Expression of GFP-tagged low affinity Na+-dependent glucose transporter in Xenopus oocytes and CHO cells.
A low-affinity and high-capacity Na(+)-dependent glucose transporter (SGLT2) was inserted into the expression vector tagging of green fluorescent protein (EGFP). The protein expression and glucose transport activity were examined in Xenopus oocytes and Chinese hamster ovary (CHO) cells. In Western blotting analysis, EGFP-tagged SGLT2 protein expressed in both Xenopus oocytes and CHO cells. We also observed the EGFP fluorescence in both cells with a confocal laser microscope. To determine the function of EGFP-tagged SGLT2, we measured the uptake of [(14)C]-alpha-methyl glucopyranoside (AMG), a specific substrate for SGLT. The AMG uptake was time-dependently increased and inhibited by phloridzin in the EGFP-tagged SGLT2-expressing cells. The K(m) value of 1.7 mM for AMG and the IC(50) of 2 microM for phloridzin consist with the renal low affinity Na(+)-dependent glucose transporter. These results indicate that EGFP-tagged SGLT2 protein functionally expressed both in Xenopus oocytes and CHO cells, and these models are useful for studying the regulatory mechanisms of glucose reabsorption.
Ikari
Akira
A
Department of Environmental Biochemistry and Toxicology, University of Shizuoka School of Pharmaceutical Sciences, Shizuoka, 422-8526 Japan. ikari@u-shizuoka-ken.ac.jp
Suketa
Yasunobu
Y
D000818:Animals; D015153:Blotting, Western; D016466:CHO Cells; D006224:Cricetinae; D005947:Glucose / Q000493:pharmacokinetics*; D049452:Green Fluorescent Proteins; D007202:Indicators and Reagents; D008164:Luminescent Proteins; D009004:Monosaccharide Transport Proteins / Q000096:biosynthesis* / Q000494:pharmacology; D009865:Oocytes; D012964:Sodium; D014981:Xenopus
D007202:Indicators and Reagents; D008164:Luminescent Proteins; D009004:Monosaccharide Transport Proteins; D049452:Green Fluorescent Proteins; D012964:Sodium; D005947:Glucose