Androgen Receptor Phosphorylation and Activity Are Regulated by an Association with Protein Phosphatase 1

Abstract

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 AR expression. More specific inhibitors of PP2A (fostriecin) and PP1 (tautomycin and siRNA against the PP1α 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 PP1α 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. 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 AR expression. More specific inhibitors of PP2A (fostriecin) and PP1 (tautomycin and siRNA against the PP1α 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 PP1α 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. Androgen receptor (AR) 3The abbreviations used are: ARandrogen receptorFBSfetal bovine serumPBSphosphate-buffered salineOAokadaic acidDAPI4′,6-diamidino-2-phenylindolePPprotein phosphatasePCaprostate cancerRLUrelative light unitDHTdihydrotestosterone. plays a central role in prostate cancer (PCa) development and progression, with androgen deprivation therapy being the standard systemic treatment for PCa (1Gelmann E.P. J. Clin. Oncol. 2002; 20: 3001-3015Crossref PubMed Scopus (769) Google Scholar). Unliganded AR associates with an Hsp90 chaperone complex and is rapidly degraded. Ligand binding stabilizes AR, enhances nuclear entry, and allows AR to recruit coactivator proteins to androgen-regulated genes. AR is phosphorylated at Ser-94 and Ser-650 in the absence of androgen, and androgen treatment further stimulates AR phosphorylation, primarily at multiple serine-proline sites (2Kuiper G.G. Brinkmann A.O. Biochemistry. 1995; 34: 1851-1857Crossref PubMed Scopus (52) Google Scholar, 3Zhou Z.X. Kemppainen J.A. Wilson E.M. Mol. Endocrinol. 1995; 9: 605-615Crossref PubMed Google Scholar, 4Gioeli D. Ficarro S.B. Kwiek J.J. Aaronson D. Hancock M. Catling A.D. White F.M. Christian R.E. Settlage R.E. Shabanowitz J. Hunt D.F. Weber M.J. J. Biol. Chem. 2002; 277: 29304-29314Abstract Full Text Full Text PDF PubMed Scopus (280) Google Scholar, 5Wong H.Y. Burghoorn J.A. Van Leeuwen M. De Ruiter P.E. Schippers E. Blok L.J. Li K.W. Dekker H.L. De Jong L. Trapman J. Grootegoed J.A. Brinkmann A.O. Biochem. J. 2004; 383: 267-276Crossref PubMed Scopus (44) Google Scholar). Similar to other steroid receptors, AR transcriptional activity and sensitivity to low levels of androgen can be enhanced by multiple kinases or kinase signaling pathways, which may contribute to tumor progression subsequent to androgen deprivation therapy. However, the kinases mediating AR phosphorylation at specific sites, and the functional importance of AR phosphorylation at particular residues, remain to be clearly defined. androgen receptor fetal bovine serum phosphate-buffered saline okadaic acid 4′,6-diamidino-2-phenylindole protein phosphatase prostate cancer relative light unit dihydrotestosterone. AR transcriptional activity may also be modulated directly or indirectly by serine/threonine phosphatases (6Ikonen T. Palvimo J.J. Kallio P.J. Reinikainen P. Jänne O.A. Endocrinology. 1994; 135: 1359-1366Crossref PubMed Scopus (155) Google Scholar). A recent study demonstrated SV40 small T-antigen-dependent loading of protein phosphatase 2A (PP2A) onto AR, with subsequent AR dephosphorylation (7Yang C.S. Vitto M.J. Busby S.A. Garcia B.A. Kesler C.T. Gioeli D. Shabanowitz J. Hunt D.F. Rundell K. Brautigan D.L. Paschal B.M. Mol. Cell. Biol. 2005; 25: 1298-1308Crossref PubMed Scopus (52) Google Scholar, 8Yang C.S. Xin H.W. Kelley J.B. Spencer A. Brautigan D.L. Paschal B.M. Mol. Cell. Biol. 2007; 27: 3390-3404Crossref PubMed Scopus (44) Google Scholar). AR also interacts with small C-terminal domain phosphatase 2 (SCP2), which is recruited by AR to the androgen-regulated PSA promoter and negatively regulates AR transcriptional activity, possibly by dephosphorylation of RNA polymerase II (9Thompson J. Lepikhova T. Teixido-Travesa N. Whitehead M.A. Palvimo