Receptor Target Genes Research Articles

Relevance of vitamin D receptor target genes for monitoring the vitamin D responsiveness of primary human cells.

Vitamin D3 has transcriptome- and genome-wide effects and activates, via the binding of its metabolite 1α,25-dihydroxyvitamin D3 to the transcription factor vitamin D receptor (VDR), several hundred target genes. Using samples from a 5-month vitamin D3 intervention study (VitDmet), we recently reported that the expression of 12 VDR target genes in peripheral blood mononuclear cells (PBMCs) as well as 12 biochemical and clinical parameters of the study participants are significantly triggered by vitamin D3. In this study, we performed a more focused selection of further 12 VDR target genes and demonstrated that changes of their mRNA expression in PBMCs of VitDmet subjects significantly correlate with alterations of 25-hydroxyvitamin D3 serum levels. Network and self-organizing map analysis of these datasets together with that of the other 24 parameters was followed by relevance calculations and identified changes in parathyroid hormone serum levels and the expression of the newly selected genes STS, BCL6, ITGAM, LRRC25, LPGAT1 and TREM1 as well as of the previously reported genes DUSP10 and CD14 as the most relevant parameters for describing vitamin D responsiveness in vivo. Moreover, parameter relevance ranking allowed the segregation of study subjects into high and low responders. Due to the long intervention period the vitamin D response was not too prominent on the level of transcriptional activation. Therefore, we performed in the separate VitDbol trial a short-term but high dose stimulation with a vitamin D3 bolus. In PBMCs of VitDbol subjects we observed direct transcriptional effects on the selected VDR target genes, such as an up to 2.1-fold increase already one day after supplementation onset. In conclusion, both long-term and short-term vitamin D3 supplementation studies allow monitoring the vitamin D responsiveness of human individuals and represent new types of human in vivo vitamin D3 investigations.

Structure‐Activity Analysis of Efavirenz‐Mediated Modulation of Pregnane X Receptor Target Genes

Efavirenz (EFV) is a non-nucleoside reverse transcriptase inhibitor used to treat HIV. EFV increases the expression of drug metabolizing enzymes via activation of pregnane X receptor (PXR) and we aimed to identify the EFV moieties that contribute to this. Hepatocytes isolated from wild-type (WT) male mice were treated with 10µM EFV or 11 EFV analogs and mRNA levels of PXR target genes were measured using qPCR. EFV increased cytochrome P450 3a11 (Cyp3a11) mRNA 2.1 ± 0.1-, 9.0 ± 4.7- and 76.6 ± 17.9-fold as compared to control at 24 h, 48 h and 72 h, respectively, and this was abrogated in hepatocytes isolated from PXR null mice. Using the EFV analogs, PXR activation was observed to be size-dependent as the smallest analog (223 Da) failed to increase Cyp3a11 mRNA, while analogs 290 Da to 424 Da in size increased Cyp3a11 mRNA similarly to EFV. Interestingly, mRNA levels of small heterodimer partner (SHP), which is transcriptionally repressed by PXR, was also increased by EFV treatment of WT mouse hepatocytes to 2.1 ± 0.6- and 2.0 ± 0.3-fold of control at 48 h and 72 h, respectively. Further, this effect was augmented in PXR null mice as the SHP mRNA levels were observed to be 4.3 ± 0.8-greater in EFV-treated mouse hepatocytes than in controls at 72 h. Since SHP is positively regulated by farnesoid X receptor (FXR) activation, hepatocytes isolated from FXR null mice were treated with EFV, however, the EFV-mediated increase in SHP mRNA abundance was not abolished in these mice. In conclusion, using a series of EFV analogs, we have demonstrated size-dependent activation of PXR and, in addition, we have identified EFV as a modulator of SHP mRNA levels. Funding: R01 GM103853 and UM1 AI068613

