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Liu CW, Wang RH, Dohadwala M, Schonthal AH, Villa-Moruzzi E, Berndt N: Inhibitory phosphorylation of PP1alpha catalytic subunit during the G (1)/S transition. J Biol Chem. 1999 Oct 8;274(41):29470-5.
We have shown earlier that, in cells expressing the retinoblastoma protein (pRB), a protein phosphatase (PP) 1alpha mutant (T320A) resistant to inhibitory phosphorylation by cyclin-dependent kinases (Cdks) causes G (1) arrest. In this study, we examined the cell cycle-dependent phosphorylation of PP1alpha in vivo using three different antibodies. PP1alpha was phosphorylated at Thr-320 during M-phase and again in late G (1)- through early S-phase. Inhibition of Cdk2 led to a small increase in PP1 activity and also prevented PP1alpha phosphorylation. In vitro, PP1alpha was a substrate for Cdk2 but not Cdk4. In pRB-deficient cells, phosphorylation of PP1alpha occurred in M-phase but not at G (1)/S. G (1)/S phosphorylation was at least partially restored after reintroduction of pRB into these cells. Consistent with this result, PP1alpha phosphorylated at Thr-320 co-precipitated with pRB during G (1)/S but was found in extracts immunodepleted of pRB in M-phase. In conjunction with earlier studies, these results indicate that PP1alpha may control pRB function throughout the cell cycle. In addition, our new results suggest that different subpopulations of PP1alpha regulate the G (1)/S and G (2)/M transitions and that PP1alpha complexed to pRB requires inhibitory phosphorylation by G (1)-specific Cdks in order to prevent untimely reactivation of pRB and permit transition from G (1)- to S-phase and/or complete S-phase. |
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