Postreceptoral adipocyte insulin resistance induced by nelfinavir is caused by insensitivity of PKB/Akt to phosphatidylinositol-3,4,5-trisphosphate

Ilana Kachko, Adva Maissel, Livnat Mazor, Ronit Ben-Romano, Robert T. Watson, June C. Hou, Jeffrey E. Pessin, Nava Bashan, Assaf Rudich

Research output: Contribution to journalArticle

11 Scopus citations

Abstract

Adipocyte insulin resistance can be caused by proximal insulin signaling defects but also from postreceptor mechanisms, which in large are poorly characterized. Adipocytes exposed for 18 h to the HIV protease inhibitor nelfinavir manifest insulin resistance characterized by normal insulin-stimulated tyrosine phosphorylation of the insulin receptor and insulin receptor substrate proteins, preserved in vitro phosphatidylinositol 3-kinase (PI 3-kinase) assay activity but impaired activation of PKB/Akt and stimulation of glucose uptake. Here we aimed to assess whether impaired PKB/Akt activation is indeed rate limiting for insulin signaling propagation in response to nelfinavir and the mechanism for defective PKB/Akt activation. Nelfinavir treatment of 3T3-L1 adipocytes impaired the insulin-stimulated translocation and membrane fusion of myc-glucose transporter (GLUT)-4-green fluorescent protein (GFP) reporter. Phosphorylation of PKB/Akt substrates including glycogen synthase kinase-3 and AS160 decreased in response to nelfinavir, and this remained true, even in cells with forced generation of phosphatidylinositol-3,4, 5-trisphohphate (PIP3) by a membrane-targeted active PI 3-kinase, confirming that impaired PKB/Akt activation was rate limiting for insulin signal propagation. Cells expressing a GFP-tagged pleckstrin homology domain of general receptors for phosphoinositides 1, which binds PIP3, revealed intact PIP3-mediated plasma membrane translocation of this reporter in nelfinavir-treated cells. However, expression of a membrane-targeted catalytic subunit of PI 3-kinase failed to induce myc-GLUT4-GFP translocation in the absence of insulin, as it did in control cells. Conversely, a membrane-targeted and constitutively active PKB/Akt mutant was normally phosphorylated on S473 and T308, confirming intact PKB/Akt kinases activity, and induced myc-GLUT4-GFP translocation. Collectively, nelfinavir uncovers a postreceptor mechanism for insulin resistance, caused by interference with the sensing of PIP3 by PKB/Akt, leading to impaired GLUT4 translocation and membrane fusion.

Original languageEnglish (US)
Pages (from-to)2618-2626
Number of pages9
JournalEndocrinology
Volume150
Issue number6
DOIs
StatePublished - Jun 2009

ASJC Scopus subject areas

  • Endocrinology

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