J.J. Jänne O.A. EMBO J. 2006; 25: 2757-2767Crossref PubMed Scopus (31) Google Scholar). We report here that protein phosphatase 1 (PP1) increases AR protein stability and markedly enhances AR-mediated transcription. We show that AR binds the catalytic subunit of PP1, PP1α, and that androgens stimulate nuclear translocation of PP1α in conjunction with AR. Moreover, we determine that PP1 selectively dephosphorylates a specific site on the AR, Ser-650. Finally, consistent with a recent report that Ser-650 phosphorylation mediates AR nuclear export (10Gioeli D. Black B.E. Gordon V. Spencer A. Kesler C.T. Eblen S.T. Paschal B.M. Weber M.J. Mol. Endocrinol. 2006; 20: 503-515Crossref PubMed Scopus (138) Google Scholar), we demonstrate that PP1 inhibition markedly decreases nuclear AR. These findings demonstrate that PP1 is a direct positive regulator of AR nuclear expression and transcriptional activity and identifies the AR-PP1 interaction as a potential therapeutic target for PCa drug development. Sources were as follows: steroids, MG115, MG132, CHX, and anti-FLAG M2 beads (Sigma); OA (Roche Applied Science); tautomycin, and fostriecin (Calbiochem); anti-AR(PG21), anti-pAR-S81, anti-PP1α, anti-PP2A, and microcystin-agarose (Upstate Biotechnology); anti-AR (N441) (Lab Vision); anti-PSA (Biodesign); anti-β-tubulin (Chemicon); normal mouse serum (NMS), normal rabbit serum (NRS), protein G, and NE-PER kit (Pierce); goat anti-rabbit Alexa 594 (Molecular Probes); serum (FBS and CDS) (Hyclone); PP1α plasmid (Origene); control and AR RNAi (Dharmacon). Two separate PP1α RNAi were from Dharmacon (SMARTpool, M-008927-00) and Santa Cruz (sc-36299). AR expression and reporter plasmids, reporter assays, and real-time RT-PCR analyses have been described (12Chen S. Xu Y. Yuan X. Bubley G.J. Balk S.P. Proc. Natl. Acad. Sci. U.S.A. 2006; 103: 15969-15974Crossref PubMed Scopus (157) Google Scholar). Cells were grown in RPMI 1640 with 10% FBS (LNCaP and C4-2) or Dulbecco's modified Eagle's medium with 5% FBS. For androgen starvation, cells were grown in medium containing 5% charcoal dextran stripped FBS (CDS). For transfection, plasmid DNA or RNAi was transfected with Lipofectamine 2000. Empty pCDNA3.1 vector (Invitrogen) was used for equalization and a CMV-Renilla luciferase reporter (Promega) was an internal control. Luciferase was measured with a Dual-Luciferase assay kit (Promega). The ratio between firefly and Renilla luciferase is relative light unit (RLU), and the results are mean and standard deviation from triplicate samples. The Flag-AR-S650A mutant was generated with the Site-directed Mutagenesis kit (Stratagene). LNCaP cell stable lines expressing Flag-AR or Flag-AR-S650A were established by selection of cells grown in RPMI 1640 with 10% FBS and 1 μg/ml puromycin. Total proteins were isolated with 2% SDS, quantified with BCA reagent (Pierce), and equal amounts were analyzed on prepoured gels (Invitrogen). Cytoplasmic-nuclear fractionation was with the NE-PER kit (Pierce). For coprecipitation, cells were harvested in Triton lysis buffer (0.5% Triton X-100, 20 mm Tris-HCl, 150 mm NaCl, 5 mm EDTA, and 2 mm dithiothreitol) with protease and phosphatase inhibitors. Supernatants were with of or anti-AR (N441) to protein AR phosphorylation, AR was by anti-AR and equal amounts of total AR were with a of phosphospecific AR (7Yang C.S. Vitto M.J. Busby S.A. Garcia B.A. Kesler C.T. Gioeli D. Shabanowitz J. Hunt D.F. Rundell K. Brautigan D.L. Paschal B.M. Mol. Cell. Biol. 2005; 25: 1298-1308Crossref PubMed Scopus (52) Google Scholar, D. Black B.E. Gordon V. Spencer A. Kesler C.T. Eblen S.T. Paschal B.M. Weber M.J. Mol. Endocrinol. 