Insulin-stimulated phosphorylation of a Rab GTPase-activating protein regulates GLUT4 translocation.
Insulin stimulates the rapid translocation of intracellular glucose transporters of the GLUT4 isotype to the plasma membrane in fat and muscle cells. The connections between known insulin signaling pathways and the protein machinery of this membrane-trafficking process have not been fully defined. Recently, we identified a 160-kDa protein in adipocytes, designated AS160, that is phosphorylated by the insulin-activated kinase Akt. This protein contains a GTPase-activating domain (GAP) for Rabs, which are small G proteins required for membrane trafficking. In the present study we have identified six sites of in vivo phosphorylation on AS160. These sites lie in the motif characteristic of Akt phosphorylation, and insulin treatment increased phosphorylation at five of the sites. Expression of AS160 with two or more of these sites mutated to alanine markedly inhibited insulin-stimulated GLUT4 translocation in 3T3-L1 adipocytes. Moreover, this inhibition did not occur when the GAP function in the phosphorylation site mutant was inactivated by a point mutation. These findings strongly indicate that insulin-stimulated phosphorylation of AS160 is required for GLUT4 translocation and that this phosphorylation signals translocation through inactivation of the Rab GAP function.
Sano
Hiroyuki
H
Department of Biochemistry, Dartmouth Medical School, Hanover, New Hampshire 03755, USA.
Kane
Susan
S
Sano
Eiko
E
Mîinea
Cristinel P
CP
Asara
John M
JM
Lane
William S
WS
Garner
Charles W
CW
Lienhard
Gustav E
GE
D016475:3T3 Cells; D000818:Animals; D001665:Binding Sites; D020690:GTPase-Activating Proteins / Q000235:genetics / Q000378:metabolism*; D051275:Glucose Transporter Type 4; D007328:Insulin / Q000494:pharmacology*; D051379:Mice; D008856:Microscopy, Fluorescence; D009004:Monosaccharide Transport Proteins / Q000378:metabolism*; D009124:Muscle Proteins; D009154:Mutation; D010766:Phosphorylation / Q000187:drug effects; D017434:Protein Structure, Tertiary; D020691:rab GTP-Binding Proteins / Q000378:metabolism*
D020690:GTPase-Activating Proteins; D051275:Glucose Transporter Type 4; D007328:Insulin; D009004:Monosaccharide Transport Proteins; D009124:Muscle Proteins; C495693:Slc2a4 protein, mouse; D020691:rab GTP-Binding Proteins