TY - JOUR
T1 - Insulin and interleukin-4 induce desensitization to the mitogenic effects of insulin-like growth factor-i
T2 - Pivotal role for insulin receptor substrate-2
AU - Haddad, Tufia C.
AU - Conover, Cheryl A.
N1 - Copyright:
Copyright 2007 Elsevier B.V., All rights reserved.
PY - 1997/8/1
Y1 - 1997/8/1
N2 - Insulin-induced desensitization to insulin-like growth factor-I (IGF-I) stimulated mitogenesis in bovine fibroblasts involves steps distal to IGF-I binding to its tyrosine kinase receptor. When quiescent cultures of bovine fibroblasts were stimulated with 10 nM IGF-I and total cell lysates immunoblotted with anti-phosphotyrosine antibody, we observed a band at ~97 kDa, representing the β-subunit of the IGF-I receptor, and a predominant tyrosyl-phosphorylated species migrating as a broad band between 170 and 190 kDa. The majority of proteins in this latter band were immunoprecipitated by antibodies against insulin receptor substrate (IRS)-2 and not by antibodies against IRS-1. Pretreatment of bovine fibroblasts with 10 nM insulin for 48 h blocked subsequent IGF-I-stimulated DNA synthesis and the IGF-I-induced increase in tyrosyl-phosphorylated IRS-2. Insulin pretreatment did not alter IRS-1 or IRS-2 expression by these cells, as assessed by metabolic labeling and direct immunoblotting with IRS antibodies. The interleukin-4 (IL-4) cytokine receptor also has IRS-2 as its major substrate for tyrosine phosphorylation. Although 10 nM IL-4 was as effective as 10 nM IGF-I in stimulating IRS-2 phosphorylation, 10 nM IL-4 did not have comparable mitogenic power in these cells. Nonetheless, pretreatment of bovine fibroblasts with IL-4 inhibited IGF-I-stimulated DNA synthesis by 50-60%, concomitant with a decrease in IGF-I-induced IRS-2 phosphorylation. Insulin- induced desensitization could be prevented if a specific inhibitor of phosphatidylinositol 3-kinase (LY294002), but not an inhibitor of mitogenactivated protein kinase (PD98059), was present during the preincubation period. LY294002 also prevented the shift in IRS-2 molecular mass in response to prolonged incubation of cells with insulin. These data indicate that, in a nontransformed cell system, IRS-2 plays a key role in cellular desensitization to IGF-I-stimulated mitogenesis most likely through a feedback mechanism in the phosphatidylinositol 3-kinase pathway. Furthermore, they suggest that signaling through IRS-2 may provide an important point of integration for hormone, growth factor, and cytokine receptor systems that regulate critical cellular growth responses.
AB - Insulin-induced desensitization to insulin-like growth factor-I (IGF-I) stimulated mitogenesis in bovine fibroblasts involves steps distal to IGF-I binding to its tyrosine kinase receptor. When quiescent cultures of bovine fibroblasts were stimulated with 10 nM IGF-I and total cell lysates immunoblotted with anti-phosphotyrosine antibody, we observed a band at ~97 kDa, representing the β-subunit of the IGF-I receptor, and a predominant tyrosyl-phosphorylated species migrating as a broad band between 170 and 190 kDa. The majority of proteins in this latter band were immunoprecipitated by antibodies against insulin receptor substrate (IRS)-2 and not by antibodies against IRS-1. Pretreatment of bovine fibroblasts with 10 nM insulin for 48 h blocked subsequent IGF-I-stimulated DNA synthesis and the IGF-I-induced increase in tyrosyl-phosphorylated IRS-2. Insulin pretreatment did not alter IRS-1 or IRS-2 expression by these cells, as assessed by metabolic labeling and direct immunoblotting with IRS antibodies. The interleukin-4 (IL-4) cytokine receptor also has IRS-2 as its major substrate for tyrosine phosphorylation. Although 10 nM IL-4 was as effective as 10 nM IGF-I in stimulating IRS-2 phosphorylation, 10 nM IL-4 did not have comparable mitogenic power in these cells. Nonetheless, pretreatment of bovine fibroblasts with IL-4 inhibited IGF-I-stimulated DNA synthesis by 50-60%, concomitant with a decrease in IGF-I-induced IRS-2 phosphorylation. Insulin- induced desensitization could be prevented if a specific inhibitor of phosphatidylinositol 3-kinase (LY294002), but not an inhibitor of mitogenactivated protein kinase (PD98059), was present during the preincubation period. LY294002 also prevented the shift in IRS-2 molecular mass in response to prolonged incubation of cells with insulin. These data indicate that, in a nontransformed cell system, IRS-2 plays a key role in cellular desensitization to IGF-I-stimulated mitogenesis most likely through a feedback mechanism in the phosphatidylinositol 3-kinase pathway. Furthermore, they suggest that signaling through IRS-2 may provide an important point of integration for hormone, growth factor, and cytokine receptor systems that regulate critical cellular growth responses.
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U2 - 10.1074/jbc.272.31.19525
DO - 10.1074/jbc.272.31.19525
M3 - Article
C2 - 9235956
AN - SCOPUS:0030743040
SN - 0021-9258
VL - 272
SP - 19525
EP - 19531
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
IS - 31
ER -