Silencing of GSTP1, a Prostate Cancer Prognostic Gene, by the Estrogen Receptor-β and Endothelial Nitric Oxide Synthase Complex

Abstract

We recently identified in prostate tumors (PCa) a transcriptional prognostic signature comprising a significant number of genes differentially regulated in patients with worse clinical outcome. Induction of up-regulated genes was due to chromatin remodeling by a combinatorial complex between estrogen receptor (ER)-␤ and endothelial nitric oxide synthase (eNOS). Here we show that this complex can also repress transcription of prognostic genes that are down-regulated in PCa, such as the glutathione transferase gene GSTP1. Silencing of GSTP1 is a common early event in prostate carcinogenesis, frequently caused by promoter hypermethylation. We validated loss of glutathione transferase (GST) P1-1 expression in vivo, in tissue microarrays from a retrospective cohort of patients, and correlated it with decreased disease-specific survival. Furthermore, we show that in PCa cultured cells ER␤/eNOS causes GSTP1 repression by being recruited at estrogen responsive elements in the gene promoter with consequential remodeling of local chromatin. Treatment with ER␤ antagonist or its natural ligand 5␣-androstane-3␤,17␤-diol, eNOS inhibitors or ER␤ small interference RNA abrogated the binding and reversed GSTP1 silencing, demonstrating the direct involvement of the complex. In vitro, GSTP1 silencing by ER␤/eNOS was specific for cells from patients with worse clinical outcome where it appeared the sole mechanism regulating GSTP1 expression because no promoter hypermethylation was present. However, in vivo chromatin immunoprecipitation assays on fresh PCa tissues demonstrated that silencing by ER␤/eNOS can coexist with promoter hypermethylation. Our findings reveal that the ER␤/eNOS complex can exert transcriptional repression and suggest that this may represent an epigenetic event favoring inactivation of the GSTP1 locus by methylation. Moreover, abrogation of ER␤/eNOS function by 3␤-adiol emphasizes the significance of circulating or locally produced sex steroid hormones or their metabolites in PCa biology with relevant clinical/therapeutic implications.

Expression of CYP1A1 and GSTP1 in Human Brain Tumor Tissues in Pakistan

“…In current study statistically significant downregulation of GSTP1 was observed in brain tumor samples compared to control samples. Similar results have also been observed in breast cancer (Erlap et al, 2013), prostate cancer (Okino et al, 2007;Re et al, 2011) and colon cancer (Ritchie et al, 2009).Whereas conflicting results for GSTP1 expression pattern have been reported in some of other studies (Masood et al, 2011;Uchida et al, 2013). In present study, significant down-regulation of GSTP1 was observed in more advance grade of brain tumor samples when compared with early grade of brain tumor.…”

MALAT1 and HOTAIR Long Non-Coding RNAs Play Opposite Role in Estrogen-Mediated Transcriptional Regulation in Prostate Cancer Cells

“…As reported in previous studies from our group, in response to estrogenic stimuli, eNOS and ERs form nuclear complexes that modulate gene transcription in HUVEC and prostate cancer cell lines6181923. Validation of the eNOS peaks within HOTAIR and MALAT1 regulatory regions as emerged by ChIP-Seq was confirmed by traditional ChIP in HUVEC, C27IM, and LNCaP cells in independent experiments (Fig.…”

“…Of note, responsiveness to ERβ selective ligand, 3βAdiol, was abrogated by both HOTAIR and MALAT1 knockdown (Supplementary Fig. S4), indicating that both lncRNAs are necessary to achieve a complete estrogen responsiveness upon treatment with this androgen metabolite capable to selectively bind ERβ2331. Similarly, an abrogation of pS2 estrogen sensitivity was observed by interference of MALAT1 (efficiency about 80%, Supplementary Fig.…”

“…The estrogen-regulated pattern of eNOS-containing complexes present along the regulatory genomic regions of a subgroup of cancer-associated lncRNAs, specifically HOTAIR and MALAT1, prompted us to investigate them as potential targets as well as effectors of the ER signaling in a non-canonical microenvironment such as that of prostate cancer. Compelling experimental data support a critical yet unresolved role of the estrogen receptor in PCa61923353637, mediated by its participation in a multiprotein complex containing eNOS and/or HIFs, along the regulatory genomic regions of a subset of genes associated with poor prognosis such as VEGF, Glut-1 and others.…”

The Histone Acetylase Activator Pentadecylidenemalonate 1b Rescues Proliferation and Differentiation in the Human Cardiac Mesenchymal Cells of Type 2 Diabetic Patients

“…Data analysis was conducted following the instructions of the SABiosciences Web site (http://www .sabiosciences.com/methylationdataanalysis.php). DNA for the methyl-DNA immunoprecipitation (MeDIp) assay was prepared from cross-linked chromatin, as previously described (15). Purified DNA (500-800 ng) was denatured and methylation analysis performed with MeDIP kit (Active Motif), according the manufacturer's instructions, using a specific monoclonal antibody to 5-methylcytosine or mouse IgG as the negative control.…”