- Open Access
Membrane androgen receptor activation triggers down-regulation of PI-3K/Akt/NF-kappaB activity and induces apoptotic responses via Bad, FasL and caspase-3 in DU145 prostate cancer cells
- Natalia Papadopoulou†1,
- Ioannis Charalampopoulos†2,
- Vasileia Anagnostopoulou1,
- Georgios Konstantinidis1,
- Michael Föller3,
- Achilleas Gravanis2,
- Konstantinos Alevizopoulos4,
- Florian Lang3 and
- Christos Stournaras1Email author
© Papadopoulou et al; licensee BioMed Central Ltd. 2008
- Received: 09 September 2008
- Accepted: 03 December 2008
- Published: 03 December 2008
Recently we have reported membrane androgen receptors-induced apoptotic regression of prostate cancer cells regulated by Rho/ROCK/actin signaling. In the present study we explored the specificity of these receptors and we analyzed downstream effectors controlling survival and apoptosis in hormone refractory DU145-prostate cancer cells stimulated with membrane androgen receptor-selective agonists.
Using membrane impermeable conjugates of serum albumin covalently linked to testosterone, we show here down-regulation of the activity of pro-survival gene products, namely PI-3K/Akt and NF-κB, in DU145 cells. Testosterone-albumin conjugates further induced FasL expression. A FasL blocking peptide abrogated membrane androgen receptors-dependent apoptosis. In addition, testosterone-albumin conjugates increased caspase-3 and Bad protein activity. The actin cytoskeleton drug cytochalasin B and the ROCK inhibitor Y-27632 inhibited FasL induction and caspase-3 activation, indicating that the newly identified Rho/Rock/actin signaling may regulate the downstream pro-apoptotic effectors in DU145 cells. Finally, other steroids or steroid-albumin conjugates did not interfere with these receptors indicating testosterone specificity.
Collectively, our results provide novel mechanistic insights pointing to specific pro-apoptotic molecules controlling membrane androgen receptors-induced apoptotic regression of prostate cancer cells and corroborate previously published observations on the potential use of membrane androgen receptor-agonists as novel anti-tumor agents in prostate cancer.
- Androgen Receptor
- Prostate Cancer Cell
- LNCaP Cell
- Prostate Cancer Cell Line
An increasing body of scientific evidence points to the existence of two types of androgen receptors: (a) intracellular androgen receptors (iARs) mediating genomic androgen signals resulting in receptor dimerization, nuclear translocation and subsequent activation of androgen-specific target genes (reviewed in ) and (b) membrane androgen receptors (mARs) triggering non-genomic signals manifested within minutes of androgen binding (reviewed in [2, 3]). Although the exact molecular identity of mAR still remains unknown, it is believed that mAR may represent either (I) a pool of iAR targeted to the plasma membrane and/or associated membrane structures (e.g. lipid rafts or caveolae) mediating rapid androgen effects in the absence of transcriptional activity (reviewed in ) or (II) an unknown G-protein coupled receptor (GPCR) (or a receptor associated with a GPCR) triggering a variety of iAR-independent signaling cascades. These cascades typically result in increased intracellular [Ca2+]i and inositol 1,4,5-triphosphate formation, are sensitive to pertussis toxin inhibition [5, 6] and cannot be blocked by anti-androgens [7, 8]. Rapid, non-genomic androgen actions have been reported in various cell types including macrophages and T cells [9, 10], LNCaP , T47D , MCF7 , DU145 , C6 , PC12  or VSMC cells .
