Evaluation Of Yeast Growth,Carbon Metabolism And Cell Cycle Under Trehalose Addition

Trehalose is an important cytoprotective agent,which stabilizes the structure of cell membrane under extreme conditions such as cold,dehydration and oxidation to protect cells,and can also acts as a backup carbon source when energy is insufficient during cell growth and development.Numerous studies have demonstrated the protective effect of trehalose on yeast cells under extreme environments.However,studies about the effect of exogenous trehalose addition in the initial fermentation medium on yeast cells are limited.In this study,CEN.PK 113-7D was selected as the experimental organism.Different concentrations of exogenous trehalose were added to the fermentation initial medium to evaluate the influence of cellular hexose transport,glucose metabolism,ATP productivity and cell cycle.This study provided clearer insights to understand the effect of exogenous trehalose on the yeast initial fermentation process in the medium.First,the yeast cells were cultured in the original YPD medium fortified with different concentrations(0 g,2 g,5 g per 100 mL)of exogenous trehalose.Then the fermentation biomass of yeast with ethanol and non-ethanol was measured,and the effects of exogenous trehalose on the intracellular glucose and trehalose content of yeast cells were analyzed.We evaluated the genes expression involved in hexose transport,gene expression was used to initially assess the effect of exogenous trehalose on glucose transport in yeast cells during fermentation.The results showed that the addition of exogenous trehalose to the initial medium had a significant effect on the growth of yeast cells,intracellular trehalose and glucose content,and the expression of key genes encoding hexose transportersIn addition,we also analyzed the influence of different concentrations of trehalose in the initial medium on the key genes for trehalose cycle,glycolysis,tricarboxylic acid cycle,respiratory chain and ATP synthesis.We also evaluate the effect of different concentrations of trehalose added to the initial medium on the intracellular ATP level of yeast cells.The results showed that the addition of different concentrations of trehalose to the initial medium could disrupt the balance in trehalose cycle and glycolysis of yeast cells.tricarboxylic acid cycle and the synthesis of ATP,and also reduces the intracellular ATP content.Finally,the effect of different concentrations of exogenous trehalose added in the initial medium on the expression of key genes encoding Gi/S,S,G2/M phase protein in yeast cells involved in the cell cycle was determined.At the same time,we also evaluated the important limiting factors of G1/S transition:protein content and cell size.Exogenous trehalose were added in the initial medium for protein chromogenic assay in order to evaluate the intracellular protein content.Exogenous trehalose on the size of yeast cell was observed through electron microscopy.PI staining were used to analyze the percentage of cell cycle distribution in yeast initial and exponential stages of growth upon incubating in medium supplemented with exogenous trehalose.The results showed that different concentrations of exogenous trehalose significantly inhibited the expression of genes related to G1/S and S cycle progression in yeast cells,while no significant effect on G2/M.The addition of exogenous trehalose to the initial medium significantly reduced the protein content of the yeast cells during the exponential phase of growth and development,and had no significant effect on cell size.In addition,exogenous trehalose has a different degree of inhibition on the cell cycle progression in the initial and middle stages of yeast cell fermentation.