Susan K Logan, Ph.D.

Androgen receptor levels are upregulated by Akt in prostate cancer
Ha, Susan; Ruoff, Rachel; Kahoud, Nicole; Franke, Thomas F; Logan, Susan K
2011 Apr;18(2):245-255, Endocrine-related cancer
Multiple lines of evidence suggest a functional link between the androgen receptor (AR) and the serine/threonine kinase Akt in the development and progression of prostate cancer. To investigate the impact of Akt activity on AR homeostasis, we treated androgen-dependent LNCaP and LAPC-4 prostate cancer cells with Akt inhibitor. Akt inhibition decreased AR expression, suggesting that Akt activity was required for regulation of AR protein levels. However, while androgen-independent LNCaP-abl cells also showed diminished AR protein levels in response to Akt inhibition, treatment of androgen-independent LNCaP-AI cells failed to alter AR protein levels upon similar treatment, suggesting that AR protein levels in these androgen-independent prostate cells were regulated by mechanisms independent of Akt activation. Regulation of AR, downstream of activated Akt, also was observed in vivo when examining transgenic mice that overexpress constitutively active mutant myristoylated (myr)-Akt1 in the prostate. Transgenic mice expressing activated myr-Akt1 exhibited higher levels of AR mRNA and protein. Expression of activated myr-Akt1 did not alter prostate cell growth and no significant size differences between prostate tissues derived from transgenic animals were observed when comparing transgenic mice with wild-type mice. Still, transgenic mice overexpressing Akt exhibited higher levels of gammaH2AX and phosphorylated Chk2 in prostate tissue. These changes in markers associated with oncogene-induced senescence confirmed significant altered signaling in the transgenic mouse model. Overall, results presented here suggest that AR levels are regulated by the Akt pathway
— id: 136566, year: 2011, vol: 18, page: 245, stat: Journal Article,

Regulation of Androgen Receptor-Mediated Transcription by RPB5 Binding Protein URI/RMP
Mita, Paolo; Savas, Jeffrey N; Djouder, Nabil; Yates, John R 3rd; Ha, Susan; Ruoff, Rachel; Schafler, Eric D; Nwachukwu, Jerome C; Tanese, Naoko; Cowan, Nicholas J; Zavadil, Jiri; Garabedian, Michael J; Logan, Susan K
2011 Sep;31(17):3639-3652, Molecular & cellular biology
Androgen receptor (AR)-mediated transcription is modulated by interaction with coregulatory proteins. We demonstrate that the unconventional prefoldin RPB5 interactor (URI) is a new regulator of AR transcription and is critical for antagonist (bicalutamide) action. URI is phosphorylated upon androgen treatment, suggesting communication between the URI and AR signaling pathways. Whereas depletion of URI enhances AR-mediated gene transcription, overexpression of URI suppresses AR transcriptional activation and anchorage-independent prostate cancer cell growth. Repression of AR-mediated transcription is achieved, in part, by URI binding and regulation of androgen receptor trapped clone 27 (Art-27), a previously characterized AR corepressor. Consistent with this idea, genome-wide expression profiling in prostate cancer cells upon depletion of URI or Art-27 reveals substantially overlapping patterns of gene expression. Further, depletion of URI increases the expression of the AR target gene NKX-3.1, decreases the recruitment of Art-27, and increases AR occupancy at the NKX-3.1 promoter. While Art-27 can bind AR directly, URI is bound to chromatin prior to hormone-dependent recruitment of AR, suggesting a role for URI in modulating AR recruitment to target genes
— id: 136514, year: 2011, vol: 31, page: 3639, stat: Journal Article,

LEF1 Identifies Androgen-Independent Epithelium in the Developing Prostate
Wu, Xinyu; Daniels, Garrett; Shapiro, Ellen; Xu, Kun; Huang, Hongying; Li, Yirong; Logan, Susan; Greco, M Alba; Peng, Yi; Monaco, Marie E; Melamed, Jonathan; Lepor, Herbert; Grishina, Irina; Lee, Peng
2011 Jun;25(6):1018-1026, Molecular endocrinology
Lymphoid enhancer-binding factor (LEF)1 is a major mediator and a target in canonical Wnt/beta-catenin pathway. Interactions between the androgen receptor (AR) and canonical Wnt pathways have been implicated in the development of the genitourinary organs. Here, we investigated the localization and role of LEF1-positive cells during development of the prostate gland in human and in the murine model. We show that during human prostate development, LEF1 is restricted to the basal epithelial layer of the urogenital sinus. During mouse development, Lef1 is also present in the urogenital mesenchyme in addition to the basal epithelial layer of the urogenital sinus. In the course of elongation and branching of the prostatic ducts, Lef1 is localized to the proliferating epithelium at the distal tips of the buds. Notably, during branching morphogenesis, domains of Lef1 and AR are mutually exclusive. We further employed the TOPGAL reporter strain to examine the dynamics of Wnt signaling in the context of prostate regression upon a 7-d treatment with a competitive AR inhibitor, bicalutamide. We found that Wnt/Lef1-positive basal cells are not dependent upon androgen for survival. Furthermore, upon bicalutamide treatment, Wnt/Lef1-positive basal progenitors repopulated the luminal compartment. We conclude that Wnt/Lef1 activity identifies an androgen-independent population of prostate progenitors, which is important for embryonic development and organ maintenance and regeneration in the adult
— id: 132604, year: 2011, vol: 25, page: 1018, stat: Journal Article,