We and others have recently characterized mAR-dependent signaling events in prostate and breast cancer cell lines [8, 11, 12, 17]. Using non-permeable androgen derivatives that do not bind to iAR, namely conjugates of testosterone covalently linked to bovine serum albumin (testosterone-BSA), we have specifically shown that activation of mAR results in actin reorganization of iAR+/mAR+ LNCaP and iAR-/mAR+ DU145 prostate cancer cell lines [8, 17]. Furthermore, we have shown that testosterone-BSA induces apoptotic regression of LNCaP and DU145 cells in vitro and in mouse xenografts in vivo [13, 18]. Finally, testosterone-BSA suppresses cell motility and potentiates paclitaxel-mediated cytotoxicity both in vitro and in vivo [12, 18]. However, the specific pro-apoptotic molecules controlling the mAR-induced apoptosis in prostate cancer cells remained unknown.
In the present study we have analyzed the specificity of mAR and the activity of downstream gene products playing a prominent role in survival and apoptosis in DU145 prostate cancer cells. Our results show that testosterone-BSA suppresses PI-3K activity, inhibits Akt function and finally inactivates the pro-survival transcription factor NF-κB. We further report mAR-dependent suppression in the phosphorylation/inactivation of the pro-apoptotic Bad protein, stimulation of FasL expression and induction of caspase-3 activity. Taken together, our results provide new mechanistic insight into specific mAR-dependent apoptosis of prostate cancer cells.
Cell culture and transfections
The DU145 human prostate cancer cell line was obtained from the American Type Culture Collection (Manassas, VA) and was studied between passages 60 and 70. DU145 cells fail to respond to androgen treatment owing to the expression of non-functional iAR , or to complete lack of iAR according to other studies [20, 21].
Preparation of steroid solution
Before each experiment testosterone-3-(O-carboxymethyl) oxime-BSA, referred to as testosterone-BSA, dihydrotestosterone (DHT), estradiol-BSA and dexamethasone (Sigma), were dissolved in serum-free culture medium at a final concentration of 10-5 M. The steroid-albumin conjugates-stock solutions were incubated for 30 min at room temperature with 0.3% charcoal and 0.03% dextran, centrifuged at 3000 × g and passed through a 0.45 μm filter to remove any potential contamination with free steroid. Testosterone-BSA, estradiol-BSA, dexamethasone and DHT solutions were used at a final concentration of 10-7 M throughout all studies. If not otherwise stated all treatments and incubations with steroids including apoptosis assays were performed in serum-containing medium.
Measurement of F/G actin ratio by Triton X-100 fractionation
The Triton X-100 soluble G-actin containing and insoluble F-actin containing fractions of cells exposed to testosterone-BSA and DHT were prepared as previously described . An increase of the triton-insoluble (F) to triton-soluble (G) actin ratio is indicative of actin polymerization.
Immunoprecipitation and Western blot analysis
DU145 cells treated or not (control cells) with testosterone-BSA were washed twice with ice-cold phosphate-buffered saline and suspended in cold lysis buffer containing 1% Nonidet P-40, 20 mM Tris (pH 7.4) and 137 mM NaCl, supplemented with protease and phosphatase inhibitors. Cleared lysates were pre-adsorbed with protein A-Sepharose beads (Amersham) for 1 h at 4°C. Equal amounts of the supernatants were subjected to immunoprecipitation using an anti-phosphotyrosine (PY20) antibody (Santa Cruz Biotechnology) and protein A-Sepharose beads. For immunoblot analysis the immunoprecipitates and equal amounts of total protein extracts were suspended in Laemmli's sample buffer and separated by SDS-PAGE. For Fas ligand expression studies cells were pretreated or not with 10-7 M cytochalasin B (Biomol Research Laboratories, PA), or 10 μM Y-27632 (Calbiochem), and stimulated with 10-7 M testosterone-BSA for the time periods indicated in the figure legends.
