The mammalian target of rapamycin controls cell motility through its effectors S6K1 and 4E-BP1

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Rapamycin and its derivatives CCI-779, RAD001 and AP23573, inhibitors of the mammalian target of rapamycin (mTOR), are in clinical trials as novel anticancer agents. Recent findings have shown that the mTOR inhibitors not only suppress tumor cell proliferation and growth, but also inhibit tumor cell motility. However, the mechanism by which rapamycin inhibits cell motility remains poorly understood. Here we show that rapamycin inhibited type I insulin-like growth factor (IGF-I)-stimulated motility of rhabdomyosarcoma cells (Rh1 and Rh30). Expression of a mutant of mTOR (mTORrr, S2035I), preventing binding of rapamycin-FKBP12 complex to mTOR, rescued rapamycin-inhibited cell motility. Further, cells expressing a kinase dead mTORrr (D2338A) remained sensitive to rapamycin, suggesting that rapamycin suppresses cell motility through inhibition of mTOR function. The kinase activity of mTOR is essential for cell motility. We also determined the roles of two downstream effector molecules of mTOR, p70 S6 kinase 1 (S6K1) and eukaryotic initiation factor 4E (eIF4E) binding protein 1 (4E-BP1), in the regulation of cell motility. Cells infected with a replication-deficient adenoviral recombinant expressing constitutively active and rapamycin-resistant mutant of S6K1 (Ad-S6K1-ED3E), but not with an adenovirus expressing wild-type S6K1 (Ad-S6K1-wt), or a control viral vector (Ad-GFP), conferred to partial resistance to rapamycin. Further, downregulation of 4E-BP1 by RNA interference attenuated rapamycin inhibition of cell motility. The results indicate that mTOR controls cell motility through both S6K1 and 4E-BP1 signaling pathways. Supported by Stiles Award and Feist-Weiller Cancer Center Award, LSUHSC-Shreveport.