GLUT-2 gene transfer into insulinoma cells confers both low and high affinity glucose-stimulated insulin release. Relationship to glucokinase activity.
The rat insulinoma cell line RIN 1046-38 loses glucose-stimulated insulin secretion as a function of time in culture. We found that the loss of glucose sensing in these cells was correlated with the loss of expression of GLUT-2 and glucokinase. Stable transfection of RIN cells with a plasmid containing the GLUT-2 cDNA conferred glucose-stimulated insulin release in intermediate but not high passage cells, with the near-maximal 3-fold increase occurring at 50 microM glucose. GLUT-2 expressing cells also exhibited a larger response to the combination of 5 mM glucose + 1 microM forskolin than untransfected cells (7.9 versus 1.6-2.7-fold, respectively). GLUT-2 expressing intermediate passage, but not high passage, RIN cells exhibited a 4-fold increase in glucokinase enzymatic activity relative to nonexpressing controls. Glucokinase activity was also increased by transfer of the GLUT-2 gene into intermediate passage RIN cells via recombinant adenovirus. Preincubation of GLUT-2 expressing intermediate passage RIN cells with 2-deoxyglucose to inhibit low Km hexokinases resulted in a glucose-stimulated insulin secretion response that was shifted toward the physiologic range. These studies indicate that GLUT-2 expression confers both a high and low affinity glucose-stimulated insulin secretion response to intermediate passage RIN cells.
Ferber
S
S
Gifford Laboratories for Diabetes Research, University of Texas, Dallas 75235.
BeltrandelRio
H
H
Johnson
J H
JH
Noel
R J
RJ
Cassidy
L E
LE
Clark
S
S
Becker
T C
TC
Hughes
S D
SD
Newgard
C B
CB
D019325:3-O-Methylglucose; D000256:Adenoviridae; D000818:Animals; D001692:Biological Transport; D002460:Cell Line; D005576:Colforsin / Q000494:pharmacology; D015870:Gene Expression; D005822:Genetic Vectors; D005941:Glucokinase / Q000096:biosynthesis / Q000378:metabolism; D005947:Glucose / Q000378:metabolism / Q000494:pharmacology*; D051273:Glucose Transporter Type 2; D006593:Hexokinase / Q000378:metabolism; D006801:Humans; D007328:Insulin / Q000378:metabolism*; D000078790:Insulin Secretion; D007340:Insulinoma / Q000378:metabolism*; D007700:Kinetics; D008757:Methylglucosides / Q000378:metabolism; D009004:Monosaccharide Transport Proteins / Q000096:biosynthesis / Q000378:metabolism*; D010190:Pancreatic Neoplasms / Q000378:metabolism*; D012333:RNA, Messenger / Q000032:analysis / Q000096:biosynthesis; D051381:Rats; D014162:Transfection; D014407:Tumor Cells, Cultured
D051273:Glucose Transporter Type 2; D007328:Insulin; D008757:Methylglucosides; D009004:Monosaccharide Transport Proteins; D012333:RNA, Messenger; D019325:3-O-Methylglucose; D005576:Colforsin; D006593:Hexokinase; D005941:Glucokinase; D005947:Glucose