Genome-scale analyses of butanol tolerance in Saccharomyces cerevisiae reveal an essential role of protein degradation

Biotechnology for Biofuels

Daniel González-Ramos¹  Marcel van den Broek¹  Antonius JA van Maris¹  Jack T Pronk²  Jean-Marc G Daran² 

[1] Kluyver Centre for Genomics of Industrial Fermentation, P.O. Box 5057, Delft 2600 GA, The Netherlands

[2] Platform for Green Synthetic Biology, P.O. Box 5057, Delft 2600 GA, The Netherlands

关键词: Multivesicular bodies;    Proteasome;    Whole genome sequencing;    Deletion collection screening;    Evolutionary engineering;    Butanol tolerance;    Saccharomyces cerevisiae;   

Others  :  798108
DOI  :  10.1186/1754-6834-6-48

 received in 2013-01-18, accepted in 2013-03-22,  发布年份 2013

【 摘 要 】

Background

n-Butanol and isobutanol produced from biomass-derived sugars are promising renewable transport fuels and solvents. Saccharomyces cerevisiae has been engineered for butanol production, but its high butanol sensitivity poses an upper limit to product titers that can be reached by further pathway engineering. A better understanding of the molecular basis of butanol stress and tolerance of S. cerevisiae is important for achieving improved tolerance.

Results

By combining a screening of the haploid S. cerevisiae knock-out library, gene overexpression, and genome analysis of evolutionary engineered n-butanol-tolerant strains, we established that protein degradation plays an essential role in tolerance. Strains deleted in genes involved in the ubiquitin-proteasome system and in vacuolar degradation of damaged proteins showed hypersensitivity to n-butanol. Overexpression of YLR224W, encoding the subunit responsible for the recognition of damaged proteins of an ubiquitin ligase complex, resulted in a strain with a higher n-butanol tolerance. Two independently evolved n-butanol-tolerant strains carried different mutations in both RPN4 and RTG1, which encode transcription factors involved in the expression of proteasome and peroxisomal genes, respectively. Introduction of these mutated alleles in the reference strain increased butanol tolerance, confirming their relevance in the higher tolerance phenotype. The evolved strains, in addition to n-butanol, were also more tolerant to 2-butanol, isobutanol and 1-propanol, indicating a common molecular basis for sensitivity and tolerance to C3 and C4 alcohols.

Conclusions

This study shows that maintenance of protein integrity plays an essential role in butanol tolerance and demonstrates new promising targets to engineer S. cerevisiae for improved tolerance.

【 授权许可】

   
2013 González-Ramos et al.; licensee BioMed Central Ltd.

【 图 表 】

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