MP31-15 GENE EXPRESSION PROFILING REVEALS MOLECULAR SUBTYPES OF BENIGN PROSTATIC HYPERPLASIA

You have accessJournal of UrologyBenign Prostatic Hyperplasia: Basic Research1 Apr 2015MP31-15 GENE EXPRESSION PROFILING REVEALS MOLECULAR SUBTYPES OF BENIGN PROSTATIC HYPERPLASIA Keyan Salari, Seth Bechis, Rongbin Ge, Jian Hong, Aleksander Otsetov, Zongwei Wang, Shahin Tabatabaei, and Aria Olumi Keyan SalariKeyan Salari More articles by this author , Seth BechisSeth Bechis More articles by this author , Rongbin GeRongbin Ge More articles by this author , Jian HongJian Hong More articles by this author , Aleksander OtsetovAleksander Otsetov More articles by this author , Zongwei WangZongwei Wang More articles by this author , Shahin TabatabaeiShahin Tabatabaei More articles by this author , and Aria OlumiAria Olumi More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2015.02.1370AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES Benign prostatic hyperplasia (BPH) affects over 80% of men by age 70 and poses a substantial burden on healthcare. At least 25-30% of men do not respond to 5-alpha reductase (5-AR) inhibitors, raising the hypothesis that there may be molecular heterogeneity in BPH underlying this variability in clinical response. In previous work, we showed that some men carry epigenetic modifications via methylation of the 5-AR promoter region that modulate gene expression and protein levels, thereby suggesting a possible mechanism of resistance to 5-AR inhibitors. In this study, we performed genome-wide gene expression profiling of prostatic tissue from patients with BPH to identify molecular subtypes of the disease with potential clinical relevance. METHODS Prostatic tissue specimens were obtained at the time of surgery from 22 patients with symptomatic BPH undergoing prostate debulking surgery. Each patient's tissue was analyzed for the presence of 5-AR type 2 protein expression by immunohistochemistry and 5-AR2 gene promoter methylation status by PCR-based methylation analysis. Total RNA was extracted from each specimen for gene expression profiling. Whole-genome gene expression profiling was performed using Illumina Human HT-12 v4 BeadChip Arrays. Gene expression microarray data was analyzed by unsupervised hierarchical clustering and Gene Set Enrichment Analysis (GSEA) using the GenePattern software platform. RESULTS Unsupervised hierarchical clustering analysis of whole-genome gene expression profiles identified two subtypes of patients with distinct gene expression signatures. Among the most differentially expressed genes were several notable androgen-regulated genes, including TMPRSS2, NKX3-1, AZGP1, KLK3 (encoding the PSA protein) and multiple other members of the kallikrein family of serine proteases (all P <0.001, FDR <0.02). By GSEA, multiple gene sets composed of androgen-regulated genes and androgen receptor target genes were found to be significantly enriched in one of the two subtypes. Interestingly, patients from this Androgen-Up subtype were also noted to have a higher frequency of methylation of the 5AR-2 gene promoter (though not statistically significant). CONCLUSIONS Patients with BPH exhibit variable prostatic gene expression patterns. Our findings suggest the presence of distinct molecular subtypes of BPH, driven by differences in androgen gene regulation. These differences may partially explain the variable response of patients to 5-AR inhibitors and thereby inform clinical strategies for devising personalized therapy for BPH patients. © 2015 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 193Issue 4SApril 2015Page: e361 Advertisement Copyright & Permissions© 2015 by American Urological Association Education and Research, Inc.MetricsAuthor Information Keyan Salari More articles by this author Seth Bechis More articles by this author Rongbin Ge More articles by this author Jian Hong More articles by this author Aleksander Otsetov More articles by this author Zongwei Wang More articles by this author Shahin Tabatabaei More articles by this author Aria Olumi More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

Developmental exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin alters DNA methyltransferase (dnmt) expression in zebrafish (Danio rerio)

DNA methylation is one of the most important epigenetic modifications involved in the regulation of gene expression. The DNA methylation reaction is catalyzed by DNA methyltransferases (DNMTs). Recent studies have demonstrated that toxicants can affect normal development by altering DNA methylation patterns, but the mechanisms of action are poorly understood. Hence, we tested the hypothesis that developmental exposure to TCDD affects dnmt gene expression patterns. Zebrafish embryos were exposed to 5nM TCDD for 1h from 4 to 5h post-fertilization (hpf) and sampled at 12, 24, 48, 72, and 96 hpf to determine dnmt gene expression and DNA methylation patterns. We performed a detailed analysis of zebrafish dnmt gene expression during development and in adult tissues. Our results demonstrate that dnmt3b genes are highly expressed in early stages of development, and dnmt3a genes are more abundant in later stages. TCDD exposure upregulated dnmt1 and dnmt3b2 expression, whereas dnmt3a1, 3b1, and 3b4 are downregulated following exposure. We did not observe any TCDD-induced differences in global methylation or hydroxymethylation levels, but the promoter methylation of aryl hydrocarbon receptor (AHR) target genes was altered. In TCDD-exposed embryos, AHR repressor a (ahrra) and c-fos promoters were differentially methylated. To characterize the TCDD effects on DNMTs, we cloned the dnmt promoters with xenobiotic response elements and conducted AHR transactivation assays using a luciferase reporter system. Our results suggest that ahr2 can regulate dnmt3a1, dnmt3a2, and dnmt3b2 expression. Overall, we demonstrate that developmental exposure to TCDD alters dnmt expression and DNA methylation patterns.

