Activation of Glut1 glucose transporter in human erythrocytes.
Glut1, the only glucose transporter isoform expressed in the human red blood cell (RBC), binds to and is inhibited by cytochalasin B (CB). In the present study we show that incubation of RBC ghost membranes with 10 microM cytochalasin E (CE) results in a 1.8-fold increase in the number of glucose-sensitive cytochalasin B binding sites. Moreover, treatment with CE was associated with no observable change in the protein composition of RBC ghosts determined by SDS-PAGE. Removal of surface ("extrinsic") proteins from RBC ghosts by treatment with 0.2 mM EDTA (pH 12) for 30 min resulted in a similar 1.8-fold increase in the number of glucose-sensitive CB binding sites. Western blot analysis showed that treatment with CE or EDTA resulted in no change in the amount or mobility of Glut1 present in ghost membranes. Glucose transport, measured as CB-inhibitable 3-O-[3H]methylglucose uptake by resealed ghosts, was stimulated in CE-treated resealed ghosts, with the t1/2 to equilibrium decreasing from 6.8 +/- 0.5 to 3.9 +/- 0.3 s (P < 0.05). Treatment of ghosts with CE or EDTA followed by Western blotting of samples in the presence or absence of beta-mercaptoethanol resulted in no change in immunoreactivity or mobility of the major Glut1 band. The above results suggest that a significant fraction of Glut1 transporters exists in an inactive form ("masked") in RBC plasma membranes and that treatment of ghosts with CE or EDTA leads to an apparent activation of Glut1.
Department of Physiology and Biophysics, Case Western Reserve University, Cleveland, Ohio, 44106-4951, USA.
J Z
JZ
Zhang
F
F
Ismail-Beigi
D001665:Binding Sites / Q000187:drug effects; D001692:Biological Transport / Q000187:drug effects; D003571:Cytochalasin B / Q000378:metabolism / Q000494:pharmacology; D003572:Cytochalasins / Q000378:metabolism / Q000494:pharmacology; D004910:Erythrocyte Membrane / Q000378:metabolism*; D005947:Glucose / Q000378:metabolism*; D051272:Glucose Transporter Type 1; D006801:Humans; D009004:Monosaccharide Transport Proteins / Q000097:blood* / Q000187:drug effects; D011485:Protein Binding
D003572:Cytochalasins; D051272:Glucose Transporter Type 1; D009004:Monosaccharide Transport Proteins; C495677:SLC2A1 protein, human; C007215:cytochalasin E; D003571:Cytochalasin B; D005947:Glucose