Impact of mutations at different serine residues on the tyrosine kinase activity of the insulin receptor.
Insulin binding to its receptor activates a cascade of signaling events which are initiated by tyrosine autophosphorylation of the receptor and activation of the tyrosine kinase activity towards the insulin receptor substrates. In addition to phosphorylation at tyrosine residues a serine phosphorylation of the insulin receptor is observed. Neither the functional significance of serine phosphorylation of the receptor nor the location of relevant regulatory sites has been determined exactly so far. We studied potential functions of serine residues in human insulin receptor (HIR) with respect to its ability to undergo insulin stimulated autophosphorylation. Using site directed mutagenesis of HIR we exchanged serine to alanine at 13 different positions in the HIR beta-subunit. Sites were chosen according to the criteria of known serine phosphorylation sites (1023/25, 1293/94, 1308/09), conserved positions in hIR, hIGF-1 receptor, hIRR, and dIR (962, 994, 1037, 1055, 1074/78, 1168, 1177/78/82, 1202, 1263, 1267). All HIR mutants were expressed in HEK 293 cells and basal and insulin stimulated autophosphorylation were determined. We found that the exchange of serine to alanine at position 994 and at position 1023/25 increased insulin stimulated receptor autophosphorylation significantly (147% +/- 12% and 129% +/- 6% of control, p < 0.01, n = 7), while all other exchanges did not significantly alter insulin stimulated HIR autophosphorylation. The data suggest that the serine residues at position 994 as well as 1023/25 might be part of inhibitory domains of the insulin receptor.
Strack
V
V
Eberhard-Karls-Universität Tübingen, Abt. IV, Germany.
Stoyanov
B
B
Bossenmaier
B
B
Mosthaf
L
L
Kellerer
M
M
Häring
H U
HU
D019943:Amino Acid Substitution; D015153:Blotting, Western; D002384:Catalysis; D002478:Cells, Cultured; D006801:Humans; D007328:Insulin / Q000378:metabolism; D008958:Models, Molecular; D010766:Phosphorylation; D017354:Point Mutation; D011972:Receptor, Insulin / Q000235:genetics / Q000378:metabolism*; D012694:Serine / Q000235:genetics / Q000378:metabolism*
D007328:Insulin; D012694:Serine; D011972:Receptor, Insulin