Role of actin in the cAMP-dependent activation of sodium/glucose cotransporter in renal epithelial cells.
We examined whether actin filaments are involved in the cAMP-dependent activation of a high affinity sodium/glucose cotransporter (SGLT1) using epithelial expression systems. The expression of enhanced green fluorescent protein-tagged SGLT1 (EGFP-SGLT1) in Madin-Darby canine kidney (MDCK) cells was revealed by Western blotting and confocal laser microscopy. 8-Br-cAMP, a membrane permeable cAMP analog, enhanced [14C]-alpha-methyl glucopyranoside ([14C]-AMG) uptake. Both basal and 8-Br-cAMP-elicited [14C]-AMG uptakes were inhibited by N-(2{[3-(4-bromophenyl)-2-propenyl]-amino}-ethyl)-5-isoquinolinesulfonamide (H-89), a protein kinase A inhibitor, and cytochalasin D, an actin filament formation inhibitor. Furthermore, cytochalasin D inhibited the distribution of EGFP-SGLT1 at the apical surface. These results suggest that the EGFP-SGLT1 protein is functionally expressed in the apical membrane of MDCK cells, and is up-regulated by a cAMP-dependent pathway requiring intact actin filaments.
Ikari
Akira
A
Department of Environmental Biochemistry and Toxicology, University of Shizuoka School of Pharmaceutical Sciences, 52-1 Yada, Shizuoka 422-8526, Japan. ikari@u-shizuoka-ken.ac.jp
Harada
Hitoshi
H
Takagi
Kuniaki
K
D015124:8-Bromo Cyclic Adenosine Monophosphate / Q000378:metabolism; D000199:Actins / Q000378:metabolism*; D000818:Animals; D000242:Cyclic AMP / Q000378:metabolism*; D015638:Cytochalasin D / Q000494:pharmacology; D004285:Dogs; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism*; D017930:Genes, Reporter; D006801:Humans; D007668:Kidney / Q000187:drug effects / Q000378:metabolism*; D008562:Membrane Glycoproteins / Q000235:genetics* / Q000378:metabolism; D009004:Monosaccharide Transport Proteins / Q000235:genetics* / Q000378:metabolism; D019384:Nucleic Acid Synthesis Inhibitors / Q000494:pharmacology; D011817:Rabbits; D051381:Rats; D051269:Sodium-Glucose Transporter 1; D014981:Xenopus
D000199:Actins; D008562:Membrane Glycoproteins; D009004:Monosaccharide Transport Proteins; D019384:Nucleic Acid Synthesis Inhibitors; C495674:SLC5A1 protein, human; C495676:Slc5a1 protein, rat; D051269:Sodium-Glucose Transporter 1; D015638:Cytochalasin D; D015124:8-Bromo Cyclic Adenosine Monophosphate; D000242:Cyclic AMP