Glucocorticoid receptor DNA binding decoy is a gas
Garabedian, Michael J; Logan, Susan K
2010 ;3(108):pe5-pe5, Science signaling
The glucocorticoid receptor (GR) is a paradigmatic DNA binding transcription factor and was described over 20 years ago as one of the first proteins identified to bind the enhancer region of genes called 'response elements.' Since that time, an immense amount of work has revealed that GR transcriptional regulation is controlled at virtually every step of its activity: ligand binding, nuclear translocation, transcriptional cofactor binding, and DNA binding. Just when the major modes of GR regulation appear known, a new study provides yet another mechanism whereby GR transcriptional activity is controlled under conditions of cell growth arrest. In this case, GR activity is repressed by a small noncoding RNA (ncRNA) from the growth arrest-specific transcript 5 gene that folds into a soluble glucocorticoid response element-like sequence and serves as a decoy for GR DNA binding. This unexpected mode of regulation by nucleic acid molecular mimicry is probably not confined to GR and should spark interest in the hunt for other ncRNAs that regulate transcription factor binding to DNA
— id: 106599, year: 2010, vol: 3, page: pe5, stat: Journal Article,

Higher Expression of Serine-213 Phosphorylated Androgen Receptor Level Is Associated With Prostate Cancer Recurrence
Jain, Shilpa; Ruoff, Rachael; Ha, Susan; Melamed, Jonathan; Wang, Jinhua; Ren, Qinghu; Lee, Peng; Logan, Susan
2010 OCT ;134(4):674-674, American journal of clinical pathology
— id: 113734, year: 2010, vol: 134, page: 674, stat: Journal Article,

High levels of Hsp90 cochaperone p23 promote tumor progression and poor prognosis in breast cancer by increasing lymph node metastases and drug resistance
Simpson, Natalie E; Lambert, W Marcus; Watkins, Renecia; Giashuddin, Shah; Huang, S Joseph; Oxelmark, Ellinor; Arju, Rezina; Hochman, Tsivia; Goldberg, Judith D; Schneider, Robert J; Reiz, Luiz Fernando Lima; Soares, Fernando Augusto; Logan, Susan K; Garabedian, Michael J
2010 Nov 1;70(21):8446-8456, Cancer research
p23 is a heat shock protein 90 (Hsp90) cochaperone located in both the cytoplasm and nucleus that stabilizes unliganded steroid receptors, controls the catalytic activity of certain kinases, regulates protein-DNA dynamics, and is upregulated in several cancers. We had previously shown that p23-overexpressing MCF-7 cells (MCF-7+p23) exhibit increased invasion without affecting the estrogen-dependent proliferative response, which suggests that p23 differentially regulates genes controlling processes linked to breast tumor metastasis. To gain a comprehensive view of the effects of p23 on estrogen receptor (ER)-dependent and -independent gene expression, we profiled mRNA expression from control versus MCF-7+p23 cells in the absence and presence of estrogen. A number of p23-sensitive target genes involved in metastasis and drug resistance were identified. Most striking is that many of these genes are also misregulated in invasive breast cancers, including PMP22, ABCC3, AGR2, Sox3, TM4SF1, and p8 (NUPR1). Upregulation of the ATP-dependent transporter ABCC3 by p23 conferred resistance to the chemotherapeutic agents etoposide and doxorubicin in MCF-7+p23 cells. MCF-7+p23 cells also displayed higher levels of activated Akt and an expanded phosphoproteome relative to control cells, suggesting that elevated p23 also enhances cytoplasmic signaling pathways. For breast cancer patients, tumor stage together with high cytoplasmic p23 expression more accurately predicted disease recurrence and mortality than did stage alone. High nuclear p23 was found to be associated with high cytoplasmic p23, therefore both may promote tumor progression and poor prognosis by increasing metastatic potential and drug resistance in breast cancer patients
— id: 114177, year: 2010, vol: 70, page: 8446, stat: Journal Article,

Sin3B expression is required for cellular senescence and is up-regulated upon oncogenic stress
Grandinetti, Kathryn B; Jelinic, Petar; DiMauro, Teresa; Pellegrino, Jessica; Fernandez Rodriguez, Ruben; Finnerty, Patricia M; Ruoff, Rachel; Bardeesy, Nabeel; Logan, Susan K; David, Gregory
2009 Aug 15;69(16):6430-6437, Cancer research
Serial passage of primary mammalian cells or strong mitogenic signals induce a permanent exit from the cell cycle called senescence. A characteristic of senescent cells is the heterochromatinization of loci encoding pro-proliferative genes, leading to their transcriptional silencing. Senescence is thought to represent a defense mechanism against uncontrolled proliferation and cancer. Consequently, genetic alterations that allow senescence bypass are associated with susceptibility to oncogenic transformation. We show that fibroblasts genetically inactivated for the chromatin-associated Sin3B protein are refractory to replicative and oncogene-induced senescence. Conversely, overexpression of Sin3B triggers senescence and the formation of senescence-associated heterochromatic foci. Although Sin3B is strongly up-regulated upon oncogenic stress, decrease in expression of Sin3B is associated with tumor progression in vivo, suggesting that expression of Sin3B may represent a barrier against transformation. Together, these results underscore the contribution of senescence in tumor suppression and suggest that expression of chromatin modifiers is modulated at specific stages of cellular transformation. Consequently, these findings suggest that modulation of Sin3B-associated activities may represent new therapeutic opportunities for treatment of cancers
— id: 101640, year: 2009, vol: 69, page: 6430, stat: Journal Article,

Genome-wide impact of androgen receptor trapped clone-27 loss on androgen-regulated transcription in prostate cancer cells
Nwachukwu, Jerome C; Mita, Paolo; Ruoff, Rachel; Ha, Susan; Wang, Qianben; Huang, S Joseph; Taneja, Samir S; Brown, Myles; Gerald, William L; Garabedian, Michael J; Logan, Susan K
2009 Apr 1;69(7):3140-3147, Cancer research
The androgen receptor (AR) directs diverse biological processes through interaction with coregulators such as AR trapped clone-27 (ART-27). Our results show that ART-27 is recruited to AR-binding sites by chromatin immunoprecipitation analysis. In addition, the effect of ART-27 on genome-wide transcription was examined. The studies indicate that loss of ART-27 enhances expression of many androgen-regulated genes, suggesting that ART-27 inhibits gene expression. Surprisingly, classes of genes that are up-regulated upon ART-27 depletion include regulators of DNA damage checkpoint and cell cycle progression, suggesting that ART-27 functions to keep expression levels of these genes low. Consistent with this idea, stable reduction of ART-27 by short-hairpin RNA enhances LNCaP cell proliferation compared with control cells. The effect of ART-27 loss was also examined in response to the antiandrogen bicalutamide. Unexpectedly, cells treated with ART-27 siRNA no longer exhibited gene repression in response to bicalutamide. To examine ART-27 loss in prostate cancer progression, immunohistochemistry was conducted on a tissue array containing samples from primary tumors of individuals who were clinically followed and later shown to have either recurrent or nonrecurrent disease. Comparison of ART-27 and AR staining indicated that nuclear ART-27 expression was lost in the majority of AR-positive recurrent prostate cancers. Our studies show that reduction of ART-27 protein levels in prostate cancer may facilitate antiandrogen-resistant disease
— id: 99292, year: 2009, vol: 69, page: 3140, stat: Journal Article,

