This Article Full Text Full Text (PDF) Supplemental Data Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Request Copyright Permission Google Scholar Articles by Goenaga, D. Articles by Burnol, A.-F. PubMed PubMed Citation Articles by Goenaga, D. Articles by Burnol, A.-F. Pubmed/NCBI databases Gene GEO Profiles HomoloGene Protein UniGene Compound via MeSH Substance via MeSH

Molecular Determinants of Grb14-Mediated Inhibition of Insulin Signaling

Institut Cochin (D.G., C.H., N.C., D.P., J.G., T.I., A.-F.B.), Université Paris Descartes, Centre National de la Recherche Scientifique (CNRS) Unité Mixte de Recherche (UMR) 8104; Institut National de la Santé et de la Recherche Médicale (D.G., C.H., N.C., D.P., J.G., T.I., A.-F.B.), Unité 567, 75014 Paris, France; Laboratoire de Régulation des Signaux de Division (K.C., E.B.-P.), Unité d’Enseignement 4020, Institut Fédératif de Recherche 118, 59655 Villeneuve d’Ascq Cedex, France; Cancer Research Program (L.J.H., R.J.D.), Garvan Institute of Medical Research, Darlinghurst 2010, Sydney, New South Wales, Australia; and Laboratoire de Cristallographie et Résonance Magnétique Nucléaire Biologiques CNRS (I.B.), UMR 8105, Faculté de Pharmacie Université Paris Descartes, Paris, France

Address all correspondence and requests for reprints to: Anne-Françoise Burnol, Institut Cochin, Département Endocrinologie, Métabolisme et Cancer, Institut National de la Santé et de la Recherche Médicale Unité 567, Centre National de la Recherche Scientifique Unité Mixte de Recherche 8104, Université Paris Descartes, 24 rue du Faubourg Saint-Jacques, 75014 Paris, France. E-mail: anne-francoise.burnol{at}inserm.fr.

Grb14 belongs to the Grb7 family of molecular adapters and was identified as an inhibitor of insulin signaling. Grb14 binds to activated insulin receptors (IR) and inhibits their catalytic activity. To gain more insight into the Grb14 molecular mechanism of action, we generated various mutants and studied the Grb14-IR interaction using coimmunoprecipitation and bioluminescence resonance energy transfer (BRET) experiments. Biological activity was further analyzed using the Xenopus oocyte model and a functional complementation assay measuring cellular proliferation rate in Grb14 knockout mouse embryonic fibroblasts. These studies identified two important interaction sites, Grb14 L404-IR L1038 and Grb14 R385-IR K1168, involving the IR C-helix and activation loop, respectively. Interestingly, the former involves residues that are likely to be crucial for the specificity of IR binding with regard to other members of the Grb7 family. In addition, mutation of the Grb14-S370 residue suggested that its phosphorylation status controlled the biological activity of the protein. We further demonstrated that insulin-induced Grb14-PDK1 interaction is required in addition to Grb14-IR binding to mediate maximal inhibition of insulin signaling. This study provides important insights into the molecular determinants of Grb14 action by demonstrating that Grb14 regulates insulin action at two levels, through IR binding and by interfering with downstream pathways. Indeed, a precise knowledge of the molecular mechanism of insulin signaling inhibition by Grb14 is a prerequisite for the development of insulin-sensitizing molecules to treat pathophysiological states such as obesity or type 2 diabetes.