PRMT1 and PRMT8 regulate retinoic acid-dependent neuronal differentiation with implications to neuropathology.

Retinoids are morphogens and have been implicated in cell fate commitment of embryonic stem cells (ESCs) to neurons. Their effects are mediated by RAR and RXR nuclear receptors. However, transcriptional cofactors required for cell and gene-specific retinoid signaling are not known. Here we show that protein arginine methyl transferase (PRMT) 1 and 8 have key roles in determining retinoid regulated gene expression and cellular specification in a multistage neuronal differentiation model of murine ESCs. PRMT1 acts as a selective modulator, providing the cells with a mechanism to reduce the potency of retinoid signals on regulatory "hotspots." PRMT8 is a retinoid receptor target gene itself and acts as a cell type specific transcriptional coactivator of retinoid signaling at later stages of differentiation. Lack of either of them leads to reduced nuclear arginine methylation, dysregulated neuronal gene expression, and altered neuronal activity. Importantly, depletion of PRMT8 results in altered expression of a distinct set of genes, including markers of gliomagenesis. PRMT8 is almost entirely absent in human glioblastoma tissues. We propose that PRMT1 and PRMT8 serve as a rheostat of retinoid signaling to determine neuronal cell specification in a context-dependent manner and might also be relevant in the development of human brain malignancy.

P,p′-Dichlorodiphenyltrichloroethane (p,p′-DDT) and p,p′-dichlorodiphenyldichloroethylene (p,p′-DDE) repress prostate specific antigen levels in human prostate cancer cell lines

Despite stringent restrictions on their use by many countries since the 1970s, the endocrine disrupting chemicals, DDT and DDE are still ubiquitous in the environment. However, little attention has been directed to p,p′-DDT and the anti-androgen, p,p′-DDE on androgen receptor (AR) target gene transcription in human cells. Inhibitors of androgenic activity may have a deleterious clinical outcome in prostate cancer screens and progression, therefore we determined whether environmentally relevant concentrations of p,p′-DDT and p,p′-DDE negatively impact AR-regulated expression of prostate-specific antigen (PSA), and other AR target genes in human LNCaP and VCaP prostate cancer cells. Quantitative real-time PCR and immuno-blotting techniques were used to measure intracellular PSA, PSMA and AR mRNA and protein levels. We have shown for the first time that p,p′-DDT and p,p′-DDE repressed R1881-inducible PSA mRNA and protein levels in a dose-dependent manner. Additionally, we used the fully automated COBAS PSA detection system to determine that extracellular PSA levels were also significantly repressed. These chemicals achieve this by blocking the recruitment of AR to the PSA promoter region at 10μM, as demonstrated by the chromatin immunoprecipitation (ChIP) in LNCaP cells. Both p,p′-DDT and p,p′-DDE repressed R1881-inducible AR protein accumulation at 10μM. Thus, we conclude that men who have been exposed to either DDT or DDE may produce a false-negative PSA test when screening for prostate cancer, resulting in an inaccurate clinical diagnosis. More importantly, prolonged exposure to these anti-androgens may mimic androgen ablation therapy in individuals with prostate cancer, thus exacerbating the condition by inadvertently forcing adaptation to this stress early in the disease.

Vitamin E in New-Generation Lipid Emulsions Protects Against Parenteral Nutrition-Associated Liver Disease in Parenteral Nutrition-Fed Preterm Pigs.