Development of phosphorylation site-specific antibodies to nuclear receptors
Torra, Ines Pineda; Staverosky, Julia A; Ha, Susan; Logan, Susan K; Garabedian, Michael J
2009 ;505:221-235, Methods in molecular biology
Protein phosphorylation is a versatile posttranslational modification that can regulate nuclear receptor function. Although the precise role of receptor phosphorylation is not fully understood, it appears that it functions to direct or refine receptor activity in response to particular physiological requirements. Identifying and characterizing specific nuclear receptor phosphorylation sites is an important step in elucidating the role(s) receptor phosphorylation plays in function. Although traditional methods of metabolic labeling and in vitro protein phosphorylation have been informative, receptor phosphorylation site-specific antibodies are simple and reliable tools to study receptor phosphorylation. This chapter will discuss how to develop nuclear receptor phosphorylation site-specific antibodies to elucidate function
— id: 92774, year: 2009, vol: 505, page: 221, stat: Journal Article,

Glucocorticoid receptor phosphorylation differentially affects target gene expression
Chen, Weiwei; Dang, Thoa; Blind, Raymond D; Wang, Zhen; Cavasotto, Claudio N; Hittelman, Adam B; Rogatsky, Inez; Logan, Susan K; Garabedian, Michael J
2008 Aug;22(8):1754-1766, Molecular endocrinology
The glucocorticoid receptor (GR) is phosphorylated at multiple sites within its N terminus (S203, S211, S226), yet the role of phosphorylation in receptor function is not understood. Using a range of agonists and GR phosphorylation site-specific antibodies, we demonstrated that GR transcriptional activation is greatest when the relative phosphorylation of S211 exceeds that of S226. Consistent with this finding, a replacement of S226 with an alanine enhances GR transcriptional response. Using a battery of compounds that perturb different signaling pathways, we found that BAPTA-AM, a chelator of intracellular divalent cations, and curcumin, a natural product with antiinflammatory properties, reduced hormone-dependent phosphorylation at S211. This change in GR phosphorylation was associated with its decreased nuclear retention and transcriptional activation. Molecular modeling suggests that GR S211 phosphorylation promotes a conformational change, which exposes a novel surface potentially facilitating cofactor interaction. Indeed, S211 phosphorylation enhances GR interaction with MED14 (vitamin D receptor interacting protein 150). Interestingly, in U2OS cells expressing a nonphosphorylated GR mutant S211A, the expression of IGF-binding protein 1 and interferon regulatory factor 8, both MED14-dependent GR target genes, was reduced relative to cells expressing wild-type receptor across a broad range of hormone concentrations. In contrast, the induction of glucocorticoid-induced leucine zipper, a MED14-independent GR target, was similar in S211A- and wild-type GR-expressing cells at high hormone levels, but was reduced in S211A cells at low hormone concentrations, suggesting a link between GR phosphorylation, MED14 involvement, and receptor occupancy. Phosphorylation also affected the magnitude of repression by GR in a gene-selective manner. Thus, GR phosphorylation at S211 and S226 determines GR transcriptional response by modifying cofactor interaction. Furthermore, the effect of GR S211 phosphorylation is gene specific and, in some cases, dependent upon the amount of activated receptor
— id: 80349, year: 2008, vol: 22, page: 1754, stat: Journal Article,

Atypical regulation of SRC-3
Garabedian, Michael J; Logan, Susan K
2008 Jul;33(7):301-304, Trends in biochemical sciences
Overexpression of steroid receptor coactivator 3 (SRC-3) is associated with an increased incidence of breast cancer. A recent study shows that SRC-3 is protected from proteasomal degradation by atypical protein kinase C (aPKC)-mediated phosphorylation in an estrogen receptor alpha (ERalpha)-dependent manner. This finding provides a novel mechanism for coupling increased SRC-3 expression with enhanced estrogen-dependent cellular proliferation
— id: 80347, year: 2008, vol: 33, page: 301, stat: Journal Article,

Down-regulation of p57Kip2 induces prostate cancer in the mouse
Jin, Ren Jie; Lho, Yongsoo; Wang, Yongqing; Ao, Mingfang; Revelo, Monica Patricia; Hayward, Simon W; Wills, Marcia L; Logan, Susan K; Zhang, Pumin; Matusik, Robert J
2008 May 15;68(10):3601-3608, Cancer research
p57(Kip2) has been considered a candidate tumor suppressor gene because of its location in the genome, biochemical activities, and imprinting status. However, little is known about the role of p57(Kip2) in tumorigenesis and cancer progression. Here, we show that the expression of p57(Kip2) is significantly decreased in human prostate cancer, and the overexpression of p57(Kip2) in prostate cancer cells significantly suppressed cell proliferation and reduced invasive ability. In addition, overexpression of p57(Kip2) in LNCaP cells inhibited tumor formation in nude mice, resulting in well-differentiated squamous tumors rather than adenocarcinoma. Furthermore, the prostates of p57(Kip2) knockout mice developed prostatic intraepithelial neoplasia and adenocarcinoma. Remarkably, this mouse prostate cancer is pathologically identical to human prostate adenocarcinoma. Therefore, these results strongly suggest that p57(Kip2) is an important gene in prostate cancer tumorigenesis, and the p57(Kip2) pathway may be a potential target for prostate cancer prevention and therapy
— id: 80348, year: 2008, vol: 68, page: 3601, stat: Journal Article,

