scholarly journals The Pleckstrin Homology (PH) Domain-Interacting Protein Couples the Insulin Receptor Substrate 1 PH Domain to Insulin Signaling Pathways Leading to Mitogenesis and GLUT4 Translocation

2002 ◽  
Vol 22 (20) ◽  
pp. 7325-7336 ◽  
Author(s):  
Janet Farhang-Fallah ◽  
Varinder K. Randhawa ◽  
Anjaruwee Nimnual ◽  
Amira Klip ◽  
Dafna Bar-Sagi ◽  
...  
Keyword(s):  
Signal Transduction ◽  
Insulin Receptor ◽  
Dominant Negative ◽  
Dominant Negative Mutant ◽  
Insulin Receptor Substrate ◽  
Glut4 Translocation ◽  
Ph Domain ◽  
Insulin Receptor Substrate 1 ◽  
Interacting Protein ◽  
Negative Mutant

ABSTRACT Receptor-mediated tyrosine phosphorylation of the insulin receptor substrate 1 (IRS-1) is required for the propagation of many of insulin's biological effects. The amino-terminal pleckstrin homology (PH) domain of IRS-1 plays a pivotal role in promoting insulin receptor (IR)-IRS-1 protein interactions. We have recently reported the isolation of a PH domain-interacting protein, PHIP, which selectively binds to the IRS-1 PH domain and is stably associated with IRS-1 in mammalian cells. Here we demonstrate that overexpression of PHIP in fibroblasts enhances insulin-induced transcriptional responses in a mitogen-activated protein kinase-dependent manner. In contrast, a dominant-negative mutant of PHIP (DN-PHIP) was shown to specifically block transcriptional and mitogenic signals elicited by insulin and not serum. In order to examine whether PHIP/IRS-1 complexes participate in the signal transduction pathway linking the IR to GLUT4 traffic in muscle cells, L6 myoblasts stably expressing a myc-tagged GLUT4 construct (L6GLUT4myc) were transfected with either wild-type or dominant-interfering forms of PHIP. Whereas insulin-dependent GLUT4myc membrane translocation was not affected by overexpression of PHIP, DN-PHIP caused a nearly complete inhibition of GLUT4 translocation, in a manner identical to that observed with a dominant-negative mutant of the p85 subunit of phosphatidylinositol 3-kinase (Δp85). Furthermore, DN-PHIP markedly inhibited insulin-stimulated actin cytoskeletal reorganization, a process required for the productive incorporation of GLUT4 vesicles at the cell surface in L6 cells. Our results are consistent with the hypothesis that PHIP represents a physiological protein ligand of the IRS-1 PH domain, which plays an important role in insulin receptor-mediated mitogenic and metabolic signal transduction.

scholarly journals Role of Insulin Receptor Substrate-1 and pp60 in the Regulation of Insulin-induced Glucose Transport and GLUT4 Translocation in Primary Adipocytes

1997 ◽  
Vol 272 (41) ◽  
pp. 25839-25844 ◽  
Author(s):  
Yasushi Kaburagi ◽  
Shinobu Satoh ◽  
Hiroyuki Tamemoto ◽  
Ritsuko Yamamoto-Honda ◽  
Kazuyuki Tobe ◽  
...  
Keyword(s):  
Insulin Receptor ◽  
Glucose Transport ◽  
Insulin Receptor Substrate ◽  
Glut4 Translocation ◽  
Insulin Receptor Substrate 1

Src homology 3 domain-dependent interaction of Nck-2 with insulin receptor substrate-1

2001 ◽  
Vol 354 (2) ◽  
pp. 315-322 ◽  
Author(s):  
Yizeng TU ◽  
Liang LIANG ◽  
Stuart J. FRANK ◽  
Chuanyue WU
Keyword(s):  
Complex Formation ◽  
Insulin Receptor ◽  
Tyrosine Phosphorylation ◽  
Molecular Basis ◽  
Insulin Receptor Substrate ◽  
Ph Domain ◽  
Insulin Receptor Substrate 1 ◽  
Sh3 Domains ◽  
Terminal Domain ◽  
Src Homology

