Comprehensive Characterization of Mutant Pichia stipitis Co-Fermenting Cellobiose and Xylose through Genomic and Transcriptomic Analyses

The development of a yeast strain capable of fermenting mixed sugars efficiently is crucial for producing biofuels and value-added materials from cellulosic biomass. Previously, a mutant Pichia stipitis YN14 strain capable of co-fermenting xylose and cellobiose was developed through evolutionary engineering of the wild-type P. stipitis CBS6054 strain, which was incapable of co-fermenting xylose and cellobiose. In this study, through genomic and transcriptomic analyses, we sought to investigate the reasons for the improved sugar metabolic performance of the mutant YN14 strain in comparison with the parental CBS6054 strain. Unfortunately, comparative whole-genome sequencing (WGS) showed no mutation in any of the genes involved in the cellobiose metabolism between the two strains. However, comparative RNA sequencing (RNA-seq) revealed that the YN14 strain had 101.2 times and 5.9 times higher expression levels of HXT2.3 and BGL2 genes involved in cellobiose metabolism, and 6.9 times and 75.9 times lower expression levels of COX17 and SOD2.2 genes involved in respiration, respectively, compared with the CBS6054 strain. This may explain how the YN14 strain enhanced cellobiose metabolic performance and shifted the direction of cellobiose metabolic flux from respiration to fermentation in the presence of cellobiose compared with the CBS6054 strain.