Proteins were transferred onto nitrocellulose membranes and blotted with rabbit polyclonal anti-PI-3K p85 (Upstate) (1:1000 dilution), rabbit polyclonal anti-phospho-Akt Ser473, anti-phospho-Akt Thr308 or anti-Akt (total) (Cell Signaling) (1:500 dilution), rabbit polyclonal anti-Fas-L (Q20, Santa Cruz) (1:200 dilution), phospho- and total Bad antibodies (Santa Cruz Biotechnology, Santa Cruz, CA, USA, 1:1000 dilution). Secondary antibodies used were horseradish peroxidase-conjugated anti-mouse IgG (Chemicon), and horseradish peroxidase-conjugated anti-rabbit IgG (Immunotech, France). Then, the membranes were exposed to Kodak X-Omat AR films. A PC-based Image Analysis program was used to quantify the intensity of each band (Image Analysis, Inc., Ontario, Canada).
NF-κB Transcription factor Assay
A non-radioactive NF-κB p50/p65 Transcription Factor Assay was used to detect specific transcription factor DNA binding activity in nuclear extracts (Chemicon, San Diego, CA, USA). A double stranded biotinylated oligonucleotide containing the consensus sequence for NF-κB binding (5'-GGGACTTTCC-3'), was mixed with cellular (nuclear) extract pre-treated or not with 10-7 M cytochalasin B. After co-incubation the active form of NF-κB contained in the nuclear extract binds to its consensus sequence. Thereafter, the extract/probe/buffer mixture was directly transferred to the streptavidin-coated plate. The biotinylated double stranded oligonucleotide bound by active NF-κB protein was immobilized, and any inactive unbound material was washed away. The bound NF-κB transcription factor subunits, p50/p65, were detected with specific primary antibodies. An HRP-conjugated secondary antibody was then used for detection and quantification in a spectrophotometric plate reader. By loading the same amount of total protein in each sample, we ensured the exact normalization for all cases.
Measurement of apoptosis
APOPercentage apoptosis assay
DU145 cells (in RPMI 1640, supplemented with 25 mM HEPES, 2 mM L-Glutamine and 10% FBS) were cultured in 96-well plates for the APOPercentage apoptosis assay (Biocolor Ltd., Belfast, Ireland). In the presence or absence of 10-7 M flutamide (Sigma), cells were stimulated or not with 10-7 M of the following steroids in serum-supplemented medium: testosterone-BSA (Testo-BSA), dihydrotestosterone (DHT), estradiol-BSA (E2-BSA) and dexamethasone (DEXA) or 10-7 M BSA for 24 hours. Untreated cells cultured in serum free medium were used as positive control for the apoptotic response.
DU145 cells (in RPMI 1640, supplemented with 25 mM HEPES, 2 mM L-Glutamine and 10% FBS) were cultured in 60 mm plates for FACS analysis and determination of Fas expression levels. After pre-treatment with a monoclonal Ab to Fas, (Fas-blocking peptide, 805-C10-C100, Alexis Biochemicals, Axxora LLC, San Diego, USA), cells were stimulated or not with 10-7 M testosterone-BSA in serum-supplemented medium for the time periods indicated in the figure legends. Untreated cells cultured in serum-free medium were used as a positive control for the apoptotic response. At the end of the respective treatment cells were harvested in PBS and stained with the Annexin V-FITC Apoptosis Detection kit I (BD Pharmingen TM, San Diego, CA) according to the manufacturer's instructions. They were analyzed within 1 h by flow cytometry using a FACSArray Apparatus (BD Biosciences) and CellQuest (BD Biosciences) and ModFit LT (Verify software, Topsham, MN) software.
The activity of caspase-3 was measured in whole cell lysates pre-treated or not with either 10-7 M cytochalasin B, or 10 μM Y-27632 and then stimulated with 10-7 M testosterone-BSA for the time periods indicated in the figure legends, using the Clontech ApoAlert® Caspase Colorimetric Assay kit according to the manufacturers' instructions. Caspase-3 activity was determined by incubating lysates with a caspase-3 substrate (the peptide DEVD conjugated to the chromophor p-nitroaniline) for 2 h at 37°C. The absorbance of each sample was measured at 405 nm by using a 96-well colorimetric plate reader.
