Evaluating Sugar Signaling during Xylose Oxidation in Saccharomyces cerevisiae

Sammanfattning: The sugar signaling state of cells oxidizing xylose was investigated through the chromosomal integration of the Weimberg pathway into previously established biosensor strains capable of monitoring the Snf3p/Rgt2p, SNF1/Mig1p, and cAMP/PKA pathways through the expression of HXT1, SUC2, and TPS1, respectively. Ultimately, two new strains were successfully developed containing both the Weimberg pathway and a biosensor for either HXT1 (TMB CB 112) or TPS1 (TMB CB 117). Unfortunately, the SUC2-based Weimberg biosensor (TMB CB 115) showed impaired growth and seemingly lacked a diauxic shift, making it unable to assimilate xylose. As for xylose utilization, the working Weimberg biosensor strains showed xylose assimilation rates similar to the benchmark Weimberg pathway strain (TMB4586), which remained lower than that of the oxidoreductase pathway biosensor strains (TMB375X series). Aside from the difference in xylose assimilation, the growing Weimberg biosensor strains showed very similar metabolite profiles to both the reference Weimberg pathway strain (TMB4586) and the oxidoreductase biosensor strains (TMB375X series). A signaling assay over a 54-hour cultivation showed clear similarities between the Weimberg biosensors (TMB CB 112, TMB CB 117) and the oxidoreductase biosensors (TMB375X series), and the induction of SUC2 after glucose depletion indicated that the full “low glucose” signal may only arise once xylose begins being actively metabolized. Finally, a separate 6-hour cultivation was performed in different key media of interest. Interestingly, the Weimberg biosensor strains showed decreased bimodalities similar to the decrease seen after integration of the oxidoreductase pathway in the TMB375X series, indicating that integration of the Weimberg pathway may have a similar effect.