Resolution of the facilitated transport of dehydroascorbic acid from its intracellular accumulation as ascorbic acid.
We performed a detailed kinetic analysis of the uptake of dehydroascorbic acid by HL-60 cells under experimental conditions that enabled the differentiation of dehydroascorbic acid transport from the intracellular reduction/accumulation of ascorbic acid. Immunoblotting and immunolocalization experiments identified GLUT1 as the main glucose transporter expressed in the HL-60 cells. Kinetic analysis allowed the identification of a single functional activity involved in the transport of dehydroascorbic acid in the HL-60 cells. Transport was inhibited in a competitive manner by both 3-O-methyl-D-glucose and 2-deoxy-D-glucose. In turn, dehydroascorbic acid competitively inhibited the transport of both sugars. A second functional component identified in experiments measuring the accumulation of ascorbic acid appears to be associated with the intracellular reduction of dehydroascorbic acid to ascorbic acid and is not directly involved in the transport of dehydroascorbic acid via GLUT1. Transport of dehydroascorbic acid by HL-60 cells was independent of the presence of external Na+, whereas the intracellular accumulation of ascorbic acid was found to be a Na(+)-sensitive process. Thus, the transport of dehydroascorbic acid via glucose transporters is a Na(+)-independent process which is kinetically and biologically separable from the reduction of dehydroascorbic acid to ascorbic acid and its subsequent intracellular accumulation.
Vera
J C
JC
Program in Molecular Pharmacology and Therapeutics, Memorial-Sloan Kettering Cancer Center, New York, New York 10021, USA.
Rivas
C I
CI
Velásquez
F V
FV
Zhang
R H
RH
Concha
I I
II
Golde
D W
DW
D019325:3-O-Methylglucose; D001205:Ascorbic Acid / Q000378:metabolism*; D001667:Binding, Competitive; D001693:Biological Transport, Active; D002460:Cell Line; D003683:Dehydroascorbic Acid / Q000378:metabolism*; D003847:Deoxyglucose / Q000378:metabolism; D051272:Glucose Transporter Type 1; D006801:Humans; D007424:Intracellular Fluid / Q000378:metabolism; D007700:Kinetics; D008757:Methylglucosides / Q000378:metabolism; D008954:Models, Biological; D009004:Monosaccharide Transport Proteins / Q000378:metabolism; D010084:Oxidation-Reduction; D012964:Sodium / Q000378:metabolism
D051272:Glucose Transporter Type 1; D008757:Methylglucosides; D009004:Monosaccharide Transport Proteins; C495677:SLC2A1 protein, human; D019325:3-O-Methylglucose; D003847:Deoxyglucose; D012964:Sodium; D001205:Ascorbic Acid; D003683:Dehydroascorbic Acid