Figure 4.

PTEN inhibits Akt kinase activity and an activated form of Akt can override a PTEN induced G₁ block. (A) Inhibition of Akt kinase by wild-type PTEN. U2-OS cells were transfected with plasmids encoding the indicated proteins. After transfection, anti-T7 immunoprecipitates were prepared and used to phosphorylate a GST-peptide substrate in vitro. Autoradiography (Upper) and anti-T7 immunoblot (Lower) of the same membrane are shown. The black arrow indicates the position of the substrate. Results are representative of two experiments. (B) Quantitation of ³²P incorporation in A with a PhosphoImager. (C) Myr-Akt overrides a PTEN-induced G₁ block. 786-O cells were transiently cotransfected with pCD19 and plasmids encoding PTEN with empty vector (pLNCX) or with plasmids encoding the indicated Akt proteins. After transfection the cells were analyzed as in Fig. 1A. The mean and SEM of two experiments are shown.