Deletion Of JJJ1 Improved Acetic Acid Tolerance And Cellulosic Ethanol Fermentation Performance Of Saccharomyces Cerevisiae Strains

Pretreatment process of lignocellulosic feedstock produces a variety of inhibitors such as weak acids, furan derivatives, and phenolic compounds that influence the growth and fermentation rate of saccharomyces cerevisiae. Acetic acid represents a major inhibitor result from degradation of hemicellulose. Undissociated acetic acids would permeate into yeast cell membrane by simple diffusion, acidify cell cytoplasm, thereby inhibit cell metabolism and cell growth, which have a serious impact on utilization of this renewable energy in the bio-ethanol production. Thus, elucidating the molecular mechanisms of acetic acid tolerance in S.cerevisiae is of great importance to the construction of more robust yeast strains on industrial bioethanol production.Our previous study shows that when yeast cells grew in lignocellulosic hydrolyzates, gene expression of JJJ1 were down-regulated. To figure out the possible role of JJJ1 under lignocellulosic hydrolyzates, in this study, we first compared the tolerance of various environmental stresses between strain BY4741 and strain BYΔJJJJ. Improved resistance to acetic acid and formic acid was observed in BYΔJJJ1. Also, BYAJJJ1 showed a better fermentation performance than that of BY4741 in 5.5g/L acetic acid,1.2g/L formic acid or lignocellulosic hydrolyzates mixed with various inhibitors (3.0g/L acetic acid,0.2g/L formic acid,1.1 g/L furfural and 0.2g/L HMF) fermentation medium. The fermentation rate was significantly enhanced in BYΔJJJ1 than in BY4741. Knockout of JJJ1 in industrial strains YJSH1 also improved its fermentation rate. Under acetic acid stress, BYAJJJ1 cells accumulated more trehalose but less acetic acid compared to its parent strain. Additionally, higher proportion of long chain fatty acids and relatively lower ergosterol, and higher activity of SOD and CAT was observed in BYAJJJ1. Using the RNA-Seq technology, we analyzed the transcriptome of strains BY4741 and BYΔJJJ1. Compared to BY4741,210 genes are differentially expressed in BYΔJJJ1. Cluster analysis of these genes on DAVID database shows genes related to the carbohydrate metabolism and membrane transport protein were down-regulated, while the cell wall-related and ribosome functional gene expressions were up-regulated in BYAJJJ1. Combined with the physiological and biochemical differences between the two strains, we speculate the up-regulated expression of cell wall-related genes remodeled cell wall, thus decreasing diffusion of acetic acid into the yeast cell. The decreased expression of glucose metabolism related genes in BYAJJJ1 might explain its slower growth rate under normal conditions, but faster growth rate under acetic acid stress.The results provide both theoretical guidance and technical support for the optimization of cellulosic ethanol fermentation and construction of more robust industrial strains of Saccharomyces cerevisiae.