Testosterone and testosterone-albumin-conjugates trigger similarly specific activation of mAR in DU145 cells
Testosterone-BSA induces long term inhibition of PI-3K activity in DU145 cells
Testosterone-BSA inhibits long term Akt activity and induces Bad de-phosphorylation in DU145 cells
Testosterone-BSA suppresses NF-κB activity in DU145 cells
Testosterone-BSA induces FasL expression in DU145 cells
mAR-stimulation by testosterone-BSA triggers caspase-3 activation in DU145 cells
Previous studies in prostate cancer cell lines have established a clear role for membrane androgen receptors in the induction of apoptotic responses via actin cytoskeleton reorganization . Furthermore, it has been proposed that specific mAR-activating ligands, namely testosterone-serum albumin conjugates, may be developed as novel drug candidates for the treatment of mAR+ prostate tumors [17, 18].
Using pharmacological inhibitors, dominant negative alleles and various functional assays, we were able to identify in previous studies a series of key denominators of mAR function in prostate cancer cell lines [8, 17]. Specifically, we have identified a FAK/PI-3K/Rac/Cdc42 pathway triggered by testosterone-BSA in LNCaP cells, resulting in actin cytoskeleton reorganization . In DU145 cells FAK and PI-3K were shown to be constitutively active, and testosterone-BSA triggers actin rearrangements via a Rho/ROCK/LIMK2/ADF-destrin signaling pathway . Interestingly, the same Rho/ROCK pathway operates in LNCaP cells, downstream of FAK/PI-3K/Rac1 . Finally, actin cytoskeleton disrupting agents and ROCK inhibitors were shown to block mAR-dependent apoptosis in both cell lines, indicating that Rho/ROCK/actin signaling is a key regulator of apoptotic responses . However the identification of the specific downstream players implicated in apoptosis remained unknown.
In the present work we characterized the specificity of mAR by using a series of BSA-conjugated and free steroid hormones, providing clear evidence for testosterone specificity. We further explored the mechanism of cell death triggered by mAR-stimulation by analyzing the expression and activity of several gene products and pathways involved in the regulation of survival and apoptosis of DU145 prostate cancer cells. Using testosterone-BSA as a specific mAR ligand, we show here that mAR activation results in almost complete down-regulation of the activity of PI-3K, Akt and NF-κB in DU145 cells. Concurrently, testosterone-BSA induces FasL expression, activates Bad and up-regulates the activity of caspase-3, indicating that mAR-stimulation affects prominent pro-apoptotic regulators [39, 40]. Importantly, these effects are blocked by actin cytoskeleton disrupting agents or the ROCK inhibitor Y-27632, providing additional evidence that actin reorganization, shown to be a prominent event in mAR-stimulated prostate cancer cells [Fig 1A and ref [7, 8, 41]], and the newly identified Rho/ROCK signaling  control testosterone-BSA-induced apoptosis in DU145 cells. These data further corroborate the hypothesis postulated by several research groups that actin dynamics reorganization is a key regulator of apoptotic responses (for reviews see [42–44]). Taken together our results offer further mechanistic insights into the control of survival and apoptosis downstream of mAR, pointing to specific pro-apoptotic molecular effectors acting most probably downstream of Rho/Rock/actin.
Although we cannot rule out that the observed changes in the expression and activity of all analyzed proteins are the consequence rather than the cause of mAR-dependent apoptosis, the data clearly underscore the key role of mAR-activating ligands in the selective elimination of DU145 cells. Moreover, these receptors are specific for testosterone and testosterone-albumin conjugates, since other steroid hormones-conjugated or not-failed to exhibit any pro-apoptotic activity. Notably, these cells typically represent an aggressive pre-clinical hormone-refractory cell line model used to assess the anti-tumor ability of chemotherapeutic drugs, as they (I) are devoid of functional intracellular androgen receptors (iARs) and (II) fail to respond to androgen treatment . Based on these results, mAR may be a novel target that can be used for the selective elimination of mAR+ prostate cancer cells independently of the functional status of the intracellular androgen receptor. Interestingly, mAR is selectively over-expressed in biopsy samples from aggressive, high-Gleason prostate tumors in comparison to samples from benign prostate hyperplasia patients or healthy subjects [45, 46].