Insulin receptor substrate-1 (IRS-1) is a multi-domain protein that mediates signal transduction from receptors for insulin and other growth factors to a variety of downstream molecules through both tyrosine-phosphorylation-dependent and -independent interactions. While the tyrosine-phosphorylation-dependent interactions mediated by IRS-1 have been well characterized, the molecular basis underlying the tyrosine-phosphorylation-independent IRS-1 interactions is largely unknown. We previously detected, in an in vitro binding assay, interactions of Nck-2 Src homology (SH) 3 domains with IRS-1. We show here that IRS-1 associates with Nck-2 in vivo. Additionally, we have investigated the molecular basis underlying the IRS-1–Nck-2 complex formation. We have found that (i) mutations at the highly conserved tryptophan within the Nck-2 SH3 domains markedly reduced the association with IRS-1, (ii) interactions mediated by multiple SH3 domains enhance the complex formation of Nck-2 with IRS-1, (iii) deletion of either the phosphotyrosine-binding/Shc and IRS-1 NPXY-binding (PTB/SAIN) domains or the Pre-C-terminal domain of IRS-1, but not the pleckstrin homology (PH) domain, reduced the Nck-2 binding, (iv) PTB/SAIN domains or the Pre-C-terminal domain alone is capable of interacting with Nck-2, and (v) the IRS-1–Nck-2 interaction occurs in the absence of other proteins and therefore is direct. These results establish that IRS-1 is a bona fide target of the Nck-2 SH3 domains and reveal that IRS-1 forms a complex with Nck-2 via direct interactions mediated by multiple domains from both binding partners.

scholarly journals Ethanol inhibits insulin receptor substrate-1 tyrosine phosphorylation and insulin-stimulated neuronal thread protein gene expression

1995 ◽  
Vol 310 (1) ◽  
pp. 125-132 ◽  
Author(s):  
Y Y Xu ◽  
K Bhavani ◽  
J R Wands ◽  
S M de la Monte
Keyword(s):  
Gene Expression ◽  
Signal Transduction ◽  
Insulin Receptor ◽  
Tyrosine Phosphorylation ◽  
Neuronal Differentiation ◽  
Growth Factor Receptor ◽  
Signal Transduction Pathway ◽  
Tumour Cell Line ◽  
Insulin Receptor Substrate ◽  
Insulin Receptor Substrate 1

Neuronal thread proteins (NTPs) are molecules that accumulate in the brains of patients with Alzheimer's disease, and may play a key role in both normal and neurodegenerative neuritic sprouting. In this investigation we determined whether NTP expression is up-regulated by insulin, an important neurotrophic factor that stimulates differentiation-associated neurite outgrowth, and studied the effects of ethanol, a known inhibitor of growth factor receptor tyrosine phosphorylation, on NTP expression and insulin-mediated signal transduction cascade in neuronal [primitive neuroectodermal tumour cell line 2; (PNET2)] cells. PNET2 cells were treated with 50 m-units/ml insulin in the presence or absence of 100 mM ethanol for 0.2-96 h, and cell proliferation and expression of NTP molecules were investigated by metabolic labelling, immunoprecipitation and immunohistochemical staining. Insulin stimulation resulted in an immediate increase in the levels of three (38, 18 and 15 kDa) of five NTP species (the others were of 26 and 21 kDa), followed by a decline in expression within 120 min; however, studies performed up to 96 h of culture demonstrated up-regulation by insulin of all five NTP species. Ethanol either abolished or severely muted the short- and long-term insulin-mediated upregulation of NTP expression, and substantially reduced insulin-mediated neuronal differentiation. The effects of ethanol on NTP gene expression were associated with impaired insulin-mediated tyrosine phosphorylation of both the insulin receptor beta subunit and the insulin receptor substrate-1 (IRS-1), resulting in decreased association of phosphatidylinositol 3-kinase with IRS-1. The findings suggest that ethanol may inhibit NTP expression associated with central nervous system neuronal differentiation by uncoupling the IRS-1-mediated insulin signal transduction pathway.

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