Figure 6.

PPM1G regulates the interaction between p27 and 14-3-3θ, and the subcellular distribution of p27. A. PPM1G dephosphorylates Akt-phosphorylated p27pT198. Flag-p27, HA-AktDD, and Myc-PPM1G were co-transfected into 293T cells, and p27pT198 levels were determined by Western blot. B. PPM1G inhibits the interaction between p27 and 14-3-3θ. Flag-p27, HA-14-3-3θ, and Myc-PPM1G were co-transfected into 293T cells, and the interaction between p27 and 14-3-3θ was determined by Co-IP. IP: p27 was immunoprecipitated with anti-Flag antibody. 14-3-3θ levels in the IP complex were determined by Western blot using HA antibody. C. Knocking down PPM1G increases cytoplasmic localization of p27. The subcellular distribution of p27 in control and PPM1G-depleted HeLa cells was determined using subcellular fractionation assay. β-tubulin and Histone H3 were used as cytoplasmic and nuclear control. P values were determined by student’s t-test. D. A working model for T198 phosphorylated cytoplasmic translocation of p27 at early G1 phase. PPM1G counteracts Akt-induced p27pT198 to reduce interaction of p27 with 14-3-3, p27 nuclear export and subsequent degradation in cytoplasm.