Comparative Genomic and Transcriptomic Analyses of 60Co-Mutagenized Scheffersomyces stipitis Strains: Identification of Candidate Genes Associated with High-Ethanol-Producing Xylose-to-Ethanol Fermentation

Sugarcane bagasse is an important renewable lignocellulosic resource, yet its bioconversion efficiency remains low, primarily because wild-type Saccharomyces cerevisiae cannot utilize xylose, which limits the industrial production of cellulosic ethanol. In this study, a high-ethanol-yielding strain 31.1 was obtained from Scheffersomyces stipitis (formerly known as Pichia stipitis) 1960 through 60Co mutagenesis and long-term domestication. Strain 31.1 exhibited an ethanol productivity of 0.78 g/(L·h), a sugar-to-ethanol conversion rate of 0.38 g/g, and a fermentation efficiency of 82.61%. Using the wild-type strain 1960 and a low-yielding strain 12.1 as controls, comparative genomics and transcriptomics were employed to elucidate the mechanism underlying the high ethanol production. Our findings are as follows: at the genomic level, there were 271 genomic structural variations. At the transcriptomic level, most genes involved in secondary metabolite synthesis, antibiotic synthesis, ribosomal pathways, amino acid biosynthesis, and oxidative phosphorylation pathways were down-regulated. Additionally, two key genes—XYL1 (xylose reductase gene) and XUT4 (high-affinity xylose transporter gene)—were significantly up-regulated. Through comprehensive integration of phenotypic comparison (e.g., fermentation performance of the high-yield strain 31.1 in yeast propagation and ethanol fermentation), comparative genomics, transcriptomics, and bioinformatics analyses of pathways involved in oxidative phosphorylation and the cell cycle (related to yeast cell growth), we identified 60 candidate key genes associated with high xylose-to-ethanol yield in S. stipitis These genes are predominantly involved in the cell cycle pathway, and several, including CDC15 and PHO4, have been experimentally validated. In conclusion, our study preliminarily reveals the mechanisms underlying the high xylose ethanol production of the high-yield strain at the genomic and transcriptomic levels.