Parenteral nutrition (PN) in preterm infants leads to PN-associated liver disease (PNALD). PNALD has been linked to serum accumulation of phytosterols that are abundant in plant oil but absent in fish oil emulsions. Whether modifying the phytosterol and vitamin E composition of soy and fish oil lipid emulsions affects development of PNALD in preterm pigs. We measured markers of PNALD in preterm pigs that received 14 days of PN that included 1 of the following: (1) Intralipid (IL, 100% soybean oil), (2) Intralipid + vitamin E (ILE, d-α-tocopherol), (3) Omegaven (OV, 100% fish oil), or (4) Omegaven + phytosterols (PS, β-sitosterol, campesterol, and stigmasterol). Serum levels of direct bilirubin, gamma glutamyl transferase, serum triglyceride, low-density lipoprotein, and hepatic triglyceride content were significantly lower (P < .05) in the ILE, OV, and PS compared to IL. Hepatic cholesterol 7-hydroxylase and organic solute transporter-α expression was lower (P < .05) and portal plasma FGF19 higher in the ILE, OV, and PS vs IL. Hepatic expression of mitochondrial carnitine palmitoyltransferase 1A and microsomal cytochrome P450 2E1 fatty acid oxidation genes was higher in ILE, OV, and PS vs IL. In vivo (13)C-CDCA clearance and expression of pregnane X receptor target genes, cytochrome P450 3A29 and multidrug resistance-associated protein 2, were higher in ILE, OV, and PS vs IL. α-tocopherol in Omegaven and added to Intralipid prevented serum and liver increases in biliary and lipidemic markers of PNALD in preterm piglets. The addition of phytosterols to Omegaven did not produce evidence of PNALD.

Loss of periostin/OSF-2 in ErbB2/Neu-driven tumors results in androgen receptor-positive molecular apocrine-like tumors with reduced Notch1 activity

IntroductionPeriostin (Postn) is a secreted cell adhesion protein that activates signaling pathways to promote cancer cell survival, angiogenesis, invasion, and metastasis. Interestingly, Postn is frequently overexpressed in numerous human cancers, including breast, lung, colon, pancreatic, and ovarian cancer.MethodsUsing transgenic mice expressing the Neu oncogene in the mammary epithelium crossed into Postn-deficient animals, we have assessed the effect of Postn gene deletion on Neu-driven mammary tumorigenesis.ResultsAlthough Postn is exclusively expressed in the stromal fibroblasts of the mammary gland, Postn deletion does not affect mammary gland outgrowth during development or pregnancy. Furthermore, we find that loss of Postn in the mammary epithelium does not alter breast tumor initiation or growth in mouse mammary tumor virus (MMTV)-Neu expressing mice but results in an apocrine-like tumor phenotype. Surprisingly, we find that tumors derived from Postn-null animals express low levels of Notch protein and Hey1 mRNA but increased expression of androgen receptor (AR) and AR target genes. We show that tumor cells derived from wild-type animals do not proliferate when transplanted in a Postn-null environment but that this growth defect is rescued by the overexpression of active Notch or the AR target gene prolactin-induced protein (PIP/GCDFP-15).ConclusionsTogether our data suggest that loss of Postn in an ErbB2/Neu/HER2 overexpression model results in apocrine-like tumors that activate an AR-dependent pathway. This may have important implications for the treatment of breast cancers involving the therapeutic targeting of periostin or Notch signaling.Electronic supplementary materialThe online version of this article (doi:10.1186/s13058-014-0513-8) contains supplementary material, which is available to authorized users.

Screening Ingredients from Herbs against Pregnane X Receptor in the Study of Inductive Herb-Drug Interactions: Combining Pharmacophore and Docking-Based Rank Aggregation.

The issue of herb-drug interactions has been widely reported. Herbal ingredients can activate nuclear receptors and further induce the gene expression alteration of drug-metabolizing enzyme and/or transporter. Therefore, the herb-drug interaction will happen when the herbs and drugs are coadministered. This kind of interaction is called inductive herb-drug interactions. Pregnane X Receptor (PXR) and drug-metabolizing target genes are involved in most of inductive herb-drug interactions. To predict this kind of herb-drug interaction, the protocol could be simplified to only screen agonists of PXR from herbs because the relations of drugs with their metabolizing enzymes are well studied. Here, a combinational in silico strategy of pharmacophore modelling and docking-based rank aggregation (DRA) was employed to identify PXR's agonists. Firstly, 305 ingredients were screened out from 820 ingredients as candidate agonists of PXR with our pharmacophore model. Secondly, DRA was used to rerank the result of pharmacophore filtering. To validate our prediction, a curated herb-drug interaction database was built, which recorded 380 herb-drug interactions. Finally, among the top 10 herb ingredients from the ranking list, 6 ingredients were reported to involve in herb-drug interactions. The accuracy of our method is higher than other traditional methods. The strategy could be extended to studies on other inductive herb-drug interactions.