Future experiments will focus on the identification of additional signaling targets downstream of mAR and the characterization of functional synergies of mAR-dependent signals with other pathways activated in prostate cancer. Characterization of the functional interplay between membrane and intracellular androgen receptors may contribute to the understanding of the apparent discrepancy in the actions of androgens inducing both proliferation and death within a given cell. Our present findings elucidating at least parts of the mAR-induced molecular pro-apoptotic machinery in DU145 cells provide novel insights in membrane GPCR mediated non-genomic androgen actions.
We would like to thank the Herakleitos EPEAEK program (supported by the European Social Fund and National Recourses) the KESY-2003 program and the Deutsche Forschungsgemeinschaft (DFG, Mercator program, GRK 1302/1 and SFB 773) for supporting this work.
- Heinlein CA, Chang C: Androgen receptor in prostate cancer. Endocr Rev. 2004, 25: 276-308. 10.1210/er.2002-0032View ArticlePubMedGoogle Scholar
- Rahman F, Christian HC: Non-classical actions of testosterone: an update. Trends Endocrinol Metab. 2007, 18 (10): 371-378. 10.1016/j.tem.2007.09.004View ArticlePubMedGoogle Scholar
- Foradori CD, Weiser MJ, Handa RJ: Non-genomic actions of androgens. Front Neuroendocrinol. 2008, 29 (2): 169-181.PubMed CentralView ArticlePubMedGoogle Scholar
- Freeman MR, Cinar B, Lu ML: Membrane rafts as potential sites of nongenomic hormonal signaling in prostate cancer. Trends Endocrinol Metab. 2005, 16: 273-279. 10.1016/j.tem.2005.06.002View ArticlePubMedGoogle Scholar
- Lieberherr M, Grosse B: Androgens increase intracellular calcium concentration and inositol 1, 4, 5-trisphosphate and diacylglycerol formation via a pertussis toxin-sensitive G-protein. J Biol Chem. 1994, 269: 7217-7223.PubMedGoogle Scholar
- Sun YH, Gao X, Tang YJ, Xu CL, Wang LH: Androgens induce increases in intracellular calcium via a G protein-coupled receptor in LNCaP prostate cancer cells. J Androl. 2006, 27: 671-678. 10.2164/jandrol.106.000554View ArticlePubMedGoogle Scholar
- Kampa M, Papakonstanti EA, Hatzoglou A, Stathopoulos EN, Stournaras C, Castanas E: The human prostate cancer cell line LNCaP bears functional membrane testosterone receptors that increase PSA secretion and modify actin cytoskeleton. Faseb J. 2002, 16: 1429-1431.PubMedGoogle Scholar
- Papakonstanti EA, Kampa M, Castanas E, Stournaras C: A rapid, nongenomic, signaling pathway regulates the actin reorganization induced by activation of membrane testosterone receptors. Mol Endocrinol. 2003, 17: 870-881. 10.1210/me.2002-0253View ArticlePubMedGoogle Scholar
- Benten WP, Lieberherr M, Stamm O, Wrehlke C, Guo Z, Wunderlich F: Testosterone signaling through internalizable surface receptors in androgen receptor-free macrophages. Mol Biol Cell. 1999, 10: 3113-3123.PubMed CentralView ArticlePubMedGoogle Scholar
- Benten WP, Lieberherr M, Giese G, Wrehlke C, Stamm O, Sekeris CE, Mossmann H, Wunderlich F: Functional testosterone receptors in plasma membranes of T cells. Faseb J. 1999, 13: 123-133.PubMedGoogle Scholar
