Phorbol esters inhibit insulin-induced receptor down-regulation in cultured human lymphocytes: association with diminished insulin receptor autophosphorylation.
Exposure of cells to phorbol 12-myristate 13-acetate (PMA) has been reported to result in resistance to the acute biological effects of insulin and an associated reduction in insulin-receptor tyrosine kinase activity. To investigate the relationship of insulin receptor autophosphorylation with a longer-term action of insulin the effect of PMA on insulin-stimulated receptor down-regulation was examined in cultured human lymphocytes (IM-9). Lymphocytes bound [3H]phorbol dibutyrate specifically with characteristics typical of binding to protein kinase C (PKC). Acute exposure (30 min) to PMA resulted in a transient decrease of insulin binding which is consistent with a decrease in receptor number. Chronic (18 h) exposure to PMA (5 nM) resulted in inhibition of insulin-induced down-regulation of its cognate receptor. Sphingosine, an inhibitor of PKC, or chronic pre-exposure to a high concentration of PMA (1 microM), which is known to inactivate PKC, blocked the effect of PMA. PMA inhibited insulin-stimulated receptor internalization by 26% and receptor degradation by 82%. Exposure of intact cells to PMA followed by insulin treatment inhibited insulin-receptor autophosphorylation subsequently assayed in vitro, as well as beta-subunit tyrosine phosphorylation in situ. In summary, PMA inhibited insulin-stimulated receptor down-regulation via activation of PKC. This was associated with an inhibition of both receptor internalization and receptor degradation. There was a concomitant inhibition of receptor tyrosine autophosphorylation consistent with a requirement of receptor kinase activation for both short-term and long-term biological effects of insulin.
Department of Medicine, Royal Victoria Hospital and McGill University, Montreal, Quebec, Canada.
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D002460:Cell Line; D015536:Down-Regulation / Q000187:drug effects*; D004789:Enzyme Activation / Q000187:drug effects; D006801:Humans; D007328:Insulin / Q000494:pharmacology*; D008214:Lymphocytes / Q000187:drug effects / Q000378:metabolism*; D015240:Phorbol 12,13-Dibutyrate / Q000378:metabolism; D010766:Phosphorylation; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D011972:Receptor, Insulin / Q000187:drug effects / Q000378:metabolism*; D013110:Sphingosine / Q000494:pharmacology; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*
D007328:Insulin; D015240:Phorbol 12,13-Dibutyrate; D011505:Protein-Tyrosine Kinases; D011972:Receptor, Insulin; D011493:Protein Kinase C; D013110:Sphingosine; D013755:Tetradecanoylphorbol Acetate