Atorvastatin induces bile acid-synthetic enzyme Cyp7a1 by suppressing FXR signaling in both liver and intestine in mice

Statins are effective cholesterol-lowering drugs to treat CVDs. Bile acids (BAs), the end products of cholesterol metabolism in the liver, are important nutrient and energy regulators. The present study aims to investigate how statins affect BA homeostasis in the enterohepatic circulation. Male C57BL/6 mice were treated with atorvastatin (100 mg/kg/day po) for 1 week, followed by BA profiling by ultra-performance LC-MS/MS. Atorvastatin decreased BA pool size, mainly due to less BA in the intestine. Surprisingly, atorvastatin did not alter total BAs in the serum or liver. Atorvastatin increased the ratio of 12α-OH/non12α-OH BAs. Atorvastatin increased the mRNAs of the BA-synthetic enzymes cholesterol 7α-hydroxylase (Cyp7a1) (over 10-fold) and cytochrome P450 27a1, the BA uptake transporters Na⁺/taurocholate cotransporting polypeptide and organic anion transporting polypeptide 1b2, and the efflux transporter multidrug resistance-associated protein 2 in the liver. Noticeably, atorvastatin suppressed the expression of BA nuclear receptor farnesoid X receptor (FXR) target genes, namely small heterodimer partner (liver) and fibroblast growth factor 15 (ileum). Furthermore, atorvastatin increased the mRNAs of the organic cation uptake transporter 1 and cholesterol efflux transporters Abcg5 and Abcg8 in the liver. The increased expression of BA-synthetic enzymes and BA transporters appear to be a compensatory response to maintain BA homeostasis after atorvastatin treatment. The Cyp7a1 induction by atorvastatin appears to be due to suppressed FXR signaling in both the liver and intestine.

Molecular modulation of estrogen-induced apoptosis by synthetic progestins in hormone replacement therapy: an insight into the women's health initiative study.

Hormone replacement therapy (HRT) is widely used to manage menopausal symptoms in women and can be comprised of an estrogen alone or an estrogen combined with a progestin. The Women's Health Initiative demonstrated in their randomized trials that estrogen alone HRT decreases the risk of breast cancer in postmenopausal women, whereas combined estrogen plus a progestin (medroxyprogesterone acetate, MPA) HRT increases this risk. Long-term estrogen-deprived MCF-7:5C cells were used to model the postmenopausal breast cancer cell environment. MPA is able to modify E2-induced apoptosis in MCF-7:5C cells. MPA, similar to dexamethasone, increases glucocorticoid receptor (GR) transcriptional activity, increases SGK1, a GR target gene, and can be blocked by RU486 (an antiglucocorticoid), suggesting that it functions through the GR. Norethindrone acetate (NETA), another progestin used in HRT, acts like an estrogen at high doses, upregulating estrogen receptor target genes and generating apoptosis in MCF-7:5C cells. The data suggest that women taking HRT comprising an estrogen plus MPA may have an increased risk of breast cancer due to MPA acting as a glucocorticoid and blunting E2-induced apoptosis in this environment. Therefore, perhaps other approved progestins (e.g., NETA) should be considered as alternatives to MPA.

Seeking genes responsible for developmental origins of health and disease from the fetal mouse liver following maternal food restriction.