- Kampa M, Nifli AP, Charalampopoulos I, Alexaki VI, Theodoropoulos PA, Stathopoulos EN, Gravanis A, Castanas E: Opposing effects of estradiol- and testosterone-membrane binding sites on T47D breast cancer cell apoptosis. Exp Cell Res. 2005, 307: 41-51. 10.1016/j.yexcr.2005.02.027View ArticlePubMedGoogle Scholar
- Kallergi G, Agelaki S, Markomanolaki H, Georgoulias V, Stournaras C: Activation of FAK/PI3K/Rac1 signaling controls actin reorganization and inhibits cell motility in human cancer cells. Cell Physiol Biochem. 2007, 20: 977-986. 10.1159/000110458View ArticlePubMedGoogle Scholar
- Hatzoglou A, Kampa M, Kogia C, Charalampopoulos I, Theodoropoulos PA, Anezinis P, Dambaki C, Papakonstanti EA, Stathopoulos EN, Stournaras C: Membrane androgen receptor activation induces apoptotic regression of human prostate cancer cells in vitro and in vivo. J Clin Endocrinol Metab. 2005, 90: 893-903. 10.1210/jc.2004-0801View ArticlePubMedGoogle Scholar
- Gatson JW, Kaur P, Singh M: Dihydrotestosterone differentially modulates the mitogen-activated protein kinase and the phosphoinositide 3-kinase/Akt pathways through the nuclear and novel membrane androgen receptor in C6 cells. Endocrinology. 2006, 147: 2028-2034. 10.1210/en.2005-1395View ArticlePubMedGoogle Scholar
- Alexaki VI, Charalampopoulos I, Kampa M, Nifli AP, Hatzoglou A, Gravanis A, Castanas E: Activation of membrane estrogen receptors induce pro-survival kinases. J Steroid Biochem Mol Biol. 2006, 98: 97-110. 10.1016/j.jsbmb.2005.08.017View ArticlePubMedGoogle Scholar
- Somjen D, Kohen F, Gayer B, Kulik T, Knoll E, Stern N: Role of putative membrane receptors in the effect of androgens on human vascular cell growth. J Endocrinol. 2004, 180: 97-106. 10.1677/joe.0.1800097View ArticlePubMedGoogle Scholar
- Papadopoulou N, Charalampopoulos I, Alevizopoulos K, Gravanis A, Stournaras C: Rho/ROCK/Actin signaling regualtes membrane androgen receptor induced apoptosis in prostate cancer cells. Exp Cell Res. 2008, 3162-3174. 314.Google Scholar
- Kampa M, Kogia C, Theodoropoulos PA, Anezinis P, Charalampopoulos I, Papakonstanti EA, Stathopoulos EN, Hatzoglou A, Stournaras C, Gravanis A, Castanas E: Activation of membrane androgen receptors potentiates the antiproliferative effects of paclitaxel on human prostate cancer cells. Mol Cancer Ther. 2006, 5: 1342-1351. 10.1158/1535-7163.MCT-05-0527View ArticlePubMedGoogle Scholar
- Alimirah F, Chen J, Basrawala Z, Xin H, Choubey D: DU-145 and PC-3 human prostate cancer cell lines express androgen receptor: implications for the androgen receptor functions and regulation. FEBS Lett. 2006, 580: 2294-2300. 10.1016/j.febslet.2006.03.041View ArticlePubMedGoogle Scholar
- Stone KR, Mickey DD, Wunderli H, Mickey GH, Paulson DF: Isolation of a human prostate carcinoma cell line (DU 145). Int J Cancer. 1978, 21: 274-281. 10.1002/ijc.2910210305View ArticlePubMedGoogle Scholar
