth in it elicited 489 genes with differential expression, which included increased expression in energy production and conversion, cellulosome production, and inorganic ion transport and metabolism and decreased expression in transcription and cell defense mechanisms.
Conclusion: These results suggest the mechanisms of tolerance for the Populus hydrolysate-tolerant mutant strain of C. thermocellum are based on increased cellular efficiency caused apparently by downregulation of non-critical genes and increasing the expression of genes in energy production and conversion rather than tolerance to specific hydrolysate components. The wild type, conversely, responds to hydrolysate media by down-regulating growth genes and up-regulating stress response genes.
| Original language | English |
|---|---|
| Article number | 215 |
| Journal | BMC Microbiology |
| Volume | 14 |
| Issue number | 1 |
| DOIs | |
| State | Published - Aug 16 2014 |
Funding
The authors thank Dawn M. Klingeman and Courtney M. Johnson for assistance with RNA purification; Dawn M. Klingeman and Charlotte M. Wilson for qPCR and PCR preparation and analysis and Qiang He and Chris Hemme for assistance with transcriptome analysis. RNA-Seq data was generated by the U.S. Department of Energy (DOE) Joint Genome Institute, which is supported by the Office of Science of the under contract no. DE-AC02-05CH11231. This research was supported by the BioEnergy Science Center, a Department of Energy Bioenergy Research Center supported by the Office of Biological and Environmental Research in the Department of Energy Office of Science. Additional support was provided by the Institute for a Secure and Sustainable Environment at the University of Tennessee. Oak Ridge National Laboratory is managed by UT-Battelle, LLC, for the DOE under Contract DE-AC05-00OR22725. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
Keywords
- Clostridium thermocellum
- Consolidated bioprocessing
- Gene expression
- Inhibitor tolerance
- Populus hydrolysate
- RNA-seq
- Transcriptomic