Low birthweight resulting from a non-optimal fetal environment is correlated epidemiologically to a higher risk of adult diseases, and which has also been demonstrated using animal models for maternal undernutrition. In this study, we subjected pregnant mice to 50% food restriction (FR), and profiled gene expression and promoter DNA methylation genome-wide using the fetal livers. The fact that effect of food restriction is opposite between before and after birth encouraged us to hunt for genes that are expressed oppositely to adult calorie restriction (CR) using the maternal livers. Among oppositely regulated genes, we identified trib1 (tribbles homolog 1). Using genetically modified mice, trib1 has been shown to have a demonstrable contribution to a risk of hypertriglyceridaemia and insulin resistance. Our data showed that the trib1 expression and its promoter DNA methylation could be affected physiologically (by maternal nutrition), and therefore might be a strong candidate gene for developmental origins of adult diseases. Furthermore, lepr (leptin receptor) gene was downregulated by maternal FR, indicating its potential role in induction of obesity and diabetes. Gene expression as well as promoter DNA methylation profiling revealed that glucocorticoid receptor target genes were regulated by maternal FR. This supports previous studies that suggest an important role of fetal glucocorticoid exposure in the mechanism of developmental origins of diseases. Our transcriptomics profiling data also suggested that maternal FR impaired development of the immune system. An inventory of candidate genes responsible for developmental origins of health and disease is presented and discussed in this study.

Abstract 2141: Dietary feeding of Kava root extract inhibits prostate carcinogenesis in the transgenic adenocarcinoma of the mouse prostate (TRAMP) model

Abstract Kava kava has been traditionally used to prepare beverages and herbal medicines in the Pacific Islands. Men living in Fiji and drinking kava have low incidence of prostate cancer (PCa). However, the PCa incidence among Fijian men who had migrated to Australia, increased by 5.1-fold. To determine the chemopreventive efficacy of kava root extracts, TRAMP mice were fed with commercial kava root extract supplemented food. In the prevention protocol, TRAMP mice were fed with 3 or 6 g (0.3% or 0.6%) kava root extract /kg food starting at 6 weeks of age and ending at 12 weeks of age, resulting in decreased numbers of high-grade prostatic intraepithelial neoplasia and moderately differentiated adenocarcinomas in kava root extract fed mice compared to the mice fed with control food (p&amp;lt;0.01). In the intervention protocol, in which TRAMP mice were fed with 0.3% or 0.6% kava root extract food starting at 6 weeks of age and ending at 24 weeks of age, the average genitourinary (GU) weight of 0.6% kava root extract fed mice was significantly reduced when compared to control food fed mice (1.98±2.07 gram vs.3.63±4.28 gram, p&amp;lt;0.05, Mann-Whitney U and Kolmogorov-Smirnov test). The percentages of large tumors (GU weight &amp;gt; 0.9 gram) were also decreased from 86.4% in the control group to 52.2% and 43.5% in 0.3 and 0.6 % kava food groups, respectively. In addition, we showed that kawain, the main component of kava root extracts, inhibited LSD1 enzyme activity, enhanced H3K9 dimethylation and decreased the mRNA expression of androgen receptor target genes, including PSA and TMPRSS2, in LNCaP cells. Taken together, these results suggested that dietary consumption of kava products have the potential to reduce the risk of prostate cancer development. Citation Format: Xuesen Li, Christopher A. Blair, Xiaolin Zi. Dietary feeding of Kava root extract inhibits prostate carcinogenesis in the transgenic adenocarcinoma of the mouse prostate (TRAMP) model. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2141. doi:10.1158/1538-7445.AM2014-2141

Cyclin A2 and its associated kinase activity are required for optimal induction of progesterone receptor target genes in breast cancer cells

A role for the cell cycle protein cyclin A2 in regulating progesterone receptor (PR) activity is emerging. This study investigates the role of cyclin A2 in regulating endogenous PR activity in T47D breast cancer cells by depleting cyclin A2 expression and measuring PR target genes using q-RT-PCR. Targets examined included genes induced by the PR-B isoform more strongly than PR-A (SGK1, FKBP5), a gene induced predominantly by PR-A (HEF1), genes induced via PR tethering to other transcription factors (p21, p27), a gene induced in part via extra-nuclear PR signaling mechanisms (cyclin D1) and PR-repressed genes (DST, IL1R1). Progestin induction of target genes was reduced following cyclin A2 depletion. However, cyclin A2 depletion did not diminish progestin target gene repression. Furthermore, inhibition of the associated Cdk2 kinase activity of cyclin A2 also reduced progestin induction of target genes, while Cdk2 enhanced the interaction between PR and cyclin A2. These results demonstrate that cyclin A2 and its associated kinase activity are important for progestin-induced activation of endogenous PR target genes in breast cancer cells.

