Insulin-induced receptor loss in cultured human lymphocytes is due to accelerated receptor degradation.
We have measured the turnover rate of the polypeptide subunits of the insulin receptor in cultured human lymphocytes (IM-9 line) and have investigated the mechanism of insulin-induced receptor loss. To estimate the rate of receptor degradation, lymphocytes were either pulse-labeled with [35S]methionine or surface labeled with Na125I and lactoperoxidase. The insulin receptor was isolated by immunoprecipitation with anti-receptor antibody, and the rate of loss of radioactivity from each receptor subunit was determined after sodium dodecyl sulfate/polyacrylamide gel electrophoresis. Two major (Mr 135,000 and 95,000) and one minor (Mr 210,000) subunits were found. By both labeling methods, the half-lives of the major insulin receptor subunits were 9--12 hr in normal media. When the cells were cultured in media containing 1 microM insulin the turnover was accelerated 2.5- to 3.5-fold (half-life approximately 3 hr). The increase in degradation rate was dependent on the insulin concentration and correlated well with the ability to "down-regulate" the receptor. Guinea pig insulin was about 2% as active as porcine insulin in accelerating degradation, and human growth hormone was without effect. The acceleration of receptor degradation induced by insulin was partially blocked by 100 microM cycloheximide. The rate of biosynthesis of the insulin receptor did not appear to be altered in the presence of 1 microM insulin after correction for the change in degradation rate. In conclusion, these data demonstrate that insulin-induced receptor loss in cultured lymphocytes is due to accelerated receptor degradation.
D000818:Animals; D013006:Growth Hormone / Q000494:pharmacology; D006168:Guinea Pigs; D006801:Humans; D007328:Insulin / Q000494:pharmacology*; D007700:Kinetics; D008214:Lymphocytes / Q000378:metabolism*; D046911:Macromolecular Substances; D011972:Receptor, Insulin / Q000187:drug effects / Q000302:isolation & purification / Q000378:metabolism*; D013045:Species Specificity; D013552:Swine
D007328:Insulin; D046911:Macromolecular Substances; D013006:Growth Hormone; D011972:Receptor, Insulin