An arginine to cysteine(252) mutation in insulin receptors from a patient with severe insulin resistance inhibits receptor internalisation but preserves signalling events.
OBJECTIVE
We examined the properties of a mutant insulin receptor (IR) with an Arg(252) to Cys (IR(R252C)) substitution in the alpha-subunit originally identified in a patient with extreme insulin resistance and acanthosis nigricans.


METHODS
We studied IR cell biology and signalling pathways in Chinese Hamster Ovary cells overexpressing this IR(R252C).


RESULTS
Our investigation showed an impairment in insulin binding to IR(R252C) related mostly to a reduced affinity of the receptor for insulin and to a reduced rate of IR(R252C) maturation; an inhibition of IR(R252C)-mediated endocytosis resulting in a decreased insulin degradation and insulin-induced receptor down-regulation; a maintenance of IR(R252C) on microvilli even in the presence of insulin; a similar autophosphorylation of mutant IR(R252C) followed by IRS 1/IRS 2 phosphorylation, p85 association with IRS 1 and IRS 2 and Akt phosphorylation similar to those observed in cells expressing wild type IR (IRwt); and finally, a reduced insulin-induced Shc phosphorylation accompanied by decreased ERK1/2 phosphorylation and activity and of thymidine incorporation into DNA in cells expressing IR(R252C) as compared to cells expressing IRwt.


CONCLUSIONS
These observations suggest that: parameters other than tyrosine kinase activation participate in or control the first steps of IR internalisation or both; IR-mediated IRS 1/2 phosphorylation can be achieved from the cell surface and microvilli in particular; Shc phosphorylation and its subsequent signalling pathway might require IR internalisation; defective IR endocytosis correlates with an enhancement of some biological responses to insulin and attenuation of others.
Hamer
I
I
Department of Morphology, University of Geneva, Switzerland.
Foti
M
M
Emkey
R
R
Cordier-Bussat
M
M
Philippe
J
J
De Meyts
P
P
Maeder
C
C
Kahn
C R
CR
Carpentier
J-L
JL
D000052:Acanthosis Nigricans / Q000235:genetics*; D000328:Adult; D019943:Amino Acid Substitution; D000818:Animals; D001120:Arginine; D016466:CHO Cells; D006224:Cricetinae; D003545:Cysteine; D004247:DNA / Q000096:biosynthesis; D006801:Humans; D007328:Insulin / Q000378:metabolism; D007333:Insulin Resistance / Q000235:genetics*; D008297:Male; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D009154:Mutation; D010766:Phosphorylation; D021122:Protein Subunits; D021381:Protein Transport; D011972:Receptor, Insulin / Q000235:genetics* / Q000378:metabolism* / Q000502:physiology; D011994:Recombinant Proteins / Q000378:metabolism; D015398:Signal Transduction / Q000502:physiology*; D013936:Thymidine / Q000378:metabolism; D014162:Transfection
D007328:Insulin; D021122:Protein Subunits; D011994:Recombinant Proteins; D004247:DNA; D001120:Arginine; D011972:Receptor, Insulin; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases; D003545:Cysteine; D013936:Thymidine