Abstract
BackgroundThe thermotolerant yeast Kluyveromyces marxianus is a potential candidate for high-temperature fermentation. When K. marxianus was used for high-temperature ethanol fermentation, a fermentation arrest was observed during the late fermentation stage and the stress responses have been investigated based on the integration of RNA-Seq and metabolite data. In order to bring new insights into the cellular responses of K. marxianus after the fermentation arrest during high-temperature ethanol fermentation, quantitative proteomic profiling and integrated analysis with transcriptomic data were performed in this study.ResultsSamples collected at 14, 16, 18, 20 and 22 h during high-temperature fermentation were subjected to isobaric tags for relative and absolute quantitation (iTRAQ)-based proteomic profiling and integrated analysis with transcriptomic data. The correlations between transcripts and proteins for the comparative group 16 h vs 14 h accounted for only 4.20% quantified proteins and 3.23% differentially expressed proteins (DEPs), respectively, much higher percentages of correlations (30.56%–59.11%) were found for other comparative groups (i.e., 18 h vs 14 h, 20 h vs 14 h, and 22 h vs 14 h). According to Spearman correlation tests between transcriptome and proteome (the absolute value of a correlation coefficient between 0.5 and 1 indicates a strong correlation), poor correlations were found for all quantified proteins (R = − 0.0355 to 0.0138), DEPs (R = − 0.0079 to 0.0233) and the DEPs with opposite expression trends to corresponding differentially expressed genes (DEGs) (R = − 0.0478 to 0.0636), whereas stronger correlations were observed in terms of the DEPs with the same expression trends as the correlated DEGs (R = 0.5593 to 0.7080). The results of multiple reaction monitoring (MRM) verification indicate that the iTRAQ results were reliable. After the fermentation arrest, a number of proteins involved in transcription, translation, oxidative phosphorylation and fatty acid metabolism were down-regulated, some molecular chaperones and proteasome proteins were up-regulated, the ATPase activity significantly decreased, and the total fatty acids gradually accumulated. In addition, the contents of palmitic acid, oleic acid, C16, C18, C22 and C24 fatty acids increased by 16.77%, 28.49%, 14.14%, 26.88%, 628.57% and 125.29%, respectively.ConclusionsThis study confirmed some biochemical and enzymatic alterations provoked by the stress conditions in the specific case of K. marxianus: such as decreases in transcription, translation and oxidative phosphorylation, alterations in cellular fatty acid composition, and increases in the abundance of molecular chaperones and proteasome proteins. These findings provide potential targets for further metabolic engineering towards improvement of the stress tolerance in K. marxianus.
Highlights
The thermotolerant yeast Kluyveromyces marxianus is a potential candidate for high-temperature fermentation
We found that when K. marxianus was used for high-temperature ethanol fermentation, there was nearly no difference in glucose consumption and ethanol production between 30 and 45 °C during the early and middle stages of fermentation, but a fermentation arrest was observed during the late fermentation stage at 45 °C [20]
Functional annotation of all the identified proteins was conducted based on the annotation information from the Gene Ontology (GO) database [39, 40] and the Kyoto Encyclopedia of Genes and Genomes (KEGG) database [41, 42]
Summary
The thermotolerant yeast Kluyveromyces marxianus is a potential candidate for high-temperature fermentation. When K. marxianus was used for high-temperature ethanol fermentation, a fermentation arrest was observed during the late fermentation stage and the stress responses have been investigated based on the integration of RNA-Seq and metabolite data. In order to bring new insights into the cellular responses of K. marxianus after the fermentation arrest during high-temperature ethanol fermentation, quantitative proteomic profiling and integrated analysis with transcriptomic data were performed in this study. Large-scale bioethanol production benefits greatly from high-temperature fermentation (≥ 40 °C), which can significantly reduce cooling costs and help prevent contamination [4, 5]. The thermotolerant yeast Kluyveromyces marxianus, which can grow at elevated temperatures (> 40 °C) [12,13,14], becomes a potential candidate for high-temperature ethanol fermentation. The cellular responses of K. marxianus to stress conditions have been investigated based on RNA sequencing (RNA-Seq)-based transcriptomic analysis [16, 19, 20]
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