Multi-Omics Dissection and Functional Validation of Candidate Regulators Modulating Stress Tolerance and Xylose...

The intrinsic weakness of the budding yeast Saccharomyces cerevisiae in xylose utilization limits its application in biological manufacturing using lignocellulosic biomass. Although the natural yeast strain S. cerevisiae YB-2625 exhibits superior innate xylose-fermenting capability, the underlying mechanisms remain largely unexplored. Here, we employed comparative multi-omics to systematically dissect the molecular basis of its high stress tolerance and superior xylose consumption. Comparative genomics revealed 73,842 single nucleotide polymorphisms (SNPs) and 5191 small insertions/deletions (InDels) in YB-2625 relative to S288C, with significant enrichment in genes associated with chromatin remodeling, transcriptional regulation, and stress signaling. Integration of genomic and transcriptomic data identified candidate variants in key regulators. Functional validation further demonstrated that Tra1, a component of the SAGA, SLIK, and NuA4 histone acetyltransferase complexes, acts as a global regulator with growth-coupled effects on stress tolerance and xylose metabolism. Deletion of TRA1 significantly reduced the final biomass in xylose medium. Moreover, deletion of RTT109 specifically impaired growth on xylose without affecting any of the tested stress tolerance phenotypes. We further examined global chromatin accessibility changes upon deletion of the histone acetyltransferase gene NGG1, a manipulation previously shown to substantially enhance xylose utilization in the engineered YB-2625 background. ATAC-seq analysis revealed that loss of Ngg1 alters chromatin accessibility at loci governing carbohydrate metabolism and stress responses, thereby establishing a direct link between epigenetic remodeling and the superior phenotype of YB-2625. Our findings provide a basis for deciphering the regulatory circuitry governing xylose utilization in recombinant yeast and for the rational engineering of robust strains for lignocellulosic bioconversion.