Targeted H3R26 deimination specifically facilitates estrogen receptor binding by modifying nucleosome structure.

Transcription factor binding to DNA in vivo causes the recruitment of chromatin modifiers that can cause changes in chromatin structure, including the modification of histone tails. We previously showed that estrogen receptor (ER) target gene activation is facilitated by peptidylarginine deiminase 2 (PAD2)-catalyzed histone H3R26 deimination (H3R26Cit). Here we report that the genomic distributions of ER and H3R26Cit in breast cancer cells are strikingly coincident, linearly correlated, and observed as early as 2 minutes following estradiol treatment. The H3R26Cit profile is unlike that of previously described histone modifications and is characterized by sharp, narrow peaks. Paired-end MNase ChIP-seq indicates that the charge-neutral H3R26Cit modification facilitates ER binding to DNA by altering the fine structure of the nucleosome. Clinically, we find that PAD2 and H3R26Cit levels correlate with ER expression in breast tumors and that high PAD2 expression is associated with increased survival in ER+ breast cancer patients. These findings provide insight into how transcription factors gain access to nucleosomal DNA and implicate PAD2 as a novel therapeutic target for ER+ breast cancer.

Jagged1 promotes osteoclast differentiation and inhibits proliferation by activating of Notch signaling pathway

Objective To study the role of Jadded1 and Notch signaling pathway played in the differentiation and proliferation of osteoclasts derived from human circulating CD14 + monocytes.Methods Peripheral blood CD14 + monocytes were isolated from healthy adult by Ficoll density gradient centrifugation combined with magnetic beads separation.Recombinant protein Jagged1 and γ-Secretase Inhibitor (DAPT) with Jagged1 were added to the culture system of CD14 + monocytes respectively.The mRNA expression of osteoclast markers tartrate-resistant acid phosphatase (TRAP),cathepsin K (CK),calctionin receptor (CTR) and Notch key target genes hairy and enhancer of split-1 (HES-1) and HES-related with YRPW motif-1 (HEY-1) were measured by quantitative real-time quantitative polymerase chain reaction (Real-time PCR).The formation of osteoclast,bone resorption,Notch expression and proliferation of CD14 + monocytes were detected by TRAP staining,scanning electron microscope,and cell counting kit-8 (CCK-8).Results High expression of Jagged1 in bone metastases specimens was observed in contrast of no expression or low expression in adjacent tissues.TRAP,CK,CTR,HES-1 and HEY-1 mRNA expression were significantly higher than the control group and DAPT group in Jadded1 group (P ＜ 0.05).TRAP + cell count,osteolytic area was significantly increased in Jagged1 group when compared with control and DAPT group,and no significant difference observed between the last two groups.Cell proliferation was inhibited in Jagged1 group also (P ＜ 0.05).Conclusion Jagged1 promotes osteoclast differentiation and inhibits proliferation by activating Notch signaling pathway. Key words: Jagged1 ; Osteoclasts ; Notch ; Differentiation ; Proliferation

SUMOylation modulates the transcriptional activity of androgen receptor in a target gene and pathway selective manner.

Androgen receptor (AR) plays an important regulatory role in prostate cancer. AR's transcriptional activity is regulated by androgenic ligands, but also by post-translational modifications, such as SUMOylation. To study the role of AR SUMOylation in genuine chromatin environment, we compared androgen-regulated gene expression and AR chromatin occupancy in PC-3 prostate cancer cell lines stably expressing wild-type (wt) or doubly SUMOylation site-mutated AR (AR-K386R,K520R). Our genome-wide gene expression analyses reveal that the SUMOylation modulates the AR function in a target gene and pathway selective manner. The transcripts that are differentially regulated by androgen and SUMOylation are linked to cellular movement, cell death, cellular proliferation, cellular development and cell cycle. Fittingly, SUMOylation mutant AR cells proliferate faster and are more sensitive to apoptosis. Moreover, ChIP-seq analyses show that the SUMOylation can modulate the chromatin occupancy of AR on many loci in a fashion that parallels their differential androgen-regulated expression. De novo motif analyses reveal that FOXA1, C/EBP and AP-1 motifs are differentially enriched at the wtAR- and the AR-K386R,K520R-preferred genomic binding positions. Taken together, our data indicate that SUMOylation does not simply repress the AR activity, but it regulates AR's interaction with the chromatin and the receptor's target gene selection.

