scholarly journals Insulin receptor substrate 1 rescues insulin action in CHO cells expressing mutant insulin receptors that lack a juxtamembrane NPXY motif.

1995 ◽  
Vol 15 (9) ◽  
pp. 4711-4717 ◽  
Author(s):  
D Chen ◽  
D J Van Horn ◽  
M F White ◽  
J M Backer
Keyword(s):  
Insulin Receptor ◽  
Tyrosine Phosphorylation ◽  
Cho Cells ◽  
Glycogen Synthesis ◽  
Insulin Receptors ◽  
Insulin Receptor Substrate ◽  
Insulin Stimulation ◽  
Insulin Receptor Substrate 1 ◽  
Npxy Motif ◽  
Stimulation Of

Insulin signals are mediated through tyrosine phosphorylation of specific proteins such as insulin receptor substrate 1 (IRS-1) and Shc by the activated insulin receptor (IR). Phosphorylation of both proteins is nearly abolished by an alanine substitution at Tyr-960 (A960) in the beta-subunit of the receptor. However, overexpression of IRS-1 in CHO cells expressing the mutant receptor (A960 cells) restored sufficient tyrosine phosphorylation of IRS-1 to rescue IRS-1/Grb-2 binding and phosphatidylinositol 3' kinase activation during insulin stimulation. Shc tyrosine phosphorylation and its binding to Grb-2 were impaired in the A960 cells and were unaffected by overexpression of IRS-1. Although overexpression of IRS-1 increased IRS-1 binding to Grb-2, ERK-1/ERK-2 activation was not rescued. These data suggest that signaling molecules other than IRS-1, perhaps including Shc, are critical for insulin stimulation of p21ras. Interestingly, overexpression of IRS-1 in the A960 cells restored insulin-stimulated mitogenesis and partially restored insulin stimulation of glycogen synthesis. Thus, IRS-1 tyrosine phosphorylation is sufficient to increase the mitogenic response to insulin, whereas insulin stimulation of glycogen synthesis appears to involve other factors. Moreover, IRS-1 phosphorylation is either not sufficient or not involved in insulin stimulation of ERK.

The role of the NPXY motif in the insulin receptor in tyrosine phosphorylation of insulin receptor substrate-1 and Shc.

Endocrinology ◽  
1995 ◽  
Vol 136 (8) ◽  
pp. 3437-3443 ◽  
Author(s):  
Y Kaburagi ◽  
R Yamamoto-Honda ◽  
K Tobe ◽  
K Ueki ◽  
M Yachi ◽  
...  
Keyword(s):  
Insulin Receptor ◽  
Tyrosine Phosphorylation ◽  
Insulin Receptor Substrate ◽  
Insulin Receptor Substrate 1 ◽  
Npxy Motif

scholarly journals The insulin receptor substrate-1-related 4PS substrate but not the interleukin-2R gamma chain is involved in interleukin-13-mediated signal transduction

Blood ◽  
1995 ◽  
Vol 86 (11) ◽  
pp. 4218-4227 ◽  
Author(s):  
LM Wang ◽  
P Michieli ◽  
WR Lie ◽  
F Liu ◽  
CC Lee ◽  
...  
Keyword(s):  
Insulin Receptor ◽  
Tyrosine Phosphorylation ◽  
Insulin Receptor Substrate ◽  
Mitogenic Response ◽  
Gamma Chain ◽  
Insulin Receptor Substrate 1 ◽  
Gamma Subunit ◽  
Interleukin 13 ◽  
Sh2 Domains ◽  
Stimulation Of

Interleukin-13 (IL-13) induced a potent mitogenic response in IL-3- dependent TF-1 cells and DNA synthesis to a lesser extent in MO7E and FDC-P1 cells. IL-13 stimulation of these lines, like IL-4 and insulin- like growth factor-1 (IGF-1), resulted in tyrosine phosphorylation of a 170-kD substrate. The tyrosine-phosphorylated 170-kD substrate strongly associated with the 85-kD subunit of phosphoinositol-3 (PI-3) kinase and with Grb-2. Anti-4PS serum readily detected the 170-kD substrate in lysates from both TF-1 and FDC-P1 cells stimulated with IL-13 or IL-4. These data provide evidence that IL-13 induces tyrosine phosphorylation of the 4PS substrate, providing an essential interface between the IL- 13 receptor and signaling molecules containing SH2 domains. IL-13 and IL-4 stimulation of murine L cell fibroblasts, which endogenously express the IL-4 receptor (IL-4R alpha) and lack expression of the IL-2 receptor gamma subunit (IL-2R gamma), resulted in tyrosine phosphorylation of insulin receptor substrate-1 (IRS-1)/4PS. Enhanced tyrosine phosphorylation of IRS-1/4PS was observed in response to IL-4, but not IL-13 treatment of L cells transfected with the IL-2R gamma chain. These results indicate that IL-13 does not use the IL-2R gamma subunit in its receptor complex and that expression of IL-2R gamma enhances, but is not absolutely required for mediating IL-4-induced tyrosine phosphorylation of IRS-1/4PS.

