An FKBP‐Rapamycin‐Sensitive, Cyclin‐Dependent Kinase Activity that Correlates with the FKBP‐Rapamycin‐Induced G1 Arrest Point in MG‐63 Cellsa

The natural products FK506 and rapamycin (FIGURE 1) bind the same intracellular receptors, a family of FK506 and rapamycin binding proteins (FKBPs), but cause distinct biological phenomena.lJFK506 inhibits signaling emanating from T cell antigen receptors on T lymphocytes3v4 and immunoglobulin E (IgE) Fc receptors on mast cells: whereas rapamycin inhibits signaling emanating from some growth factor receptors, including interleukin-2 receptors on lymphocyte^^-^ and insulin receptors on hepatocytes6v7 (FIGURE 2).Three lines of evidence suggest that these natural products are prodrugs that become activated upon binding to an FKBP, i.e., the binding of each ligand by FKBP results in a gain of function rather than a loss of function.First, FK506 and rapamycin reciprocally antagonize each other's a ~t i o n .~, ~ Second, 506BD, a nonnatural FKBP ligand (FIGURE l), has no known biological activity but can reverse the actions of both FK506 and rapamycin.*Third, a strain of Saccharomyces cerevisiae with a null FKBP12 mutation is viable, but insensitive to both FK506 and r a p a m y ~i n .~-~~The gain-of-function model was confirmed when calcineurin, a Ca2+, calmodulin protein phosphatase, was shown to bind the FKBP12-FK506 complex, but neither component alone nor the FKBP-rapamycin complex.13Evidence that calcineurin is an in vivo target of FKBP-FK506 complexes has been generated by four independent methods.The ability of analogues of FK506 to inhibit TCR-mediated signaling in vivo correlates with their ability to inhibit calcineurin's phosphatase activity in vitro.l4 Second, the phosphatase activity of calcineurin in lymphocyte extracts was shown to be inhibited by pretreatment of the cells with FK506 but not rapamycin.15Third, overexpression of calcineurin in Jurkat T cells