Category:Team Red B 2019

In Figure 2, wild type C. japonicus and cel3b, cel3Acel3B, and 4BG mutants were grown on cellobiose and glucose (control). The four strains showed no significant changes in glucose consumption during growth (Fig. 2A & 2B). However, the cel3b mutant showed a decreased rate of cellobiose consumption (Fig. 2C & 2D). This shows that cel3b has a role in efficient cellobiose utilization in C. japonicus.

B-glucuronidase activity was used to determine relative gene expression levels of uidA+ transcription fusion proteins, Smc00887 and Smc00888 (Fig 2). Smc00888 encodes a histidine kinase. During stationary phase, SMc00888 locus expression decreased three-fold in the cbrA::Tn5 mutant compared to WT.

To demonstrate specificity of the sweet taste receptor, the pathway of G-protein coupled receptors linked to the receptor was modified to increase intracellular calcium which was observed using the fluorescent FURA-2 calcium indicator dye. An increase in cellular calcium was observed (normalized to responses observed from the highest concentration per compound) in the sweet compounds GA-2, saccharin, acesulfame-K, and sucrose in the receptor combination T1R2+3 (Fig 6b), matching the biological thresholds observed in vivo (Fig 6a). Dose responses were not observed for bitter or umami compounds (Fig 6b). As intracellular calcium was not shown to increase upon exposure to any compound other than sugar and artificial sweeteners, the function of T1Rs as a sweet taste receptors is maintained.

Dose responses (normalized to responses observed from the highest concentration per compound) were observed for the sweet compounds GA-2, saccharin, acesulfame-K, and sucrose in the receptor combination T1R2+3 (Fig 6b), matching the biological thresholds observed in vivo (Fig 6a). Dose responses were not observed for bitter or umami compounds (Fig 6b)