scholarly journals Mitogenic and Metabolic Effects of Type I IGF Receptor Overexpression in Insulin Receptor-Deficient Hepatocytes

Endocrinology ◽  
2001 ◽  
Vol 142 (8) ◽  
pp. 3354-3360 ◽  
Author(s):  
Jane J. Kim ◽  
Byung-Chul Park ◽  
Yoshiaki Kido ◽  
Domenico Accili
Keyword(s):  
Cell Proliferation ◽  
Insulin Receptor ◽  
Glycogen Synthesis ◽  
Insulin Receptors ◽  
Metabolic Effects ◽  
Insulin Receptor Substrate ◽  
Type I ◽  
Wild Type ◽  
Igf I ◽  
Stimulation Of

Abstract We have previously shown that hepatocytes lacking insulin receptors (Ir−/−) fail to mediate metabolic responses, such as stimulation of glycogen synthesis, while retaining the ability to proliferate in response to IGFs. In this study we have asked whether overexpression of type I IGF receptors would rescue the metabolic response of Ir−/− hepatocytes. After IGF-I stimulation, insulin receptor substrate-1 and -2 phosphorylation and PI3K activity were restored to levels similar to or greater than those seen in wild-type cells. Rates of cell proliferation in response to IGF-I increased approximately 2-fold, whereas glycogen synthesis was restored to wild-type levels, but was comparatively smaller than that elicited by overexpression of insulin receptors. In summary, overexpression of IGF-I receptors in Ir−/− hepatocytes normalized insulin receptor substrate-2 phosphorylation and glycogen synthesis to wild-type levels, whereas it increased cell proliferation above wild-type levels. Moreover, stimulation of glycogen synthesis was submaximal compared with the effect of insulin receptor overexpression. We conclude that IGF-I receptors are more efficiently coupled to cell proliferation than insulin receptors, but are less potent than insulin receptors in stimulating glycogen synthesis. The data are consistent with the possibility that there exist intrinsic signaling differences between insulin and IGF-I receptors.

scholarly journals The Pleckstrin Homology and Phosphotyrosine Binding Domains of Insulin Receptor Substrate 1 Mediate Inhibition of Apoptosis by Insulin

1998 ◽  
Vol 18 (11) ◽  
pp. 6784-6794 ◽  
Author(s):  
Lynne Yenush ◽  
Christine Zanella ◽  
Tohru Uchida ◽  
Dolores Bernal ◽  
Morris F. White
Keyword(s):  
Insulin Receptor ◽  
Tyrosine Phosphorylation ◽  
Insulin Receptors ◽  
Insulin Receptor Substrate ◽  
Phosphorylation Sites ◽  
Pleckstrin Homology ◽  
Insulin Stimulation ◽  
Kinase Cascade ◽  
Irs Proteins ◽  
Ptb Domains

ABSTRACT Insulin and insulin-like growth factor 1 (IGF-1) evoke diverse biological effects through receptor-mediated tyrosine phosphorylation of insulin receptor substrate (IRS) proteins. We investigated the elements of IRS-1 signaling that inhibit apoptosis of interleukin 3 (IL-3)-deprived 32D myeloid progenitor cells. 32D cells have few insulin receptors and no IRS proteins; therefore, insulin failed to inhibit apoptosis during IL-3 withdrawal. Insulin stimulated mitogen-activated protein kinase in 32D cells expressing insulin receptors (32DIR) but failed to activate the phosphatidylinositol 3 (PI 3)-kinase cascade or to inhibit apoptosis. By contrast, insulin stimulated the PI 3-kinase cascade, inhibited apoptosis, and promoted replication of 32DIR cells expressing IRS-1. As expected, insulin did not stimulate PI 3-kinase in 32DIR cells, which expressed a truncated IRS-1 protein lacking the tail of tyrosine phosphorylation sites. However, this truncated IRS-1 protein, which retained the NH2-terminal pleckstrin homology (PH) and phosphotyrosine binding (PTB) domains, mediated phosphorylation of PKB/akt, inhibition of apoptosis, and replication of 32DIR cells during insulin stimulation. These results suggest that a phosphotyrosine-independent mechanism mediated by the PH and PTB domains promoted antiapoptotic and growth actions of insulin. Although PI 3-kinase was not activated, its phospholipid products were required, since LY294002 inhibited these responses. Without IRS-1, a chimeric insulin receptor containing a tail of tyrosine phosphorylation sites derived from IRS-1 activated the PI 3-kinase cascade but failed to inhibit apoptosis. Thus, phosphotyrosine-independent IRS-1-linked pathways may be critical for survival and growth of IL-3-deprived 32D cells during insulin stimulation.