- Mitchell S, Abel P, Ware M, Stamp G, Lalani E: Phenotypic and genotypic characterization of commonly used human prostatic cell lines. BJU Int. 2000, 85 (7): 932-944. 10.1046/j.1464-410x.2000.00606.xView ArticlePubMedGoogle Scholar
- Papakonstanti EA, Stournaras C: Actin cytoskeleton architecture and signaling in osmosensing. Methods Enzymol. 2007, 428: 227-240. 10.1016/S0076-6879(07)28012-7View ArticlePubMedGoogle Scholar
- Cairns P, Okami K, Halachmi S, Halachmi N, Esteller M, Herman JG, Jen J, Isaacs WB, Bova GS, Sidransky D: Frequent inactivation of PTEN/MMAC1 in primary prostate cancer. Cancer Res. 1997, 57: 4997-5000.PubMedGoogle Scholar
- Pesche S, Latil A, Muzeau F, Cussenot O, Fournier G, Longy M, Eng C, Lidereau R: PTEN/MMAC1/TEP1 involvement in primary prostate cancers. Oncogene. 1998, 16: 2879-2883. 10.1038/sj.onc.1202081View ArticlePubMedGoogle Scholar
- Feilotter HE, Nagai MA, Boag AH, Eng C, Mulligan LM: Analysis of PTEN and the 10q23 region in primary prostate carcinomas. Oncogene. 1998, 16: 1743-1748. 10.1038/sj.onc.1200205View ArticlePubMedGoogle Scholar
- Vivanco I, Sawyers CL: The phosphatidylinositol 3-Kinase AKT pathway in human cancer. Nat Rev Cancer. 2002, 2: 489-501. 10.1038/nrc839View ArticlePubMedGoogle Scholar
- Cardone MH, Roy N, Stennicke HR, Salvesen GS, Franke TF, Stanbridge E, Frisch S, Reed JC: Regulation of cell death protease caspase-9 by phosphorylation. Science. 1998, 282: 1318-1321. 10.1126/science.282.5392.1318View ArticlePubMedGoogle Scholar
- Datta SR, Dudek H, Tao X, Masters S, Fu H, Gotoh Y, Greenberg ME: Akt phosphorylation of BAD couples survival signals to the cell-intrinsic death machinery. Cell. 1997, 91: 231-241. 10.1016/S0092-8674(00)80405-5View ArticlePubMedGoogle Scholar
- Alessi DR, James SR, Downes CP, Holmes AB, Gaffney PR, Reese CB, Cohen P: Characterization of a 3-phosphoinositide-dependent protein kinase which phosphorylates and activates protein kinase Balpha. Curr Biol. 1997, 7: 261-269. 10.1016/S0960-9822(06)00122-9View ArticlePubMedGoogle Scholar
- Stokoe D, Stephens LR, Copeland T, Gaffney PR, Reese CB, Painter GF, Holmes AB, McCormick F, Hawkins PT: Dual role of phosphatidylinositol-3, 4, 5-trisphosphate in the activation of protein kinase B. Science. 1997, 277: 567-570. 10.1126/science.277.5325.567View ArticlePubMedGoogle Scholar
- Vanhaesebroeck B, Alessi DR: The PI3K-PDK1 connection: more than just a road to PKB. Biochem J. 2000, 346 (Pt 3): 561-576. 10.1042/0264-6021:3460561PubMed CentralPubMedGoogle Scholar
- Zha J, Harada H, Yang E, Jockel J, Korsmeyer SJ: Serine phosphorylation of death agonist BAD in response to survival factor results in binding to 14-3-3 not BCL-X(L). Cell. 1996, 87: 619-628. 10.1016/S0092-8674(00)81382-3View ArticlePubMedGoogle Scholar
- Hayden MS, Ghosh S: Signaling to NF-kappaB. Genes Dev. 2004, 18: 2195-2224. 10.1101/gad.1228704View ArticlePubMedGoogle Scholar
- Chen ZJ, Parent L, Maniatis T: Site-specific phosphorylation of IkappaBalpha by a novel ubiquitination-dependent protein kinase activity. Cell. 1996, 84: 853-862. 10.1016/S0092-8674(00)81064-8View ArticlePubMedGoogle Scholar
