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Figure 3 | BMC Evolutionary Biology

Figure 3

From: Evolutionary ancestry and novel functions of the mammalian glucose transporter (GLUT) family

Figure 3

Effects of pH and glucose gradients on GLUT12-mediated glucose transport. (a) Facilitative transport occurs down a glucose gradient at pH 7.4 and pH 5.0. (b) Active glucose transport occurs against a glucose gradient at pH 5.0 only. Representative experiments are shown. Bars represent mean 2-deoxy-D-glucose uptake with data point range plotted (n = 3). P values < 0.05 were considered significant and are shown on the graphs. (c, d) Immunofluoresence detection demonstrates apical localization of endogenous GLUT12 under all conditions used for glucose uptake assays (X-Y and X-Z planes shown). GLUT12, red; polarization of MDCK cells was determined by immunofluorescent labelling of the tight junction marker ZO-1, green. Bar, 20 μm. (e, f) Flow cytometry recorded intracellular pH of MDCK cells under conditions used in glucose uptake assays. Mean and SD were calculated (n = 6). A representative experiment is shown. # Significant decrease in pH compared to all other treatment groups. P = 0.0001. (g) Glucose uptake assays were performed in MDCK cells expressing endogenous GLUT12 and retrovirally transfected GLUT3. 2-deoxy-D-glucose transport with a glucose gradient at both pH 7.4 and pH 5.0 was significantly increased in MDCK cells retrovirally expressing GLUT3, in comparison to MDCK cells expressing endogenous GLUT12 only. (h) In cells retrovirally expressing GLUT3 no significant increase in 2-deoxy-D-glucose transport against a glucose gradient at pH 7.4 or pH 5.0 was recorded. Representative experiments are shown. Bars represent fold increase in 2-deoxy-D-glucose uptake with data point range plotted (n = 3). P values of < 0.05 were considered significant and are shown on the graphs. (i, j) Immunofluorescence detection of retrovirally transfected GLUT3 at the apical membrane of MDCK cells under conditions used for glucose uptake assays. GLUT3, red; tight junctional marker ZO-1, green. Bar, 20 μm.

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