Early alteration of insulin stimulation of PI 3-kinase in muscle and adipocyte from gold thioglucose obese mice

1995 ◽  
Vol 268 (4) ◽  
pp. E604-E612 ◽  
Author(s):  
S. J. Heydrick ◽  
N. Gautier ◽  
C. Olichon-Berthe ◽  
E. Van Obberghen ◽  
Y. Le Marchand-Brustel
Keyword(s):  
Insulin Receptor ◽  
Tyrosine Phosphorylation ◽  
Glycogen Synthesis ◽  
Lipid Synthesis ◽  
Obese Mice ◽  
Insulin Stimulation ◽  
Gold Thioglucose ◽  
Stimulation Of

The activation of phosphatidylinositol 3-kinase (PIK) was studied in vivo and in vitro in soleus muscle and adipocytes from young (8 wk) and old (30 wk) gold thioglucose obese mice. Insulin resistance assessed from muscle glucose transport and glycogen synthesis was present both in young and old obese mice. Adipocyte lipid synthesis and muscle glycolysis or glucose oxidation are not defective in young obese mice but become resistant later on. After incubation with 50 nM insulin, muscle antiphosphotyrosine-immunoprecipitable PIK activity was stimulated 5- to 10-fold in both young and old animals. This response was impaired by 56 and 75% in muscles from young and old obese mice, respectively. Insulin stimulation of receptor tyrosine kinase activity was only slightly decreased in muscle of young obese mice, whereas insulin receptor substrate 1 (IRS-1) tyrosine phosphorylation was blunted. The altered PIK stimulation in muscle, which is present both in vivo and in vitro, is thus characterized by a reduced association of PIK activity with IRS-1 and appears to result from a diminished IRS-1 tyrosine phosphorylation. In adipocytes isolated from lean mice, antiphosphotyrosine-immunoprecipitable PIK increased 25-fold within 10 min of incubation with insulin. This stimulation was markedly altered both in young and old obese mice, whereas lipogenesis was insulin resistant only in old obese animals. In adipocytes from young obese mice, insulin's stimulatory effect on the phosphorylation of insulin receptor beta-subunit, pp60, and an exogenous substrate was normal, whereas IRS-1 tyrosine phosphorylation was markedly depressed.(ABSTRACT TRUNCATED AT 250 WORDS)

Role of IRS-1-GRB-2 complexes in insulin signaling

1994 ◽  
Vol 14 (6) ◽  
pp. 3577-3587
Author(s):  
M G Myers ◽  
L M Wang ◽  
X J Sun ◽  
Y Zhang ◽  
L Yenush ◽  
...  
Keyword(s):  
Map Kinase ◽  
Insulin Receptor ◽  
Insulin Receptors ◽  
Growth Factor Signaling ◽  
Insulin Stimulation ◽  
Mitogen Activated Protein ◽  
D Cells ◽  
Stimulation Of

GRB-2 is a small SH2- and SH3 domain-containing adapter protein that associates with the mammalian SOS homolog to regulate p21ras during growth factor signaling. During insulin stimulation, GRB-2 binds to the phosphorylated Y895VNI motif of IRS-1. Substitution of Tyr-895 with phenylalanine (IRS-1F-895) prevented the IRS-1-GRB-2 association in vivo and in vitro. The myeloid progenitor cell line, 32-D, is insensitive to insulin because it contains few insulin receptors and no IRS-1. Coexpression of IRS-1 or IRS-1F-895 with the insulin receptor was required for insulin-stimulated mitogenesis in 32-D cells, while expression of the insulin receptor alone was sufficient to mediate insulin-stimulated tyrosine phosphorylation of Shc and activation of p21ras and mitogen-activated protein (MAP) kinase. The Shc-GRB-2 complex formed during insulin stimulation is a possible mediator of p21ras and MAP kinase activation in IRS-1-deficient 32-D cells. Interestingly, IRS-1, but not IRS-1F-895, enhanced the stimulation of MAP kinase by insulin in 32-D cells expressing insulin receptors. Thus, IRS-1 contributes to the stimulation of MAP kinase by insulin, probably through formation of the IRS-1-GRB-2 complex at Tyr-895. Our results suggest that the Shc-GRB-2 complex and the activation of p21ras-dependent signaling pathways, including MAP kinase, are insufficient for insulin-stimulated mitogenesis and that the essential function(s) of IRS-1 in proliferative signaling is largely unrelated to IRS-1-GRB-2 complex formation.

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