scholarly journals APS, an adapter protein with a PH and SH2 domain, is a substrate for the insulin receptor kinase

1999 ◽  
Vol 341 (3) ◽  
pp. 665-668 ◽  
Author(s):  
Zamal AHMED ◽  
Beverley J. SMITH ◽  
Kei KOTANI ◽  
Peter WILDEN ◽  
Tahir S. PILLAY
Keyword(s):  
Insulin Receptor ◽  
Sh2 Domain ◽  
Activation Loop ◽  
Adapter Proteins ◽  
Adapter Protein ◽  
Receptor Kinase ◽  
Kinase Activation ◽  
Tyrosine Residues ◽  
Insulin Receptor Kinase ◽  
Receptor Binding Protein

APS (adapter protein with a PH and SH2 domain) is the newest member of a family of tyrosine kinase adapter proteins including SH2-B and Lnk. We previously identified SH2-B as an insulin-receptor-binding protein and substrate [Kotani, Wilden and Pillay (1998) Biochem J. 335, 103-109]. Here we show that APS interacts with the insulin receptor kinase activation loop through its SH2 domain and insulin stimulates the tyrosine-phosphorylation of APS. Furthermore, the phosphorylation of activation-loop tyrosine residues 1158 and 1162 are required for this interaction.

scholarly journals SH2-Bα is an insulin-receptor adapter protein and substrate that interacts with the activation loop of the insulin-receptor kinase

1998 ◽  
Vol 335 (1) ◽  
pp. 103-109 ◽  
Author(s):  
Kei KOTANI ◽  
Peter WILDEN ◽  
Tahir S. PILLAY
Keyword(s):  
Growth Factor ◽  
Insulin Receptor ◽  
Receptor Binding ◽  
Binding Protein ◽  
Growth Factor Receptor ◽  
Sh2 Domain ◽  
Activation Loop ◽  
Adapter Protein ◽  
Insulin Signalling Pathway ◽  
Receptor Binding Protein

We identified SH2-Bα as an insulin-receptor-binding protein based on interaction screening in yeast hybrid systems and co-precipitation in cells. SH2-Bα contains pleckstrin-homology (‘PH’) and Src homology 2 (SH2) domains and is closely related to APS (adapter protein with a PH domain and an SH2 domain) and lnk, adapter proteins first identified in lymphocytes. SH2-Bα is ubiquitously expressed and is present in rat epididymal adipose tissue, liver and skeletal muscle, physiological sites of insulin action. On SDS/PAGE, SH2-Bα migrates at a molecular mass of 98 kDa, although the predicted size of SH2-Bα is 79.6 kDa. Insulin causes an electrophoretic mobility shift. SH2-Bα can be immunoprecipitated using anti-(insulin receptor) antibody from insulin-stimulated cells. Anti-phosphotyrosine antibody or the growth factor receptor-binding protein 2 (Grb2) SH2 domain precipitate SH2-Bα after insulin stimulation, suggesting that SH2-Bα is tyrosine-phosphorylated and may be a substrate for the insulin receptor. The SH2-Bα SH2 domain did not interact with insulin-receptor substrate (IRS) proteins or epidermal-growth-factor receptor. Mutation of the juxtamembrane and C-terminus of the insulin receptor did not abolish the interaction with the SH2 domain. This was further confirmed using a panel of activation-loop single point mutants where mutation of Tyr1158, Tyr1162 and Tyr1163 abolished interaction. Thus SH2-Bα is a likely component in the insulin-signalling pathway and may function as an alternative signalling protein by interacting with the activation loop of the insulin-receptor cytoplasmic domain.

scholarly journals APS, an adapter protein with a PH and SH2 domain, is a substrate for the insulin receptor kinase

1999 ◽  
Vol 341 (3) ◽  
pp. 665 ◽  
Author(s):  
Zamal AHMED ◽  
Beverley J. SMITH ◽  
Kei KOTANI ◽  
Peter WILDEN ◽  
Tahir S. PILLAY
Keyword(s):  
Insulin Receptor ◽  
Sh2 Domain ◽  
Adapter Protein ◽  
Receptor Kinase ◽  
Insulin Receptor Kinase

scholarly journals Hydrogen peroxide stimulates tyrosine phosphorylation of the insulin receptor and its tyrosine kinase activity in intact cells

1988 ◽  
Vol 250 (1) ◽  
pp. 95-101 ◽  
Author(s):  
O Koshio ◽  
Y Akanuma ◽  
M Kasuga
Keyword(s):  
Insulin Receptor ◽  
Kinase Activity ◽  
Intact Cell ◽  
Insulin Receptors ◽  
Beta Subunit ◽  
Receptor Kinase ◽  
Intact Cells ◽  
Tyrosine Residues ◽  
Insulin Receptor Kinase ◽  
Insulin Receptor Kinase Activity

