Direct inhibition of the signaling functions of the mammalian target of rapamycin by the phosphoinositide 3‐kinase inhibitors, wortmannin and LY294002.

The immunosuppressant, rapamycin, inhibits cell growth by interfering with the function of a novel kinase, termed mammalian target of rapamycin (mTOR). The putative catalytic domain of mTOR is similar to those of mammalian and yeast phosphatidylinositol (PI) 3‐kinases. This study demonstrates that mTOR is a component of a cytokine‐triggered protein kinase cascade leading to the phosphorylation of the eukaryotic initiation factor‐4E (eIF‐4E) binding protein, PHAS‐1, in activated T lymphocytes. This event promotes G1 phase progression by stimulating eIF‐4E‐dependent translation initiation. A mutant YAC‐1 T lymphoma cell line, which was selected for resistance to the growth‐inhibitory action of rapamycin, was correspondingly resistant to the suppressive effect of this drug on PHAS‐1 phosphorylation. In contrast, the PI 3‐kinase inhibitor, wortmannin, reduced the phosphorylation of PHAS‐1 in both rapamycin‐sensitive and ‐resistant T cells. At similar drug concentrations (0.1–1 microM), wortmannin irreversibly inhibited the serine‐specific autokinase activity of mTOR. The autokinase activity of mTOR was also sensitive to the structurally distinct PI 3‐kinase inhibitor, LY294002, at concentrations (1–30 microM) nearly identical to those required for inhibition of the lipid kinase activity of the mammalian p85‐p110 heterodimer. These studies indicate that the signaling functions of mTOR, and potentially those of other high molecular weight PI 3‐kinase homologs, are directly affected by cellular treatment with wortmannin or LY294002.