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.
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 (Deltap85). 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.
Farhang-Fallah
Janet
J
Department of Biology. Department of Pathology and Molecular Medicine, McMaster University, Hamilton, Ontario L8N 3Z5, Canada.
Randhawa
Varinder K
VK
Nimnual
Anjaruwee
A
Klip
Amira
A
Bar-Sagi
Dafna
D
Rozakis-Adcock
Maria
M
D016475:3T3 Cells; D000199:Actins / Q000378:metabolism; D000818:Animals; D001692:Biological Transport; D001798:Blood Proteins / Q000235:genetics / Q000378:metabolism*; D019556:COS Cells; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D002522:Chlorocebus aethiops; D003599:Cytoskeleton / Q000378:metabolism; D015870:Gene Expression; D017930:Genes, Reporter; D051275:Glucose Transporter Type 4; D007328:Insulin / Q000378:metabolism / Q000494:pharmacology; D055504:Insulin Receptor Substrate Proteins; D047908:Intracellular Signaling Peptides and Proteins; D008156:Luciferases / Q000235:genetics; D051379:Mice; D009004:Monosaccharide Transport Proteins / Q000378:metabolism*; D009124:Muscle Proteins; D010750:Phosphoproteins / Q000235:genetics / Q000378:metabolism*; D016760:Proto-Oncogene Proteins c-fos / Q000235:genetics; D011972:Receptor, Insulin / Q000378:metabolism*; D027602:Serum Response Element; D015398:Signal Transduction; D014158:Transcription, Genetic; D015533:Transcriptional Activation
D000199:Actins; D001798:Blood Proteins; D002352:Carrier Proteins; D051275:Glucose Transporter Type 4; D007328:Insulin; D055504:Insulin Receptor Substrate Proteins; D047908:Intracellular Signaling Peptides and Proteins; C526295:Irs1 protein, mouse; D009004:Monosaccharide Transport Proteins; D009124:Muscle Proteins; C418612:PHIP protein, human; D010750:Phosphoproteins; D016760:Proto-Oncogene Proteins c-fos; C495693:Slc2a4 protein, mouse; C039077:platelet protein P47; D008156:Luciferases; D011972:Receptor, Insulin