Transcriptome changes in adaptive evolution of xylose-fermenting industrial Saccharomyces cerevisiae strains with δ-integration of different xylA genes

Abstract

It is of utmost importance to construct industrial xylose-fermenting Saccharomyces cerevisiae strains for lignocellulosic bioethanol production. In this study, two xylose isomerase-based industrial S. cerevisiae strains, O7 and P5, were constructed by δ-integration of the xylose isomerase (XI) gene xylA from the fungus Orpinomyces sp. and from the bacterium Prevotella ruminicola, respectively. The xylose consumption of the strains O7 and P5 at 48-h fermentation was 17.71 and 26.10 g/L, respectively, in synthetic medium with xylose as the sole sugar source. Adaptive evolution further improved the xylose fermentation capacity of the two strains to 51.0 and 28.9% in average, respectively. The transcriptomes of these two strains before and after evolution were analyzed using RNA-Seq. The expression levels of the genes involved in cell integrity, non-optimal sugar utilization, and stress response to environment were significantly up-regulated after evolution and did not depend on the origin of xylA; the expression levels of the genes involved in transmembrane transport, rRNA processing, cytoplasmic translation, and other processes were down-regulated. The expression of genes involved in central carbon metabolism was fine-tuned after the evolution. The analysis of transcription factors (TFs) indicated that most of the genes with significant differential expression were regulated by the TFs related to cell division, DNA damage response, or non-optimal carbon source utilization. The results of this study could provide valuable references for the construction of efficient xylose-fermenting XI strains.

Metabolic engineering considerations for the heterologous expression of xylose-catabolic pathways in Saccharomyces cerevisiae

“…First, the deletion of GRE3 encoding aldose reductase and/or the deletion of SOR1 encoding sorbitol (xylitol) dehydrogenase were proposed to reduce xylitol accumulation [ 45 , 46 ]. Also, extra copies of xylA and/or XYL3 [ 22 , 39 , 42 ] were often accompanied with the overexpression of the PP pathway genes such as TAL1 to improve xylose consumption rates. In Fig 5B , the necessity and contribution of each factors above were evaluated.…”

“…reduce xylitol accumulation [45,46]. Also, extra copies of xylA and/or XYL3 [22,39,42] were often accompanied with the overexpression of the PP pathway genes such as TAL1 to improve xylose consumption rates. In Fig 5B, the necessity and contribution of each factors above were evaluated.…”

Genomic and phenotypic characterization of a refactored xylose-utilizing Saccharomyces cerevisiae strain for lignocellulosic biofuel production

“…The most pronounced difference between the XI- and XR/XDH-based strains was the transcription levels of the genes involved in the nonoxidative PP pathway. As shown in Additional file 4 : Figure S1, nonoxidative PP pathway enzymes were downregulated (or rearranged) to reduce their transcriptional burdens in XI-based strains [ 24 , 41 ]. Since the limited metabolic flux through the PP pathway is a known bottleneck for efficient xylose fermentation, strain engineering for enhanced xylose metabolism often involves overexpression of the genes in the nonoxidative PP pathway [ 15 , 24 ].…”

Combination of mutagenesis and adaptive evolution to engineer salt‐tolerant and aroma‐producing yeast for soy sauce fermentation

“…To evaluate the accumulation of metabolically compatible solutes, yeast strains were grown to the midlog phase (OD 660 of 2) in YEPD medium, and then the cells were harvested, washed with ice‐cold water, resuspended in 3 mL of water and boiled for 10 min. After the cell suspensions were centrifuged at 8000 × g for 5 min, the amounts of glycerol and trehalose were determined using an HPLC system (SCL‐10A VP; Shimadzu) equipped with an AMINEX HPX‐87H column (300 × 7.8 mm) (Bio‐Rad, Hercules, CA< USA) and an RID‐10A refractive index detector (Shimadzu) as described previously 19 . For glycerol detection, the HPLC system was operated at 65 °C with a mobile phase of 5 mmol L −1 H 2 SO 4 , a flow rate of 0.6 mL min −1 and an injection volume of 50 μL.…”