Wortmannin inhibits the effects of insulin and serum on the activities of glycogen synthase kinase-3 and mitogen-activated protein kinase

Abstract

We have previously shown that insulin causes inactivation of glycogen synthase kinase-3 (GSK-3) in Chinese hamster ovary cells over-expressing the human insulin receptor (CHO.T cells). We now show that serum and phorbol ester also cause rapid inactivation of GSK-3, both in CHO.T cells and in the nontransfected parental cell line, CHO.K1 cells. Rapamycin was without effect on the inactivation of GSK-3 by insulin, serum or phorbol ester, indicating that the p70 S6 kinase pathway is not involved. In contrast, wortmannin, a potent inhibitor of phosphatidylinositol 3-kinase, blocked the effects of both insulin and serum on GSK-3 activity, and also substantially reduced the activation of both p90 S6 kinase (by insulin) and mitogen-activated protein (MAP) kinase (by insulin and serum). These findings imply (i) that GSK-3 activity is regulated by a cascade involving MAP kinase and p90 S6 kinase and (ii) that wortmannin affects an early step in the MAP kinase pathway. One can infer from this that GSK-3 may be an important regulatory enzyme for the control of several biosynthetic pathways, key enzymes in which are regulated by GSK-3-mediated phosphorylation. Wortmannin had a smaller effect on the activation of MAP kinase by phorbol ester, indicating that phorbol esters may stimulate MAP kinase by a different or additional mechanism to that employed by insulin or serum. Wortmannin had very little effect on the inactivation of GSK-3 by phorbol ester: possible reasons for this are discussed.

ErbB kinases and NDF signaling in human prostate cancer cells

“…Representative ®elds are shown. Magni®cation=2006 p70 S6K activity may not always be necessary for the physiological e ects of NDF, since several recent reports have shown p70 S6K to be dispensable for PI3-K-mediated suppression of apoptosis (Dudek et al, 1997;Kau mann-Zeh et al, 1997;Kulik et al, 1997;Yao and Cooper, 1996), glycogen synthesis (Welsh et al, 1994;Yamamoto-Honda et al, 1995), and gene expression (Sutherland et al, 1995). Both the STAT and PLCg pathways can be activated by TGF-a treatment in prostate lines.…”

Defects in insulin signal transduction in liver and muscle of pregnant rats

“…Distinct experimental approaches have led to the conclusion that PI 3-kinase is necessary for insulin-stimulated GLUT4 translocation [38][39][40][41][42][43][44]. Evidence from other sources has also demonstrated a correlation between PI 3-kinase activity and glycogen metabolism [45,46]. Thus, it is reasonable to speculate that the IRS-1/PI 3-kinase pathway may be linked to the activation of glucose transport in muscle and glycogen synthesis in liver and muscle, and that a reduction in this association in pregnancy may have a role in the insulin resistance in liver and muscle of pregnant rats.…”

The Ras/Rac1/Cdc42/SEK/JNK/c-Jun Cascade Is a Key Pathway by Which Agonists Stimulate DNA Synthesis in Primary Cultures of Rat Hepatocytes

“…After 20 min, 40 l of the reaction mixtures were spotted onto a 2-cm circle of P81 paper (Whatman) and immediately placed into 180 mM phosphoric acid. Papers were washed four times (10 min each) with phosphoric acid and once with acetone, and 32 P-labeled incorporation into peptide substrate compared with identical samples containing enzyme but without substrate determined by liquid scintillation spectroscopy (Welsh et al, 1994).…”