Intracellular trehalose accumulation via the Agt1 transporter promotes freeze-thaw tolerance in Saccharomyces cerevisiae.

AIM: This study is to investigate the use of a constitutively expressed trehalose transport protein to directly control intracellular trehalose levels and protect baker's yeast (Saccharomyces cerevisiae) cells against freeze-thaw stress in vivo. METHODS AND RESULTS: We used a constitutively overexpressed Agt1 transporter to investigate the role of trehalose in the freeze-thaw tolerance of yeast cells by regulating intracellular trehalose concentrations independently of intracellular biosynthesis. Using this method, we found that increasing intracellular trehalose in yeast cells improved cell survival rate after 8 days of freezing at -80 and -20°C. We also observed that freeze-thaw tolerance promoted by intracellular trehalose only occurs in highly concentrated cell pellets rather than cells in liquid suspension. CONCLUSIONS: Trehalose is sufficient to provide freeze-thaw tolerance using our Agt1 overexpression system. Freeze-thaw tolerance can be further enhanced by deletion of genes encoding intracellular trehalose degradation enzymes. SIGNIFICANCE AND IMPACT OF STUDY: These findings are relevant to improving the freeze-thaw tolerance of baker's yeast in the frozen baked goods industry through engineering strains that can accumulate intracellular trehalose via a constitutively expressed trehalose transporter and inclusion of trehalose into the growth medium.