Androgen Receptor Phosphorylation and Activity Are Regulated by an Association with Protein Phosphatase 1
Androgen Receptor Phosphorylation and Activity Are Regulated by an Association with Protein Phosphatase 1
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DOI：
10.1074/jbc.m109.043133
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发表时间：
2009-09-18
影响因子：
4.8
通讯作者：
Balk, Steven P.
中科院分区：
文献类型：
--
作者：
Chen, Shaoyong;Kesler, Cristina T.;Balk, Steven P.
雄激素受体（AR）在多个位点被磷酸化以响应配体结合，但调节AR磷酸化的功能后果和机制仍有待建立。我们最初观察到，冈田酸，主要PPP家族丝氨酸/苏氨酸磷酸酶PP 2A和蛋白磷酸酶1（PP 1）的抑制剂，AR表达细胞类型依赖性的影响。PP 2A（fostriecin）和PP 1（针对PP 1 α催化亚基的互变霉素和siRNA）的更特异性抑制剂表明，PP 1和蛋白磷酸酶2A对AR蛋白和转录活性具有相反的作用。PP 1抑制增强了蛋白酶体介导的AR降解，而PP 1 α过表达增加了AR表达并显著增强了AR转录活性。共沉淀实验证明了AR-PP 1相互作用，而免疫荧光和核质分级显示雄激素刺激的前列腺癌细胞中AR和PP 1的核转位。用磷酸特异性AR抗体进行的研究表明，PP 1抑制显著增加了Ser-650的磷酸化，该位点位于AR铰链区，可介导核输出。值得注意的是，PP 1抑制导致野生型AR的核定位显著降低，但不改变S650 A突变体AR的总水平或核水平。这些发现揭示了PP 1通过Ser-650的去磷酸化在调节AR蛋白稳定性和核定位中的关键作用。此外，AR可能作为PP 1调节亚基发挥作用，并介导PP 1向染色质的募集，在染色质中它可以调节转录和剪接。
Androgen receptor (AR) is phosphorylated at multiple sites in response to ligand binding, but the functional consequences and mechanisms regulating AR phosphorylation remain to be established. We observed initially that okadaic acid, an inhibitor of the major PPP family serine/threonine phosphatases PP2A and protein phosphatase 1 (PP1), had cell type-dependent effects on ARexpression. More specific inhibitors of PP2A (fostriecin) and PP1 (tautomycin and siRNA against the PP1 alpha catalytic subunit) demonstrated that PP1 and protein phosphatase 2A had opposite effects on AR protein and transcriptional activity. PP1 inhibition enhanced proteasome-mediated AR degradation, while PP1 alpha overexpression increased AR expression and markedly enhanced AR transcriptional activity. Coprecipitation experiments demonstrated an AR-PP1 interaction, while immunofluorescence and nuclear-cytoplasmic fractionation showed androgen-stimulated nuclear translocation of both AR and PP1 in prostate cancer cells. Studies with phosphospecific AR antibodies showed that PP1 inhibition dramatically increased phosphorylation of Ser-650, a site in the AR hinge region shown to mediate nuclear export. Significantly, PP1 inhibition caused a marked decrease in nuclear localization of the wild-type AR, but did not alter total or nuclear levels of a S650A mutant AR. These findings reveal a critical role of PP1 in regulating AR protein stability and nuclear localization through dephosphorylation of Ser-650. Moreover, AR may function as a PP1 regulatory subunit and mediate PP1 recruitment to chromatin, where it can modulate transcription and splicing.