Regulation of prostate cell growth through androgen receptor cofactors
Logan, SK; Nwachukwu, JC; Mita, P; Taneja, SS; Garabedian, MJ
2008 ;179(4):190-190, Journal of urology
— id: 104579, year: 2008, vol: 179, page: 190, stat: Journal Article,

The Heterochromatin Protein 1 Family is Regulated in Prostate Development and Cancer
Shapiro, Ellen; Huang, Hongying; Ruoff, Rachel; Lee, Peng; Tanese, Naoko; Logan, Susan K
2008 Apr 22;179(6):2435-2439, Journal of urology
PURPOSE: The HP1 family of evolutionarily conserved proteins regulates heterochromatin packaging, in addition to a less defined role in the regulation of euchromatic genes. To examine the possible role of HP1 proteins in fetal prostate development and prostate cancer the protein expression of HP1alpha, beta and gamma was evaluated in human archival tissue. MATERIALS AND METHODS: Tissue sections from human prostate cancer and fetal prostate were examined using antibodies against HP1 isoforms to evaluate HP1 modulation in cancer and development. Western blot analysis of HP1 proteins was also performed in extracts of cultured prostate cancer cells. RESULTS: HP1alpha, beta and gamma are differentially regulated in various cellular compartments in prostate development. HP1alpha is not expressed at 14 or 24 weeks of prostate development but it is expressed in adult prostate tissue. HP1beta is highly expressed at 14 and 24 weeks, and it appears predominantly in epithelial cells compared to HP1gamma, which is expressed at equal levels in epithelial and stromal cells. All 3 HP1 isoforms show altered expression in prostate cancer compared to that in normal adult prostate tissue. CONCLUSIONS: HP1 proteins are tightly regulated during prostate development. In the adult prostate HP1alpha, beta and gamma antibodies detect high levels of HP1 antigen in a contiguous layer of epithelial cells. However, the detection of HP1 in prostate cancer ranges from undetectable to inconsistent staining of noncontiguous epithelial cells
— id: 78573, year: 2008, vol: 179, page: 2435, stat: Journal Article,

Transcriptional regulation of the androgen receptor cofactor androgen receptor trapped clone-27
Nwachukwu, Jerome C; Li, Wenhui; Pineda-Torra, Ines; Huang, Hong Ying; Ruoff, Rachel; Shapiro, Ellen; Taneja, Samir S; Logan, Susan K; Garabedian, Michael J
2007 Dec;21(12):2864-2876, Molecular endocrinology
Cofactors modulate nuclear receptor activity and impact human health and disease, yet surprisingly little is known about their transcriptional regulation. Androgen receptor trapped clone-27 (ART-27) is a cofactor that binds to androgen receptor (AR) amino terminus and modulates AR-dependent transcription. Interestingly, ART-27 displays both a cell type- and developmental stage-specific expression pattern. However, the cis-acting elements and trans-acting factors affecting ART-27 gene expression have not been elucidated. We found that ART-27 gene expression is repressed and its promoter is histone H3-K27 tri-methylated in human embryonic kidney cells, but not prostate cells, and the histone deacetylase inhibitor, trichostatin A, relieves this inhibition. The DNA response elements that control the induction of ART-27 gene expression were also characterized. The major cis-acting element corresponds to a consensus cAMP-responsive element (CRE) and binds the CRE-binding protein (CREB) as shown by EMSA and chromatin immunoprecipitation assays. Furthermore, ART-27 promoter activity is induced upon CREB overexpression. Epidermal growth factor, which activates CREB via phosphorylation, also induces ART-27 expression, whereas a reduction in CREB phosphorylation or expression blocks this induction in prostate cells. In human prostate development, both epithelial and stromal cells express CREB; however, active phosphorylated CREB is restricted to epithelial cells where ART-27 is expressed. Based on these findings, we propose a transcriptional regulatory circuit for the developmental expression of ART-27 that includes repression by chromatin modification through a trichostatin A-sensitive factor and activation upon growth factor stimulation via CREB
— id: 94948, year: 2007, vol: 21, page: 2864, stat: Journal Article,

ART-27: A novel coactivator with tumor suppressor function in the prostate
Feig, JL; Ha, S; Ruoff, R; Logan, SK
2006 MAR 26 ;231(4):658-658, Abstracts of papers (American Chemical Society)
— id: 68761, year: 2006, vol: 231, page: 658, stat: Journal Article,

EXPRESSION AND REGULATION OF GLUCOCORTICOID RECEPTOR IN HUMAN PLACENTAL VILLOUS FIBROBLASTS
Lee, Men-Jean; Wang, Zhen; Yee, Herman; Ma, Yuehong; Swenson, Nicole; Yang, Liubin; Kadner, Susan S; Baergen, Rebecca N; Logan, Susan K; Garabedian, Michael J; Guller, Seth
2005 Nov;146(11):4619-4626, Endocrinology
The human placenta is a glucocorticoid (GC)-responsive organ consisting of multiple cell types including smooth muscle cells, fibroblasts, and trophoblast that demonstrate changes in gene expression following hormone treatment. However, little is known about the relative expression or activity of the GC receptor (GR) among the various placental cell types. Normal term human placentas were examined by immunohistochemistry using either GR phosphorylation site-specific antibodies that are markers for various activation states of the GR, or a GR antibody that recognizes the receptor independent of its phosphorylation state (total GR). We found strong total GR and phospho-GR immunoreactivity in stromal fibroblasts of terminal villi, as well as perivascular fibroblasts and vascular smooth muscle cells of the stem villi. Lower levels of both total GR and phospho-GR were found within cytotrophoblast cells relative to fibroblasts, whereas syncytiotrophoblast showed very little total GR or phospho-GR immunoreactivity. This pattern holds true by immunoblot analysis of extracts from cell fractions cultured ex vivo. In cultured placental fibroblasts, phosphorylation of GR increased upon short-term GC treatment, consistent with a role for GR phosphorylation in receptor transactivation. Total GR levels were reduced by nearly 90% following long-term hormone treatment; however, this down-regulation was independent of changes in GR mRNA levels. These findings demonstrate that GR levels in fibroblasts can be modulated by changes in hormone exposure. Such cell type-specific differences in GR protein expression and phosphorylation may provide the means of differentially regulating the GC response among the cells of the human placenta
— id: 57670, year: 2005, vol: 146, page: 4619, stat: Journal Article,