2006; 20: 503-515Crossref PubMed Scopus (138) Google Scholar). For cells were in for in and in Triton for in cells were in bovine serum for and in for 2 in cells were in goat anti-rabbit Alexa 594 for 2 in and with 4′,6-diamidino-2-phenylindole for 5 in cells were on and observed with the We initially protein levels of transfected AR in a of cell lines treatment with okadaic acid a inhibitor of multiple phosphatases PP1 and PP2A Chem. 2002; 9: PubMed Scopus Google Scholar). OA increased AR protein levels in transfected and which was by increases in phosphorylation of by a The site is phosphorylated in response to androgen by serine/threonine and is with AR and transcriptional activity D. Ficarro S.B. Kwiek J.J. Aaronson D. Hancock M. Catling A.D. White F.M. Christian R.E. Settlage R.E. Shabanowitz J. Hunt D.F. Weber M.J. J. Biol. Chem. 2002; 277: 29304-29314Abstract Full Text Full Text PDF PubMed Scopus (280) Google Scholar, S. Xu Y. Yuan X. Bubley G.J. Balk S.P. Proc. Natl. Acad. Sci. U.S.A. 2006; 103: 15969-15974Crossref PubMed Scopus (157) Google Scholar). These results were consistent with a report that PP2A can sites in the AR domain PP2A interaction with AR in study was on loading by SV40 small (7Yang C.S. Vitto M.J. Busby S.A. Garcia B.A. Kesler C.T. Gioeli D. Shabanowitz J. Hunt D.F. Rundell K. Brautigan D.L. Paschal B.M. Mol. Cell. Biol. 2005; 25: 1298-1308Crossref PubMed Scopus (52) Google Scholar, 8Yang C.S. Xin H.W. Kelley J.B. Spencer A. Brautigan D.L. Paschal B.M. Mol. Cell. Biol. 2007; 27: 3390-3404Crossref PubMed Scopus (44) Google Scholar). However, OA had the opposite on AR protein expression and phosphorylation in where the androgen in AR protein expression and phosphorylation were both markedly by OA Significantly, OA caused a decrease in expression of the AR in the LNCaP and PCa cell lines decrease in AR protein was by that it was proteasome-mediated The of the AR is consistent with increased The effects of OA to inhibitors of PP1 and the major phosphatases that are by a PP2A specific increased AR protein expression in LNCaP cells and increased the expression of transfected AR in and cells not that PP2A negatively regulates AR expression in cells that not SV40 to the PP1 inhibitor a decrease in AR protein expression in LNCaP cells in medium with serum with or or in medium with serum expression of transfected AR in and cells that the of on AR in LNCaP cells was to PP1 we used siRNA to PP1 activity. cells have that PP1 catalytic and which are the direct of M. 2004; PubMed Scopus Google Scholar, J. Sci. 2002; PubMed Google Scholar). that LNCaP cells catalytic with expression of and being not we the protein of we used siRNA against to determine PP1α was AR expression in LNCaP cells. but not a control siRNA or an AR the levels of PP1α in LNCaP cells and in the cell Significantly, PP1α also caused a decrease in AR levels in the LNCaP cells in the or absence of and in the cells. a further we overexpression of PP1α expression of AR of PP1α markedly increased the expression of AR in both and and increases were observed in the absence and of and not study the of PP1 on AR expression in PCa cells expressing AR, LNCaP cells were with PP1α and AR to AR from AR in cells. shown in PP1α also increased Flag-AR in LNCaP the was not as marked as in and cells a of PP1 activity in LNCaP results that PP1 was AR expression and that PP1α was a major to have shown that the AR is rapidly while the AR is stable but a Z.X. Kemppainen J.A. Wilson E.M. Mol. Endocrinol. 1995; 9: PubMed Google Scholar). with treatment to protein in cells caused a in AR protein levels with a in the absence of androgen inhibitors increased transfected AR protein expression in the absence and of ligand consistent with that AR is through a L. Biochem. PubMed Scopus Google Scholar). determine AR protein expression by increased degradation, we the of protein was with shown in the of to cells with caused a further decrease in transfected AR protein that PP1 inhibition was AR protein determine was through the cells were to treatment in with shown in the decrease in AR expression be by that was of AR Similar experiments were in LNCaP cells to