Control of p70 S6 kinase by kinase activity of FRAP in vivo

WHEN complexed with the intracellular protein FKBP12, rapamy-cin is a potent immunosuppressant 1,2 and an inhibitor of a mitogen-stimulated signalling pathway that leads to activation of p70 S6 kinase 3-6 (p70 S6k ) and cyclin-dependent kinases 7-10 (CDKs). A recently cloned FKBP12-rapamycin-associated protein (FRAP/ RAFT) is the likely mediator of these effects 11,12 . Using FRAP variants that do not bind FKBP12-rapamycin, we demonstrate here that FRAP is a rapamycin-sensitive regulator of p70 S6k in vivo and that the kinase activity of FRAP is required for this regulation. In addition, we show that FRAP autophosphorylates in vitro . Consistent with an essential role for FRAP kinase activity in vivo , autophosphorylation of FRAP is inhibited by FKBP12-rapamycin. Deletion studies indicate that the kinase activity of FRAP alone is not sufficient for control of p70 S6k and that an amino-terminal domain in FRAP is also required.