H-35 rat hepatoma cells were labelled with [32P]orthophosphate and their insulin receptors isolated on wheat germ agglutinin (WGA)-agarose and anti-(insulin receptor) serum. The incubation of these cells with 10 mM-H2O2 for 10 min increased the phosphorylation of both the serine and tyrosine residues of the beta subunit of the insulin receptor. Next, insulin receptors were purified on WGA-agarose from control and H2O2-treated H-35 cells and the purified fractions incubated with [gamma-32P]ATP and Mn2+. Phosphorylation of the beta subunit of insulin receptors obtained from H2O2-treated cells was 150% of that of control cells. The kinase activity of the WGA-purified receptor preparation obtained from H2O2-treated cells, as measured by phosphorylation of src-related synthetic peptide, was increased about 4-fold over control cells. These data suggest that in intact cell systems, H2O2 may increase the insulin receptor kinase activity by inducing phosphorylation of the beta subunit of insulin receptor.

scholarly journals Insulin activation of insulin receptor kinase in erythrocytes is not altered in non-insulin-dependent diabetes and not influenced by hyperglycemia

2000 ◽  
Vol 166 (2) ◽  
pp. 275-281 ◽  
Author(s):  
HH Klein ◽  
R Muller ◽  
M Drenckhan ◽  
M Schutt ◽  
B Batge ◽  
...  
Keyword(s):  
Insulin Receptor ◽  
Insulin Receptors ◽  
Insulin Binding ◽  
Diabetic Control ◽  
Insulin Dependent Diabetes ◽  
Receptor Kinase ◽  
Kinase Activation ◽  
Insulin Dependent ◽  
Insulin Receptor Kinase

Recent studies suggest that high glucose concentrations impair insulin receptor phosphorylation and kinase activation in certain cell models. To examine whether such an effect of glucose can also be demonstrated in vivo, insulin receptor kinase activation was studied in erythrocytes from 11 patients with non-insulin-dependent diabetes (NIDDM), before and after reduction of hyperglycemia (from 14.6+/-1.6 to 6.6+/-0.5 mmol/l fasting plasma glucose within 8.6+/-0.6 days). For the measurement of receptor kinase activation, cells were incubated with insulin (0-400 nmol/l), solubilized and insulin receptors immobilized to microwells coated with anti-insulin receptor antibody. Kinase activity towards insulin receptor substrate-1 and insulin binding were then measured in these wells. Kinase activities (expressed as amol phosphate transferred per min and per fmol insulin binding activity) were similar before (2.4+/-0.4 and 32.2+/-2.0 amol/min per fmol with 0 and 400 nmol/l insulin, respectively) and after improvement of metabolic control (2.4+/-0.5 and 32.0+/-2.3 amol/min per fmol with 0 and 400 nmol/l insulin, respectively). Moreover, activities were also similar in 22 hyperglycemic patients with NIDDM (2.1+/-0.3 and 35.1+/-1.4 amol/min per fmol with 0 and 400 nmol/l insulin, respectively) compared with those in 21 non-diabetic control individuals (2.1+/-0.3 and 34.2+/-1.2 amol/min per fmol with 0 and 400 nmol/l insulin, respectively). We conclude that insulin activation of erythrocyte insulin receptor kinase is not impaired in NIDDM and is not influenced by hyperglycemia.

scholarly journals APS Facilitates c-Cbl Tyrosine Phosphorylation and GLUT4 Translocation in Response to Insulin in 3T3-L1 Adipocytes

2002 ◽  
Vol 22 (11) ◽  
pp. 3599-3609 ◽  
Author(s):  
Jun Liu ◽  
Akiko Kimura ◽  
Christian A. Baumann ◽  
Alan R. Saltiel
Keyword(s):  
Insulin Receptor ◽  
Tyrosine Phosphorylation ◽  
Insulin Treatment ◽  
Adapter Protein ◽  
Receptor Kinase ◽  
Wild Type ◽  
Insulin Stimulation ◽  
Intracellular Vesicles ◽  
Cbl Protein ◽  
Insulin Receptor Kinase

ABSTRACT APS is a Cbl-binding protein that is tyrosine phosphorylated by the insulin receptor kinase. Insulin-stimulated phosphorylation of tyrosine 618 in APS is necessary for its association with c-Cbl and the subsequent tyrosine phosphorylation of Cbl by the insulin receptor in both 3T3-L1 adipocytes and CHO-IR cells. When overexpressed in these cells, wild-type APS but not an APS/Y618F mutant facilitated the tyrosine phosphorylation of coexpressed Cbl and its association with Crk upon insulin stimulation. APS-facilitated phosphorylation occurred on tyrosines 371, 700, and 774 in the Cbl protein. APS also interacted directly with the c-Cbl-associated protein (CAP) and colocalized with the protein in cells. The association was dependent on the SH3 domains of CAP and was independent of insulin treatment. Overexpression of the APS/Y618F mutant in 3T3-L1 adipocytes blocked the insulin-stimulated tyrosine phosphorylation of endogenous Cbl and binding to Crk. Moreover, the translocation of GLUT4 from intracellular vesicles to the plasma membrane was also inhibited by overexpression of the APS/Y618F mutant. These data suggest that APS serves as an adapter protein linking the CAP/Cbl pathway to the insulin receptor and, further, that APS-facilitated Cbl tyrosine phosphorylation catalyzed by the insulin receptor is a crucial event in the stimulation of glucose transport by insulin.

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