the effects of PP1 inhibition on AR. of LNCaP cells with for or caused a marked decrease in AR protein expression that was by the of that of AR in LNCaP was through the be that a in AR which a decrease in AR levels not observed in for and caused a further decrease in AR levels in LNCaP that PP1 inhibition was of AR protein AR was not observed in LNCaP cells with that was of the AR Similar results were in LNCaP cells in steroid medium FBS medium with or FBS medium a potential direct interaction between PP1 and AR, we experiments in cells with Flag-AR and Significantly, levels of PP1α, but were by from the cells in FBS or medium PP1α also be with AR from LNCaP cells further the interaction between PP1α and AR, we used which binds to PPP family phosphatases and been used to PP1 and PP2A and proteins N. T. 1994; PubMed Scopus Google Scholar). For experiments we used beads to PP1 from LNCaP cells grown in FBS we AR as as levels of PP1α with the beads AR was with the we from the to PP1 and determine the AR shown in of PP1α from the markedly AR levels AR and PP1α the and the AR by beads was markedly in the that AR was not with Similar results were from LNCaP cells in or The of AR was increased in the of but total AR levels were also enhanced by the binding not to be Finally, the interaction is not on PP1 catalytic activity, as it was not by PP1 inhibition with findings the that is a direct or by a PP1 regulatory subunit) interaction between AR and PP1α, and that interaction not The AR is in the and ligand binding stimulates nuclear of AR in LNCaP cells to further is a AR-PP1 interaction, we was nuclear translocation of PP1α in response to Significantly, the nuclear of a of PP1α in LNCaP and PCa but not in cells of nuclear proteins from LNCaP cells showed a nuclear of AR, by nuclear of PP1α These results further for an AR-PP1 interaction and that PP1α may with AR in the and in of AR nuclear The an interaction and nuclear translocation that PP1 may in of AR nuclear we PP1α can modulate AR transcriptional activity. shown in of PP1α enhanced the and transcriptional activity of the AR in LNCaP cells. determine PP1 AR to low levels of androgen, LNCaP cells were a of shown in AR activity be by in the absence of was enhanced further cells were with PP1α, was a in activity in the absence of did not to be a on as was on a control Renilla luciferase reporter not Significantly, PP1α did not the of for The of AR activity by in LNCaP cells transfected with reporter to amounts of AR, and can be increased by with AR. we LNCaP cells transfected with AR and an or of AR, PP1α, increased the response to at with PP1α increased activity, but dramatically enhanced reporter activity by the in the absence of PP1 of activity a response be clearly at but was observed at in the absence and of by which PP1 may be androgen-stimulated transcription is by AR protein expression as shown in PP1α not markedly total AR levels in LNCaP to LNCaP cells were with PP1α, reporter and shown in of PP1α and AR in LNCaP to a marked in AR reporter activity. in the was a in expression of the that increased AR protein is not the for enhanced reporter expression. Finally, to study the effects of PP1 are LNCaP and cells PCa cell in medium were transfected with AR and a of luciferase reporter or PP1α, and activity was shown in and of PP1α enhanced AR of androgen-regulated reporter genes. the marked increases in AR reporter expression in AR and LNCaP in conjunction with increases in AR protein in that PP1α can AR transcriptional activity in to effects on AR protein expression. with the with reporter showed that the PP1 inhibitor markedly decrease transcriptional activity Moreover, we that treatment of LNCaP cells expression of the androgen-regulated PSA and and with we also that PP1α siRNA PSA levels in LNCaP cells the effects of on AR protein AR transcriptional activity, we a response in LNCaP cells. the absence of