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The B56?3 regulatory subunit-containing protein phosphatase 2A outcompetes Akt to regulate p27KIP1 subcellular localization by selectively dephosphorylating phospho-Thr157 of p27KIP1.

ABSTRACT: The B56?-containing protein phosphatase 2A (PP2A-B56?) has been postulated to have tumor suppressive functions. Here, we report regulation of p27KIP1 subcellular localization by PP2A-B56?3. B56?3 overexpression enhanced nuclear localization of p27KIP1, whereas knockdown of B56?3 decreased p27KIP1 nuclear localization. B56?3 overexpression decreased phosphorylation at Thr157 (phospho-Thr157), whose phosphorylation promotes cytoplasmic localization of p27KIP1, whereas B56?3 knockdown significantly increased the level of phospho-Thr157. In vitro, PP2A-B56?3 catalyzed dephosphorylation of phospho-Thr157 in a dose-dependent and okadaic acid-sensitive manner. B56?3 did not increase p27KIP1 nuclear localization by down-regulating the upstream kinase Akt activity and outcompeted a myristoylated constitutively active Akt (Aktca) in regulating Thr157 phosphorylation and subcellular localization of p27KIP1. In addition, results of interaction domain mapping revealed that both the N-terminal and C-terminal domains of p27 and a domain at the C-terminus of B56?3 are required for interaction between p27 and B56?3. Furthermore, we demonstrated that p27KIP1 levels are positively correlated with B56? levels in both non-tumor and tumor parts of a set of human colon tissue specimens. However, positive correlation between nuclear p27KIP1 levels and B56? levels was found only in the non-tumor parts, but not in tumor parts of these tissues, implicating a dysregulation in PP2A-B56?3-regulated p27KIP1 nuclear localization in these tumor tissues. Altogether, this study provides a new mechanism by which the PP2A-B56?3 holoenzyme plays its tumor suppressor role.

SUBMITTER: Lai TY

PROVIDER: S-EPMC4826225 | biostudies-literature | 2016 Jan

REPOSITORIES: biostudies-literature

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The B56γ3 regulatory subunit-containing protein phosphatase 2A outcompetes Akt to regulate p27KIP1 subcellular localization by selectively dephosphorylating phospho-Thr157 of p27KIP1.

Lai Tai-Yu TY Yen Chia-Jui CJ Tsai Hung-Wen HW Yang Yu-San YS Hong Wei-Fu WF Chiang Chi-Wu CW

Oncotarget 20160101 4

The B56γ-containing protein phosphatase 2A (PP2A-B56γ) has been postulated to have tumor suppressive functions. Here, we report regulation of p27KIP1 subcellular localization by PP2A-B56γ3. B56γ3 overexpression enhanced nuclear localization of p27KIP1, whereas knockdown of B56γ3 decreased p27KIP1 nuclear localization. B56γ3 overexpression decreased phosphorylation at Thr157 (phospho-Thr157), whose phosphorylation promotes cytoplasmic localization of p27KIP1, whereas B56γ3 knockdown significantly ...[more]

PMID: 26684356

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Project description:The Cyclin-dependent kinase inhibitor 1B (p27Kip1) is a key protein in the decision between proliferation and cell cycle exit. Quiescent cells show nuclear p27Kip1, but this protein is exported to the cytoplasm in response to proliferating signals. We recently reported that catalase treatment increases the levels of p27Kip1 in vitro and in vivo in a murine model. In order to characterize and broaden these findings, we evaluated the regulation of p27Kip1 by hydrogen peroxide (H(2)O(2)) in human melanoma cells and melanocytes. We observed a high percentage of p27Kip1 positive nuclei in melanoma cells overexpressing or treated with exogenous catalase, while non-treated controls showed a cytoplasmic localization of p27Kip1. Then we studied the levels of p27Kip1 phosphorylated (p27p) at serine 10 (S10) and at threonine 198 (T198) because phosphorylation at these sites enables nuclear exportation of this protein, leading to accumulation and stabilization of p27pT198 in the cytoplasm. We demonstrated by western blot a decrease in p27pS10 and p27pT198 levels in response to H(2)O(2) removal in melanoma cells, associated with nuclear p27Kip1. Melanocytes also exhibited nuclear p27Kip1 and lower levels of p27pS10 and p27pT198 than melanoma cells, which showed cytoplasmic p27Kip1. We also showed that the addition of H(2)O(2) (0.1 µM) to melanoma cells arrested in G1 by serum starvation induces proliferation and increases the levels of p27pS10 and p27pT198 leading to cytoplasmic localization of p27Kip1. Nuclear localization and post-translational modifications of p27Kip1 were also demonstrated by catalase treatment of colorectal carcinoma and neuroblastoma cells, extending our findings to these other human cancer types. In conclusion, we showed in the present work that H(2)O(2) scavenging prevents nuclear exportation of p27Kip1, allowing cell cycle arrest, suggesting that cancer cells take advantage of their intrinsic pro-oxidant state to favor cytoplasmic localization of p27Kip1.

Dataset Information

The B56?3 regulatory subunit-containing protein phosphatase 2A outcompetes Akt to regulate p27KIP1 subcellular localization by selectively dephosphorylating phospho-Thr157 of p27KIP1.

Publications

The B56γ3 regulatory subunit-containing protein phosphatase 2A outcompetes Akt to regulate p27KIP1 subcellular localization by selectively dephosphorylating phospho-Thr157 of p27KIP1.

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