Androgen receptor mutations identified in prostate cancer and androgen insensitivity syndrome display aberrant ART-27 coactivator function
Li, Wenhui; Cavasotto, Claudio N; Cardozo, Timothy; Ha, Susan; Dang, Thoa; Taneja, Samir S; Logan, Susan K; Garabedian, Michael J
2005 May 26;19(9):2273-2282, Molecular endocrinology
The transcriptional activity of the androgen receptor (AR) is modulated by interactions with coregulatory molecules. It has been proposed that aberrant interactions between AR and its coregulators may contribute to diseases related to AR activity, such as prostate cancer and androgen insensitivity syndrome (AIS); however, evidence linking abnormal receptor:cofactor interactions to disease is scant. The Androgen Receptor Trapped clone-27 (ART-27) is a recently identified AR N-terminal coactivator that is associated with AR-mediated growth inhibition. Here we analyze a number of naturally occurring AR mutations identified in prostate cancer and AIS for their ability to affect AR response to ART-27. Although the vast majority of AR mutations appeared capable of increased activation in response to ART-27, an AR mutation identified in prostate cancer (AR P340L) and AIS (AR E2K) show reduced transcriptional responses to ART-27, whereas their response to the p160 class of coactivators was not diminished. Relative to the wild-type receptor, less ART-27 protein associated with the AR E2K substitution, consistent with reduced transcriptional response. Surprisingly, more ART-27 associated with AR P340L, despite the fact that the mutation decreased transcriptional activation in response to ART-27. Our findings suggest that aberrant AR-coactivator association interferes with normal ART-27 coactivator function resulting in suppression of AR activity and may contribute to the pathogenesis of diseases related to alterations in AR activity, such as prostate cancer and AIS
— id: 56038, year: 2005, vol: 19, page: 2273, stat: Journal Article,

Cell-specific regulation of androgen receptor phosphorylation in vivo
Taneja, Samir S; Ha, Susan; Swenson, Nicole K; Huang, Hong Ying; Lee, Peng; Melamed, Jonathan; Shapiro, Ellen; Garabedian, Michael J; Logan, Susan K
2005 Dec 9;280(49):40916-40924, Journal of biological chemistry
The biological ramifications of phosphorylation of the androgen receptor (AR) are largely unknown. To examine the phosphorylation of AR at serine 213, a putative substrate for Akt, a phosphorylation site-specific antibody was generated. The use of this antibody indicated that AR Ser-213 is phosphorylated in vivo and that phosphorylation is tightly regulated in a cell type-specific manner. Furthermore, Ser-213 phosphorylation took place with rapid kinetics and was inhibited by the phosphatidylinositol 3-kinase inhibitor LY294002. Phosphorylation occurred in response to R1881 and dihydrotestosterone but weakly if at all in response to testosterone. It did not occur in response to AR antagonists or growth factor stimulation in the absence of an AR agonist. Transcription assays using an AR-responsive reporter gene construct showed that activated phosphatidylinositol 3-kinase inhibited transcription mediated by wild type AR but not that of a mutant AR variant (S213A), which could not be phosphorylated at Ser-213. By immunohistochemistry, the AR Ser(P)-213 antigen was detected in prostate epithelial but not stromal cells despite the fact that an antibody recognizing both phosphorylated and non-phosphorylated forms of AR demonstrates that AR is present in both cell types as expected. In fetal tissue the AR-Ser(P)-213 antigen was present in epithelial cells of the urogenital sinus when endogenous androgen levels were high and activated Akt was prevalent, but absent at a later stage of development when endogenous androgen levels were low and Akt activation was minimal. Immunoreactivity was evident in differentiated cells lining the lumen of the urogenital sinus but not in rapidly dividing, Ki67 positive cells within the developing prostate or stromal tissue, suggesting that site-specific phosphorylation of AR Ser-213 by cellular kinases occurs in a non-proliferating cellular milieu
— id: 61359, year: 2005, vol: 280, page: 40916, stat: Journal Article,

ART-27, an androgen receptor coactivator regulated in prostate development and cancer
Taneja, Samir S; Ha, Susan; Swenson, Nicole K; Torra, Ines Pineda; Rome, Serge; Walden, Paul D; Huang, Hong Ying; Shapiro, Ellen; Garabedian, Michael J; Logan, Susan K
2004 Apr 2;279(14):13944-13952, Journal of biological chemistry
Androgen receptor trapped clone-27 (ART-27) is a newly described transcriptional coactivator that binds to the N terminus of the androgen receptor (AR). Given the vital importance of AR signaling in prostate growth and differentiation, we investigated the role of ART-27 in these processes. Immunohistochemical studies indicate that ART-27 protein is expressed in differentiated epithelial cells of adult human prostate and breast tissue. In prostate, ART-27 is abundant in AR-positive prostate luminal epithelial cells, in contrast to the stroma, where cells express AR but not ART-27. The use of a rat model of androgen depletion/reconstitution indicates that ART-27 expression is associated with the elaboration of differentiated prostate epithelial cells. Interestingly, regulated expression of ART-27 in the androgen-sensitive LNCaP prostate cancer cell line inhibits androgen-mediated cellular proliferation and enhances androgen-mediated transcription of the prostate-specific antigen (PSA) gene. Consistent with a growth suppressive function, we show that ART-27 expression levels are negligible in human prostate cancer. Importantly, examination of ART-27 protein expression in early fetal prostate development demonstrates that ART-27 is detected only when the developing prostate gland has proceeded from a solid mass of undifferentiated cells to a stage in which differentiated luminal epithelial cells are evident. Thus, ART-27 is an AR cofactor shown to be subject to both cell type and developmental regulation in humans. Overall, the results suggest that decreased levels of ART-27 protein in prostate cancer tissue may occur as a result of de-differentiation, and indicate that ART-27 is likely to regulate a subset of AR-responsive genes important to prostate growth suppression and differentiation
— id: 44732, year: 2004, vol: 279, page: 13944, stat: Journal Article,

