Mutation in a conserved motif next to the insulin receptor key autophosphorylation sites de-regulates kinase activity and impairs insulin action.
We have recently reported two non-insulin-dependent diabetic patients exhibiting a heterozygous point mutation (R1152-Q) next to the key tyrosine autophosphorylation sites (Y1146, Y1150, Y1151) of the insulin receptor. In the present study, we demonstrate that the Q1152 mutation alters a previously unrecognized consensus sequence in the insulin receptor family of tyrosine kinases. To define the effect of this alteration on insulin receptor function, the mutant insulin receptor (Q1152) was constructed and overexpressed in NIH-3T3 cells. In spite of normal insulin binding, "in vivo" and "in vitro" autophosphorylation as well as transphosphorylation by the wild-type receptor (WT) were deficient in Q1152 as compared with the transfected WT receptors. Insulin-stimulated kinase activity toward poly(Glu, Tyr) 4:1 and the endogenous substrates p120 and p175 were also impaired in Q1152. However, insulin-independent kinase activity of Q1152 was 2-5-fold higher than that of WT. While insulin stimulated 2-deoxyglucose uptake and glycogen synthase activity in WT-transfected cells with a sensitivity proportional to receptor number, no insulin stimulation was observed in Q1152 cells. Similar to the kinase, insulin-independent glycogen synthase activity and 2-deoxyglucose uptake were 2-fold higher in Q1152 than in either WT or parental cells. We conclude that the Q1152 mutation deregulates insulin receptor kinase and generates insulin insensitivity in cells. Alterations in this highly conserved region of the insulin receptor may contribute to non-insulin dependent diabetes mellitin pathogenesis in humans.
Formisano
P
P
Centro di Endocrinologia ed Oncologia Sperimentale del Consiglio Nazionale delle Ricerche, Milano, Italy.
Sohn
K J
KJ
Miele
C
C
Di Finizio
B
B
Petruzziello
A
A
Riccardi
G
G
Beguinot
L
L
Beguinot
F
F
D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D004277:DNA, Single-Stranded; D006801:Humans; D007328:Insulin / Q000378:metabolism*; D051379:Mice; D008969:Molecular Sequence Data; D009154:Mutation; D010766:Phosphorylation; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D011972:Receptor, Insulin / Q000235:genetics / Q000378:metabolism*; D012689:Sequence Homology, Nucleic Acid; D014162:Transfection
D004277:DNA, Single-Stranded; D007328:Insulin; D011505:Protein-Tyrosine Kinases; D011972:Receptor, Insulin