Receptor-mediated internalization of insulin. Potential role of pp120/HA4, a substrate of the insulin receptor kinase.
pp120/HA4 is a hepatocyte membrane glycoprotein phosphorylated by the insulin receptor tyrosine kinase. In this study, we have investigated the role of pp120/HA4 in insulin action. Transfection of antisense pp120/HA4 cDNA in H35 hepatoma cells resulted in inhibition of pp120/HA4 expression and was associated with a 2-3-fold decrease in the rate of insulin internalization. Furthermore, insulin internalization in NIH 3T3 fibroblasts co-transfected with insulin receptors and pp120/HA4 was increased 2-fold compared with cells expressing insulin receptors alone. In contrast, no effect on internalization was observed in cells overexpressing a naturally occurring splice variant of pp120/HA4 that lacks the phosphorylation sites in the intracellular domain. Insulin internalization was also unaffected in cells expressing three site-directed mutants of pp120/HA4 in which the sites of phosphorylation by the insulin receptor kinase had been removed (Y488F, Y488F/Y513F, and S503A). Our data suggest that pp120/HA4 is part of a complex of proteins required for receptor-mediated internalization of insulin. It is possible that this function is regulated by insulin-induced phosphorylation of the intracellular domain of pp120/HA4.
Formisano
P
P
Diabetes Branch, NIDDK, National Institutes of Health, Bethesda, Maryland 20892, USA.
Najjar
S M
SM
Gross
C N
CN
Philippe
N
N
Oriente
F
F
Kern-Buell
C L
CL
Accili
D
D
Gorden
P
P
D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies; D002352:Carrier Proteins / Q000096:biosynthesis / Q000378:metabolism*; D002453:Cell Cycle; D002455:Cell Division; D006913:Hydroxysteroid Dehydrogenases; D007328:Insulin / Q000378:metabolism*; D007700:Kinetics; D008099:Liver / Q000378:metabolism; D008562:Membrane Glycoproteins / Q000378:metabolism*; D051379:Mice; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010446:Peptide Fragments / Q000737:chemistry; D010766:Phosphorylation; D010982:Platelet-Derived Growth Factor / Q000378:metabolism; D016372:RNA, Antisense; D051381:Rats; D011972:Receptor, Insulin / Q000378:metabolism*; D017479:Receptors, Platelet-Derived Growth Factor / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D013936:Thymidine / Q000378:metabolism; D014162:Transfection; D045002:3-alpha-Hydroxysteroid Dehydrogenase (B-Specific)
D000906:Antibodies; D002352:Carrier Proteins; D006913:Hydroxysteroid Dehydrogenases; D007328:Insulin; D008562:Membrane Glycoproteins; D010446:Peptide Fragments; D010982:Platelet-Derived Growth Factor; D016372:RNA, Antisense; D011972:Receptor, Insulin; D017479:Receptors, Platelet-Derived Growth Factor; D011994:Recombinant Proteins; D013936:Thymidine; C037643:bile acid binding proteins; C435496:AKR1C2 protein, human; D045002:3-alpha-Hydroxysteroid Dehydrogenase (B-Specific)