ART-27, a novel androgen receptor (AR) interacting protein, is a potential. mediator of prostate eptihelial differentiation
Logan, S; Ha, S; Rome, S; Garabedian, MJ; Taneja, SS
2002 ;167(4):56-56, Journal of urology
— id: 104583, year: 2002, vol: 167, page: 56, stat: Journal Article,

Identification and characterization of ART-27, a novel coactivator for the androgen receptor N terminus
Markus, Steven M; Taneja, Samir S; Logan, Susan K; Li, Wenhui; Ha, Susan; Hittelman, Adam B; Rogatsky, Inez; Garabedian, Michael J
2002 Feb;13(2):670-682, Molecular biology of the cell
The androgen receptor (AR) is a ligand-regulated transcription factor that stimulates cell growth and differentiation in androgen-responsive tissues. The AR N terminus contains two activation functions (AF-1a and AF-1b) that are necessary for maximal transcriptional enhancement by the receptor; however, the mechanisms and components regulating AR transcriptional activation are not fully understood. We sought to identify novel factors that interact with the AR N terminus from an androgen-stimulated human prostate cancer cell library using a yeast two-hybrid approach designed to identify proteins that interact with transcriptional activation domains. A 157-amino acid protein termed ART-27 was cloned and shown to interact predominantly with the AR(153-336), containing AF-1a and a part of AF-1b, localize to the nucleus and increase the transcriptional activity of AR when overexpressed in cultured mammalian cells. ART-27 also enhanced the transcriptional activation by AR(153-336) fused to the LexA DNA-binding domain but not other AR N-terminal subdomains, suggesting that ART-27 exerts its effect via an interaction with a defined region of the AR N terminus. ART-27 interacts with AR in nuclear extracts from LNCaP cells in a ligand-independent manner. Interestingly, velocity gradient sedimentation of HeLa nuclear extracts suggests that native ART-27 is part of a multiprotein complex. ART-27 is expressed in a variety of human tissues, including sites of androgen action such as prostate and skeletal muscle, and is conserved throughout evolution. Thus, ART-27 is a novel cofactor that interacts with the AR N terminus and plays a role in facilitating receptor-induced transcriptional activation
— id: 39704, year: 2002, vol: 13, page: 670, stat: Journal Article,

Src and Pyk2 mediate G-protein-coupled receptor activation of epidermal growth factor receptor (EGFR) but are not required for coupling to the mitogen-activated protein (MAP) kinase signaling cascade
Andreev J; Galisteo ML; Kranenburg O; Logan SK; Chiu ES; Okigaki M; Cary LA; Moolenaar WH; Schlessinger J
2001 Jun 8;276(23):20130-20135, Journal of biological chemistry
The epidermal growth factor receptor (EGFR) and the non-receptor protein tyrosine kinases Src and Pyk2 have been implicated in linking a variety of G-protein-coupled receptors (GPCR) to the mitogen-activated protein (MAP) kinase signaling cascade. In this report we apply a genetic strategy using cells isolated from Src-, Pyk2-, or EGFR-deficient mice to explore the roles played by these protein tyrosine kinases in GPCR-induced activation of EGFR, Pyk2, and MAP kinase. We show that Src kinases are critical for activation of Pyk2 in response to GPCR-stimulation and that Pyk2 and Src are essential for GPCR-induced tyrosine phosphorylation of EGFR. By contrast, Pyk2, Src, and EGFR are dispensable for GPCR-induced activation of MAP kinase. Moreover, GPCR-induced MAP kinase activation is normal in fibroblasts deficient in both Src and Pyk2 (Src-/-Pyk2-/- cells) as well as in fibroblasts deficient in all three Src kinases expressed in these cells (Src-/-Yes-/-Fyn-/- cells). Finally, experiments are presented demonstrating that, upon stimulation of GPCR, activated Pyk2 forms a complex with Src, which in turn phosphorylates EGFR directly. These experiments reveal a role for Src kinases in Pyk2 activation and a role for Pyk2 and Src in tyrosine phosphorylation of EGFR following GPCR stimulation. In addition, EGFR, Src family kinases, and Pyk2 are not required for linking GPCRs with the MAP kinase signaling cascade
— id: 21223, year: 2001, vol: 276, page: 20130, stat: Journal Article,

Activation of phospholipase C gamma by PI 3-kinase-induced PH domain-mediated membrane targeting
Falasca M; Logan SK; Lehto VP; Baccante G; Lemmon MA; Schlessinger J
1998 Jan 15;17(2):414-422, EMBO journal
Signaling via growth factor receptors frequently results in the concomitant activation of phospholipase C gamma (PLC gamma) and phosphatidylinositol (PI) 3-kinase. While it is well established that tyrosine phosphorylation of PLC gamma is necessary for its activation, we show here that PLC gamma is regulated additionally by the lipid products of PI 3-kinase. We demonstrate that the pleckstrin homology (PH) domain of PLC gamma binds to phosphatidylinositol 3,4,5-trisphosphate [PdtIns(3,4,5)P3], and is targeted to the membrane in response to growth factor stimulation, while a mutated version of this PH domain that does not bind PdtIns(3,4,5)P3 is not membrane targeted. Consistent with these observations, activation of PI 3-kinase causes PLC gamma PH domain-mediated membrane targeting and PLC gamma activation. By contrast, either the inhibition of PI 3-kinase by overexpression of a dominant-negative mutant or the prevention of PLC gamma membrane targeting by overexpression of the PLC gamma PH domain prevents growth factor-induced PLC gamma activation. These experiments reveal a novel mechanism for cross-talk and mutual regulation of activity between two enzymes that participate in the control of phosphoinositide metabolism
— id: 7559, year: 1998, vol: 17, page: 414, stat: Journal Article,