tautomycin, AR protein levels were and PSA expression was at caused a decrease in total AR protein at but AR levels at AR phosphorylation at also at the of relative to total AR was at PSA levels were markedly by at low and did not at to the increases in total AR Similar results were in which have AR activity in the absence of and to levels of the PP1α and demonstrate that PP1α can markedly AR transcriptional activity, and that mechanisms in to AR protein may contribute to to tautomycin, the PP2A inhibitor fostriecin increased AR protein and markedly increased both and PSA in LNCaP cells that PP2A negatively regulates AR in cells that not SV40 and that protein phosphatases have effects on AR expression and phosphorylation The results showed that PP1α both AR and it transcriptional activity, but it was not were direct effects of PP1 on AR effects of PP1 on other proteins that AR stability and activity. we used a of phosphospecific antibodies at AR phosphorylation sites to determine AR phosphorylation at specific sites was increased by PP1 inhibition (7Yang C.S. Vitto M.J. Busby S.A. Garcia B.A. Kesler C.T. Gioeli D. Shabanowitz J. Hunt D.F. Rundell K. Brautigan D.L. Paschal B.M. Mol. Cell. Biol. 2005; 25: 1298-1308Crossref PubMed Scopus (52) Google Scholar, D. Black B.E. Gordon V. Spencer A. Kesler C.T. Eblen S.T. Paschal B.M. Weber M.J. Mol. Endocrinol. 2006; 20: 503-515Crossref PubMed Scopus (138) Google Scholar). The AR was from or control LNCaP and equal amounts of total AR protein were with phosphospecific AR treatment in a marked in the phosphorylation of Ser-650, and a in and with at or A and and not that PP1 may selectively AR at Ser-650, we dephosphorylation of other sites or the that PP1 inhibition is kinases that target Ser-650 D. Ficarro S.B. Kwiek J.J. Aaronson D. Hancock M. Catling A.D. White F.M. Christian R.E. Settlage R.E. Shabanowitz J. Hunt D.F. Weber M.J. J. Biol. Chem. 2002; 277: 29304-29314Abstract Full Text Full Text PDF PubMed Scopus (280) Google Scholar, D. Black B.E. Gordon V. Spencer A. Kesler C.T. Eblen S.T. Paschal B.M. Weber M.J. Mol. Endocrinol. 2006; 20: 503-515Crossref PubMed Scopus (138) Google Scholar). a study of sites by PP2A in the AR that Ser-650 phosphorylation was not by consistent with Ser-650 being a site by PP1 (7Yang C.S. Vitto M.J. Busby S.A. Garcia B.A. Kesler C.T. Gioeli D. Shabanowitz J. Hunt D.F. Rundell K. Brautigan D.L. Paschal B.M. Mol. Cell. Biol. 2005; 25: 1298-1308Crossref PubMed Scopus (52) Google Scholar). The functional of AR phosphorylation at sites not been but recent results that AR nuclear export can be enhanced by Ser-650 phosphorylation and by OA treatment (10Gioeli D. Black B.E. Gordon V. Spencer A. Kesler C.T. Eblen S.T. Paschal B.M. Weber M.J. Mol. Endocrinol. 2006; 20: 503-515Crossref PubMed Scopus (138) Google Scholar, Kelley J.B. Gioeli D. C.S. Spencer A. Paschal B.M. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). we treatment the nuclear localization of AR in LNCaP cells. treatment of LNCaP cells in medium a marked in nuclear AR and a decrease in AR The of nuclear with a marked decrease in the ratio of nuclear to AR. also caused a in nuclear AR in cells that were with inhibitors consistent with a direct on AR localization that is increased results that PP1 inhibition is AR Ser-650 phosphorylation, nuclear and Finally, to determine effects of PP1 inhibition were on increased Ser-650 phosphorylation, we an S650A mutant AR. For experiments we generated LNCaP cells transfected with wild-type or S650A AR. observed with the AR, treatment of cells expressing the wild-type AR caused a marked of nuclear Flag-AR and a decrease in total Flag-AR expression did not decrease levels of the in the or decrease total levels that effects of PP1 inhibition are by increased Ser-650 We initially observed cell effects on AR of a phosphatase a specific PP1 inhibitor and PP1 we that proteasome-mediated AR