Proteomic profiling and integrated analysis with transcriptomic data bring new insights in the stress responses of Kluyveromyces marxianus after an arrest during high-temperature ethanol fermentation

Abstract

Abstract Background 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. Results Samples 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. Conclusions This 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.

Keywords

• Kluyveromyces marxianus
• Proteome
• Correlation analysis
• High-temperature fermentation
• Stress response