Antagonism of glucocorticoid receptor transcriptional activation by the c-Jun N-terminal kinase
Rogatsky I; Logan SK; Garabedian MJ
1998 Mar 3;95(5):2050-2055, Proceedings of the National Academy of Sciences of the United States of America
The mitogen-activated protein kinases ERK (extracellular signal-regulated kinase), JNK (c-Jun N-terminal kinase), and p38 phosphorylate and activate transcription factors that promote proliferative and inflammatory responses, whereas glucocorticoid receptor (GR) activation inhibits cell growth and inflammation. We demonstrate that JNK and ERK but not p38 phosphorylate GR in vitro primarily at Ser-246. Selective activation of either ERK or JNK in vivo inhibits GR-mediated transcriptional activation, which depends on receptor phosphorylation at Ser-246 by JNK but not ERK. Thus, JNK inhibits GR transcriptional activation by direct receptor phosphorylation, whereas ERK does so indirectly. We propose that phosphorylation of GR by JNK or of a GR cofactor by ERK provides mechanisms to ensure the rapid inhibition of GR-dependent gene expression when it conflicts with mitogenic or proinflammatory signals
— id: 7764, year: 1998, vol: 95, page: 2050, stat: Journal Article,

Phosphatidylinositol 3-kinase mediates epidermal growth factor-induced activation of the c-Jun N-terminal kinase signaling pathway
Logan SK; Falasca M; Hu P; Schlessinger J
1997 Oct;17(10):5784-5790, Molecular & cellular biology
The signaling events which mediate activation of c-Jun N-terminal kinase (JNK) are not yet well characterized. To broaden our understanding of upstream mediators which link extracellular signals to the JNK pathway, we investigated the role of phosphatidylinositol (PI) 3-kinase in epidermal growth factor (EGF)-mediated JNK activation. In this report we demonstrate that a dominant negative form of PI 3-kinase as well as the inhibitor wortmannin blocks EGF-induced JNK activation dramatically. However, wortmannin does not have an effect on JNK activation induced by UV irradiation or osmotic shock. In addition, a membrane-targeted, constitutively active PI 3-kinase (p110beta) was shown to produce in vivo products and to activate JNK, while a kinase-mutated form of this protein showed no activation. On the basis of these experiments, we propose that PI 3-kinase activity plays a role in EGF-induced JNK activation in these cells
— id: 7203, year: 1997, vol: 17, page: 5784, stat: Journal Article,

T-antigen inhibits metalloproteinase expression and invasion in human placental cells transformed with temperature-sensitive simian virus 40
Logan SK; Hansell EJ; Damsky CH; Werb Z
1996 Jul;15(2):81-89, Matrix biology
Cellular transformation frequently induces invasive behavior in cells. The effects of simian virus (SV) 40 T-antigen on the relationship between metalloproteinase expression and cell invasion were tested in human placental trophoblast-like cells transformed with a temperature-sensitive form of the SV40 virus, tsA30. As a comparison, metalloproteinase expression was also tested in human fibroblasts transformed with wild-type SV40 T-antigen. When tsA30.1 cells were cultured at the nonpermissive temperature for T-antigen expression, 40 degrees C, they expressed abundant metalloproteinases, including the 72 kDa gelatinase A (MMP-2), the 92 kDa gelatinase B (MMP-9) and stromelysin-1 (MMP-3). In contrast, tsA 30.1 cells cultured at the permissive temperature of 33 degrees C produced T-antigen and showed markedly decreased amounts of these proteinases. A similar suppression was seen in the human fibroblasts transformed with wild-type T-antigen. The tsA30 cells cultured at either temperature expressed a similar amount of tissue inhibitor of metalloproteinases-1 and -2. Cell invasion assays were performed to determine whether the altered ratio of proteinases to inhibitors under these conditions affected the extracellular matrix-degrading and invasive characteristics of the cells. In their differentiated state at the nonpermissive temperature for T-antigen expression, tsA30.1 cells were highly invasive, whereas at the permissive temperature they were not invasive. Therefore, the expression of T-antigen suppressed metalloproteinase production and changed the cells from an invasive to a noninvasive phenotype. We conclude that in tsA30.1 cells, SV 40 T-antigen expression suppresses metalloproteinase production, thereby decreasing the rate of degradation of the extracellular matrix. Taken together, these data indicate that invasive behavior is related to proteinase gene expression rather than to transformation by T-antigen. Function-blocking antibody to beta 1 integrins did not affect adhesion of tsA30.1 cells but inhibited invasion at the nonpermissive temperature, even though they continued to secrete proteinases. This observation indicates that beta 1 integrin-mediated cell migration is required along with proteinases for cells to be invasive
— id: 57671, year: 1996, vol: 15, page: 81, stat: Journal Article,

Synergistic transcriptional activation of the tissue inhibitor of metalloproteinases-1 promoter via functional interaction of AP-1 and Ets-1 transcription factors
Logan, S K; Garabedian, M J; Campbell, C E; Werb, Z
1996 Jan 12;271(2):774-782, Journal of biological chemistry
The tissue inhibitor of metalloproteinases-1 (TIMP-1) is an inhibitor of the extracellular matrix-degrading metalloproteinases. We characterized response elements that control TIMP-1 gene expression. One contains a binding site that selectively binds c-Fos and c-Jun in vitro and confers a response to multiple AP-1 family members in vivo. Adjacent to this is a binding site for Ets domain proteins. Although c-Ets-1 alone did not activate transcription from this element, it enhanced transcription synergistically with AP-1 either in the context of the natural promoter or when the sequence was linked upstream of a heterologous promoter. Furthermore, a complex of c-Jun and c-Fos interacted with c-Ets-1 in vitro. These results suggest that AP-1 tethers c-Ets-1 to the TIMP-1 promoter via protein-protein interaction to achieve Ets-dependent transcriptional regulation. Collectively, our results indicate that TIMP-1 expression is controlled by several DNA response elements that respond to variations in the level and activity of AP-1 and Ets transcriptional regulatory proteins
— id: 106246, year: 1996, vol: 271, page: 774, stat: Journal Article,

