Differential roles of insulin receptor substrates in the anti-apoptotic function of insulin-like growth factor-1 and insulin.
Insulin-like growth factor-1 (IGF-1) and insulin are known to prevent apoptosis. The signaling network of IGF-1 and insulin occurs via multiple pathways involving different insulin receptor substrates (IRSs). To define their roles in the anti-apoptotic function of IGF-1 and insulin, we established brown pre-adipocyte cell lines from wild-type and IRS knockout (KO) animals. In response to 16 h of serum deprivation, IRS-1-deficient cells showed a significant decrease in response to IGF-1 protection from apoptosis, whereas no changes were observed in the IRS-2, IRS-3, or IRS-4 KO cells. Five hours after serum withdrawal, cells already began to undergo apoptosis. At this early time point, IGF-1 and insulin were able to protect both wild-type and IRS-1 KO cells from death by 85-90%. After a longer period of serum deprivation, the protective ability of insulin and IGF-1 was decreased, and this was especially reduced in the IRS-1 KO cells. Reconstitution of these cells with IRS-1, IRS-2, IRS-3, or IRS-1/IRS-2 chimeras restored the anti-apoptotic effects of IGF-1, whereas overexpression of IRS-4 had no effect at long time points and actually reduced the effect of IGF-1 at the short time point. The biochemical basis of the defect in anti-apoptosis was not dependent on phosphorylation of mitogen-activated protein kinase; whereas phosphoinositide 3-kinase activity was decreased by 30% in IRS-1 KO cells. Akt phosphorylation was slightly reduced in these cells. Phosphorylation of the transcription factors cAMP response element-binding protein and FKHR by IGF-1 and insulin was markedly reduced in IRS-1 KO cells. In addition, both IGF-1 and insulin prevented caspase-3 cleavage in the wild-type cells, and this effect was greatly reduced in the IRS-1-deficient cells. These findings suggest that the IRS proteins may play differential roles in the anti-apoptotic effects of IGF-1 and insulin in brown pre-adipocytes, with IRS-1 being predominant, possibly acting through caspase-3-, CREB-, and FKHR-dependent mechanisms.
Tseng
Yu-Hua
YH
Research Division, Joslin Diabetes Center, Department of Medicine, Harvard Medical School, Boston, Massachusetts 02215, USA.
Ueki
Kohjiro
K
Kriauciunas
Kristina M
KM
Kahn
C Ronald
CR
D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D002460:Cell Line; D002478:Cells, Cultured; D016895:Culture Media, Serum-Free; D007328:Insulin / Q000494:pharmacology / Q000502:physiology*; D055504:Insulin Receptor Substrate Proteins; D007334:Insulin-Like Growth Factor I / Q000494:pharmacology*; D047908:Intracellular Signaling Peptides and Proteins; D007700:Kinetics; D051379:Mice; D018345:Mice, Knockout; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D010750:Phosphoproteins / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D011972:Receptor, Insulin / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D011994:Recombinant Proteins / Q000494:pharmacology; D014162:Transfection
D016895:Culture Media, Serum-Free; D007328:Insulin; D055504:Insulin Receptor Substrate Proteins; D047908:Intracellular Signaling Peptides and Proteins; C526295:Irs1 protein, mouse; C526298:Irs2 protein, mouse; C108872:Irs3 protein, mouse; C491531:Irs4 protein, mouse; D010750:Phosphoproteins; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D007334:Insulin-Like Growth Factor I; D011972:Receptor, Insulin; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases