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. 2008 Jan 17;149(5):2451–2458. doi: 10.1210/en.2007-1595

Figure 8.

Figure 8

Overexpression of PKC-ζ impairs insulin-stimulated activation of PI3K associated with IRS-4. NIH-3T3IR cells were transiently cotransfected with HA-tagged IRS-1 or myc-tagged IRS-4 and either an empty control vector or PKC-ζ. Transfected cells were treated with 100 nm insulin for 0, 2, or 60 min. A, Experiments were as described in Fig. 6A. B, Cell lysates were immunoprecipitated (IP) with anti-myc antibody and subsequently immunoblotted (IB) with either anti-p85 antibody (top panel) or anti-IRS-4 antibody (bottom panel). IRS-4-associated PI3K activity was determined in parallel using a lipid kinase assay measuring [32P]-phosphatidylinositol phosphate (PIP) product (middle panel). Results from five independent lipid kinase experiments were quantified by PhosphorImager and expressed as mean ± sem. Overexpression of PKC-ζ significantly reduced insulin-stimulated PI3K activity associated with both IRS-1 and IRS-4 at 2 and 60 min when compared with control cells (*, P = 0.04; §, P = 0.02; ¶, P = 0.03; #, P = 0.04).