Med3-mediated NADPH generation to help tolerate hyperosmotic stress.

Hyperosmotic stress tolerance is crucial for in producing value-added products from renewable feedstock. The limited understanding of its tolerance mechanism has impeded the application of these microbial cell factories. Previous studies have shown that Med3 plays a role in hyperosmotic stress in . However, the specific function of Med3 in hyperosmotic stress tolerance remains unclear. In this study, we showed that the deletion of the mediator Med3 impairs growth under hyperosmotic stress. Phenotypic analyses and yeast two-hybrid assays revealed that Med3 interacts with the transcription factor Stb5 to regulate the expression of the genes and , which are involved in NADPH production under hyperosmotic stress conditions. The deletion of resulted in a decrease in intracellular NADPH content, leading to increased oxidative stress and elevated levels of intracellular reactive oxygen species under hyperosmotic stress, thereby impacting bud formation. These findings highlight the significant role of Med3 as a regulator in maintaining NADPH generation and redox homeostasis in during hyperosmotic stress.IMPORTANCEHyperosmotic stress tolerance in the host strain is a significant challenge for fermentation performance in industrial production. In this study, we showed that the mediator Med3 is essential for yeast growth under hyperosmotic conditions. Med3 interacts with the transcription factor Stb5 to regulate the expression of genes involved in the NADPH-generation system during hyperosmotic stress. Adequate NADPH ensures the timely removal of excess reactive oxygen species and supports bud formation under these conditions. This work highlights the crucial role of Med3 as a regulator in maintaining NADPH generation and redox homeostasis in during hyperosmotic stress.