was increased in response to PP1 in cell overexpression of PP1α increased AR expression and transcriptional activity. the PP2A specific inhibitor fostriecin enhanced AR protein expression and a role for PP2A in AR in cells that not SV40 small (7Yang C.S. Vitto M.J. Busby S.A. Garcia B.A. Kesler C.T. Gioeli D. Shabanowitz J. Hunt D.F. Rundell K. Brautigan D.L. Paschal B.M. Mol. Cell. Biol. 2005; 25: 1298-1308Crossref PubMed Scopus (52) Google Scholar). an between AR and PP2A not be a and interaction, we an AR-PP1 interaction by that was of treatment also the nuclear translocation of both AR and PP1α, a interaction and that AR may function as a PP1 regulatory subunit for nuclear of Significantly, PP1 inhibition caused a and specific in Ser-650 Moreover, consistent with recent that Ser-650 phosphorylation stimulates AR nuclear export (10Gioeli D. Black B.E. Gordon V. Spencer A. Kesler C.T. Eblen S.T. Paschal B.M. Weber M.J. Mol. Endocrinol. 2006; 20: 503-515Crossref PubMed Scopus (138) Google Scholar), we that PP1 inhibition markedly nuclear AR. Finally, effects of PP1 inhibition on AR levels and nuclear localization were by a S650A that are to phosphorylation at results demonstrate that PP1 interacts with AR and enhances that AR to PP1 nuclear and that PP1 enhances AR nuclear expression by dephosphorylation of Ser-650. The in proteasome-mediated of AR in response to PP1 inhibition by may be to increased nuclear export and subsequent to in the However, Ser-650 is also in a that may directly target AR for through or T. A. Acad. Sci. 2004; PubMed Scopus Google Scholar, S. J. Biol. Chem. 2002; 277: Full Text Full Text PDF PubMed Scopus Google Scholar, A. Jänne O.A. Palvimo J.J. J. Biol. Chem. 2002; 277: Full Text Full Text PDF PubMed Scopus Google Scholar). Significantly, report that Ser-650 phosphorylation enhances the interaction between AR and the S. 2006; PubMed Scopus (44) Google Scholar). PP1 binding is not in AR by ligand binding or with other proteins may PP1 to and indirectly AR localization or C.S. Xin H.W. Kelley J.B. Spencer A. Brautigan D.L. Paschal B.M. Mol. Cell. Biol. 2007; 27: 3390-3404Crossref PubMed Scopus (44) Google Scholar). experiments that was a direct interaction between AR and PP1α, an interaction by a PP1 regulatory protein be of PP1 catalytic to regulatory is in by a M. 2004; PubMed Scopus Google Scholar, J. Sci. 2002; PubMed Google Scholar, D.F. P. D. EMBO J. PubMed Scopus Google Scholar, P. M. M. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). The AR site that which is in the DNA binding domain and is of the DNA that site is not for AR-PP1 binding not However, further are to determine AR is directly with the PP1 catalytic subunit through site or other A. X. E. A. Biol. 2004; PubMed Scopus Google Scholar). it that binding is through a PP1 regulatory prostate cancer overexpression of PP1 markedly AR-mediated while PP1 inhibition AR transcriptional activity. The of in total AR protein expression were and to for effects on transcription. nuclear AR clearly a further for the of AR transcriptional activity by PP1 is to with multiple proteins that and K. T. M. M. A. M. S. J. Biol. Chem. 2002; 277: Full Text Full Text PDF PubMed Scopus Google Scholar, J. Biol. Chem. 2002; 277: Full Text Full Text PDF PubMed Scopus Google Scholar, E. S. M. Biol. PubMed Scopus Google Scholar, A. M. P. Van A. V. M. M. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar, C.S. C.S. J. Biol. Chem. 2005; Full Text Full Text PDF PubMed Scopus Google Scholar, M.J. Mol. Cell. Full Text Full Text PDF PubMed Scopus Google Scholar). in to mediating Ser-650 dephosphorylation and nuclear AR, PP1 may be recruited to AR target and function as a coactivator of AR transcriptional activity. Moreover, nuclear recruitment of PP1 by AR a by which AR both transcription and of specific genes. These are We Gioeli of for the AR We and Xin Yuan for and the Flag-AR for and for