Integrin switching regulates normal trophoblast invasion
Damsky CH; Librach C; Lim KH; Fitzgerald ML; McMaster MT; Janatpour M; Zhou Y; Logan SK; Fisher SJ
1994 Dec;120(12):3657-3666, Development
Cells invade extracellular matrices in a regulated manner at specific times and places during normal development. A dramatic example is trophoblast invasion of the uterine wall. Previous studies have shown that differentiation of trophoblasts to an invasive phenotype is accompanied by temporally and spatially regulated switching of their integrin repertoire. In the first trimester human placenta, alpha 6 integrins are restricted to cytotrophoblast (CTB) stem cells and downregulated in invasive CTBs, whereas alpha 5 beta 1 and alpha 1 beta 1 integrins are upregulated in differentiating and invasive CTBs. The goal of the present study was to determine whether these changes have functional consequences for CTB invasiveness. Using an in vitro invasion model, we determined first that aggregates of invading first trimester CTBs in vitro undergo the same pattern of integrin switching as was observed in situ, thereby validating the utility of the model. We then showed that antibody perturbation of interactions involving laminin or collagen type IV and their integrin alpha 1/beta 1 receptor inhibited invasion by CTBs, whereas perturbing interactions between fibronectin and the alpha 5/beta 1 fibronectin receptor accelerated invasion. Finally, we report that later gestation CTBs, which display greatly decreased invasive capacity, are also unable to upregulate alpha 1 beta 1 complexes, providing further evidence that this integrin is critical for CTB invasion. This gestational regulation is transcriptional. These data indicate that integrin switching observed during differentiation in situ has significant functional consequences for CTB invasion. The data suggest further that differentiating CTBs upregulate counterbalancing invasion-accelerating and invasion-restraining adhesion mechanisms. We propose that this contributes to regulating the depth of CTB invasion during normal implantation
— id: 57672, year: 1994, vol: 120, page: 3657, stat: Journal Article,

Human placental cells transformed with temperature-sensitive simian virus 40 are immortalized and mimic the phenotype of invasive cytotrophoblasts at both permissive and nonpermissive temperatures
Logan SK; Fisher SJ; Damsky CH
1992 Nov 1;52(21):6001-6009, Cancer research
Establishment of the human placenta is essential for subsequent development of the embryo. Previous studies from our laboratories have demonstrated that chorionic villus cytotrophoblast stem cells undergo a stepwise differentiation process in vivo that results in their ability to invade the uterine wall. This process can be mimicked by isolated primary first-trimester cytotrophoblasts in vitro. Efforts to study the regulation of this differentiation pathway have been hampered by the inability of the isolated cytotrophoblast to replicate in culture. We therefore performed experiments to determine the suitability of the temperature-sensitive simian virus 40-transformed cell line SPA 255-26 (SPA-26), derived from early-gestation cytotrophoblasts, for studying the cytotrophoblast differentiation pathway that results in uterine invasion. Our results show that this cell line exhibits many properties of differentiated early-gestation cytotrophoblasts at both permissive and nonpermissive temperature. These cells were invasive in vitro and expressed the repertoire of hormones, adhesion molecules, and proteinases characteristic of an advanced stage of cytotrophoblast differentiation in vivo. Thus, these cells should be useful in studying the regulation of the adhesive and invasive behavior of human cytotrophoblasts
— id: 57673, year: 1992, vol: 52, page: 6001, stat: Journal Article,

Ovarian follicle cell enhancers from the Drosophila yolk protein genes: different segments of one enhancer have different cell-type specificities that interact to give normal expression
Logan SK; Wensink PC
1990 Apr;4(4):613-623, Genes & development
This paper examines ovarian transcription of the divergently oriented yolk protein genes 1 and 2 (yp1 and yp2) of Drosophila melanogaster. We report germ line transformation results demonstrating that yp1 and yp2 are transcribed in the same subpopulations of ovarian follicle cells. Our results show that this expression pattern is directed by two enhancers: ovarian enhancer 1, located between the genes, and ovarian enhancer 2, located within the first exon of yp2. Analysis of the expression pattern resulting from alterations in ovarian enhancer 1 demonstrates that different segments of this enhancer have different positive or negative effects on the cell-type specificity of transcription
— id: 57674, year: 1990, vol: 4, page: 613, stat: Journal Article,

DNA regions that regulate the ovarian transcriptional specificity of Drosophila yolk protein genes
Logan SK; Garabedian MJ; Wensink PC
1989 Sep;3(9):1453-1461, Genes & development
Yolk protein genes 1 and 2 (yp1 and yp2) of Drosophila melanogaster are divergently transcribed neighboring genes. Both are transcribed in only two tissues, the ovarian follicle cells and the fat bodies of adult females. Previous work has identified a yolk protein enhancer between the genes that is sufficient to direct transcription in one of the tissues, female fat bodies. Using germ-line transformation methods, we identify two cis-acting regions with positive effects on transcription in ovaries. One, a 301-bp region located between the genes, influences both genes and is an enhancer determining the stage and cell type specificity of ovarian transcription. The other, a 105-bp region located in the first exon of yp2, acts across the yp2 promoter region to stimulate yp1 transcription in ovaries. Additional observations suggest how a single enhancer influences both promoters
— id: 57675, year: 1989, vol: 3, page: 1453, stat: Journal Article,