Using phosphoglucose isomerase-deficient ( pgi1 Δ) Saccharomyces cerevisiae to map the impact of sugar phosphate levels on d -glucose and d -xylose sensing

Background Despite decades of engineering efforts, recombinant Saccharomyces cerevisiae are still less efficient at converting d -xylose sugar to ethanol compared to the preferred sugar d -glucose. Using GFP-based biosensors reporting for the three main sugar sensing routes, we recently demonstrated that the sensing response to high concentrations of d -xylose is similar to the response seen on low concentrations of d -glucose. The formation of glycolytic intermediates was hypothesized to be a potential cause of this sensing response. In order to investigate this, glycolysis was disrupted via the deletion of the phosphoglucose isomerase gene ( PGI1 ) while intracellular sugar phosphate levels were monitored using a targeted metabolomic approach. Furthermore, the sugar sensing of the PGI1 deletants was compared to the PGI1 -wildtype strains in the presence of various types and combinations of sugars. Results Metabolomic analysis revealed systemic changes in intracellular sugar phosphate levels after deletion of PGI1 , with the expected accumulation of intermediates upstream of the Pgi1p reaction on d -glucose and downstream intermediates on d -xylose. Moreover, the analysis revealed a preferential formation of d -fructose-6-phosphate from d -xylose, as opposed to the accumulation of d -fructose-1,6-bisphosphate that is normally observed when PGI1 deletants are incubated on d -fructose. This may indicate a role of PFK27 in d -xylose sensing and utilization. Overall, the sensing response was different for the PGI1 deletants, and responses to sugars that enter the glycolysis upstream of Pgi1p ( d -glucose and d -galactose) were more affected than the response to those entering downstream of the reaction ( d -fructose and d -xylose). Furthermore, the simultaneous exposure to sugars that entered upstream and downstream of Pgi1p ( d -glucose with d -fructose, or d -glucose with d -xylose) resulted in apparent synergetic activation and deactivation of the Snf3p/Rgt2p and cAMP/PKA pathways, respectively. Conclusions Overall, the sensing assays indicated that the previously observed d -xylose response stems from the formation of downstream metabolic intermediates. Furthermore, our results indicate that the metabolic node around Pgi1p and the level of d -fructose-6-phosphate could represent attractive engineering targets for improved d -xylose utilization.