- Hatakeyama S, Kitagawa M, Nakayama K, Shirane M, Matsumoto M, Hattori K, Higashi H, Nakano H, Okumura K, Onoe K, Good RA: Ubiquitin-dependent degradation of IkappaBalpha is mediated by a ubiquitin ligase Skp1/Cul 1/F-box protein FWD1. Proc Natl Acad Sci USA. 1999, 96: 3859-3863. 10.1073/pnas.96.7.3859PubMed CentralView ArticlePubMedGoogle Scholar
- Spencer E, Jiang J, Chen ZJ: Signal-induced ubiquitination of IkappaBalpha by the F-box protein Slimb/beta-TrCP. Genes Dev. 1999, 13: 284-294. 10.1101/gad.13.3.284PubMed CentralView ArticlePubMedGoogle Scholar
- Ghosh S, Karin M: Missing pieces in the NF-kappaB puzzle. Cell. 2002, 109 (Suppl): S81-96. 10.1016/S0092-8674(02)00703-1View ArticlePubMedGoogle Scholar
- Li Q, Verma IM: NF-kappaB regulation in the immune system. Nat Rev Immunol. 2002, 2: 725-734. 10.1038/nri910View ArticlePubMedGoogle Scholar
- Rokhlin OW, Bishop GA, Hostager BS, Waldschmidt TJ, Sidorenko SP, Pavloff N, Kiefer MC, Umansky SR, Glover RA, Cohen MB: Fas-mediated apoptosis in human prostatic carcinoma cell lines. Cancer Res. 1997, 57 (9): 1758-68.PubMedGoogle Scholar
- Caraglia M, Marra M, Leonetti C, Meo G, D'Alessandro AM, Baldi A, Santini D, Tonini G, Bertieri R, Zupi G, Budillon A, Abbruzzese A: R115777 (Zarnestra)/Zoledronic acid (Zometa) cooperation on inhibition of prostate cancer proliferation is paralleled by Erk/Akt inactivation and reduced Bcl-2 and Bad phosphorylation. J Cell Physiol. 2007, 211 (2): 533-43. 10.1002/jcp.20960View ArticlePubMedGoogle Scholar
- Papadopoulou N, Papakonstanti E, Kallergi G, Alevizopoulos K, Stournaras C: Membrane androgen receptor activation in tumor cells: Molecular signaling and clinical impact. IUBMB life. 2008.Google Scholar
- Gourlay CW, Ayscough KR: The actin cytoskeleton: a key regulator of apoptosis and ageing?. Nat Rev Mol Cell Biol. 2005, 6: 583-589. 10.1038/nrm1682View ArticlePubMedGoogle Scholar
- Franklin-Tong VE, Gourlay CW: A role for actin in regulating apoptosis/programmed cell death: evidence spanning yeast, plants and animals. Biochem J. 2008, 413 (3): 389-404. 10.1042/BJ20080320View ArticlePubMedGoogle Scholar
- Papakonstanti EA, Stournaras C: Cell responses regulated by early reorganization of actin cytoskeleton. FEBS Lett. 2008, 582: 2120-2127. 10.1016/j.febslet.2008.02.064View ArticlePubMedGoogle Scholar
- Stathopoulos EN, Dambaki C, Kampa M, Theodoropoulos PA, Anezinis P, Delakas D, Delides GS, Castanas E: Membrane androgen binding sites are preferentially expressed in human prostate carcinoma cells. BMC Clin Pathol. 2003, 3: 1- 10.1186/1472-6890-3-1PubMed CentralView ArticlePubMedGoogle Scholar
- Dambaki C, Kogia C, Kampa M, Darivianaki K, Nomikos M, Anezinis P, Theodoropoulos PA, Castanas E, Stathopoulos EN: Membrane testosterone binding sites in prostate carcinoma as a potential new marker and therapeutic target: study in paraffin tissue sections. BMC Cancer. 2005, 5: 148- 10.1186/1471-2407-5-148PubMed CentralView ArticlePubMedGoogle Scholar
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