Bl. Ing et al., CHARACTERIZATION OF A MUTANT GLUT4 LACKING THE N-GLYCOSYLATION SITE -STUDIES IN TRANSFECTED RAT ADIPOSE-CELLS, Biochemical and biophysical research communications, 218(1), 1996, pp. 76-82
GLUT4, the insulin-responsive glucose transporter expressed primarily
in muscle and adipose tissue, contains a single N-glycosylation sire.
We characterized a mutant GLUT4 lacking the N-glycosylation site (Asn(
57) --> Gin) in primary cultures of rat adipose cells, We transiently
transfected cells with expression vectors for epitope-tagged GLUT4 con
taining either wild-type (GLUT4-HA) or mutant (GLN57-HA) cDNA sequence
s, Expression of GLN57-HA in adipose cells was similar to 10-fold lowe
r than for GLUT4-HA even though mRNA levels for bath recombinant trans
porters were comparable, Biosynthetic labeling studies showed markedly
decreased incorporation of [S-35]-methionine/cysteine into GLN57-HA r
elative to GLUT4-HA consistent with either a decreased synthetic rate
or accelerated degradation of GLN57-HA. Interestingly, transient trans
fection of GLUT4-HA and GLN57-HA in COS-7 cells (which do not express
endogenous GLUT4) resulted in comparable levels of protein expression
for both transporters. Thus, in the physiologically relevant adipose c
ell, glycosylation of GLUT4 appears to play an important functional ro
le. (C) 1996 Academic Press, Inc.