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

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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

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Nelfinavir
Adipocytes
Insulin Resistance
Phosphatidylinositol 3-Kinase
Insulin
Green Fluorescent Proteins
Membrane Fusion
Protein Transport
Membranes
Phosphorylation
HIV Protease Inhibitors
Glycogen Synthase Kinase 3
Insulin Receptor Substrate Proteins
Facilitative Glucose Transport Proteins
Insulin Receptor
Phosphatidylinositols
Tyrosine
phosphatidylinositol 3,4,5-triphosphate
Catalytic Domain
Phosphotransferases

ASJC Scopus subject areas

  • Endocrinology

Cite this

Postreceptoral adipocyte insulin resistance induced by nelfinavir is caused by insensitivity of PKB/Akt to phosphatidylinositol-3,4,5-trisphosphate. / Kachko, Ilana; Maissel, Adva; Mazor, Livnat; Ben-Romano, Ronit; Watson, Robert T.; Hou, June C.; Pessin, Jeffrey E.; Bashan, Nava; Rudich, Assaf.

In: Endocrinology, Vol. 150, No. 6, 06.2009, p. 2618-2626.

Research output: Contribution to journalArticle

Kachko, Ilana ; Maissel, Adva ; Mazor, Livnat ; Ben-Romano, Ronit ; Watson, Robert T. ; Hou, June C. ; Pessin, Jeffrey E. ; Bashan, Nava ; Rudich, Assaf. / Postreceptoral adipocyte insulin resistance induced by nelfinavir is caused by insensitivity of PKB/Akt to phosphatidylinositol-3,4,5-trisphosphate. In: Endocrinology. 2009 ; Vol. 150, No. 6. pp. 2618-2626.
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AU - Ben-Romano, Ronit

AU - Watson, Robert T.

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AU - Pessin, Jeffrey E.

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