The direct effect of estrogen on cell viability and apoptosis in human gastric cancer cells.

Epidemiology researches indicated that gastric cancer is a male-predominant disease; both expression level of estrogen and expression pattern of estrogen receptors (ERs) influence its carcinogenesis. But the direct effect of estrogen on gastric cancer cells is still unclear. This study aimed to explore the direct effect of β-estradiol (E2) on gastric cancer cells. SGC7901 and BGC823 were treated with a serial of concentrations of E2. The survival rates of both the cell lines were significantly reduced, and the reduction of viability was due to apoptosis triggered by E2 treatment. Caspase 3 was activated in response to the increasing E2 concentration in both SGC7901 and BGC823. Cleaved Caspase 3 fragments were detected, and the expression levels of Bcl-2 and Bcl-xL were reduced. Apoptosis was further confirmed by flow cytometry. The expression level of PEG10, an androgen receptor target gene, was reduced during E2 treatment. Both ERα and ERβ were expressed in these cell lines, and the result of bioinformatics analysis of gastric cancer from GEO datasets indicated that the expression levels of both ERα and ERβ were significantly higher in noncancerous gastric tissues than in gastric cancer tissues. Our research indicated that estrogen can reduce cell viability and promote apoptosis in gastric cancer cells directly; ERs expression level is associated with gastric cancer. Our research will help to understand the mechanism of gender disparity in gastric cancer.

Transgenic expression of dominant-active IDOL in liver causes diet-induced hypercholesterolemia and atherosclerosis in mice.

The E3 ubiquitin ligase inducible degrader of the low-density lipoprotein receptor (IDOL) triggers lysosomal degradation of the low-density lipoprotein receptor. The tissue-specific effects of the IDOL pathway on plasma cholesterol and atherosclerosis have not been examined. Given that the liver is the primary determinant of plasma cholesterol levels, we sought to examine the consequence of effect of chronic liver-specific expression of a dominant-active form of IDOL in mice. We expressed a degradation-resistant, dominant-active form of IDOL (super IDOL [sIDOL]) in C57Bl/6J mice from the liver-specific albumin promoter (L-sIDOL transgenics). L-sIDOL mice were fed a Western diet for 20 or 30 weeks and then analyzed for plasma lipid levels and atherosclerotic lesion formation. L-sIDOL mice showed dramatic reductions in hepatic low-density lipoprotein receptor protein and increased plasma low-density lipoprotein cholesterol levels on both chow and Western diets. Moreover, L-sIDOL mice developed marked atherosclerotic lesions when fed a Western diet. Lesion formation in L-sIDOL mice was more robust than in apolipoprotein E*3 Leiden mice and did not require the addition of cholate to the diet. Western diet-fed L-sIDOL mice had elevated expression of liver X receptor target genes and proinflammatory genes in their aortas. Liver-specific expression of dominant-active IDOL is associated with hypercholesterolemia and a marked elevation in atherosclerotic lesions. Our results show that increased activity of the IDOL pathway in the liver can override other low-density lipoprotein receptor regulatory pathways leading to cardiovascular disease. L-sIDOL mice are a robust, dominantly inherited, diet-inducible model for the study of atherosclerosis.

P0040 Expression analysis of oestrogen receptor target genes in renal cell carcinoma

<h3>Background</h3> The aim of this study was to explore differentially expressed genes (DEGs) and target genes of oestrogen receptors in renal cell carcinoma. <h3>Methods</h3> Data (GSE12090) were downloaded from the gene expression omnibus database. After data preprocessing using the affy package, DEGs were selected with the significance analysis of microarray algorithm within the siggenes package. DEGs were then subjected to function and pathway enrichment analysis by using DAVID (Database for Annotation Visualization and Integrated Discovery) software. Furthermore, the transcriptional regulatory networks between the DEGs and transcription factors were constructed. Finally, the oestrogen receptor target genes were subjected to Gene Ontology enrichment analysis. <h3>Findings</h3> A total of 215 DEGs were identified between the chromophobe renal cell carcinoma samples and the oncocytoma samples, including 126 up-regulated and 89 down-regulated genes. Function enrichment showed that 25% of the DEGs were significantly enriched into plasma membrane. Among those DEGs, 105 were regulated by oestrogen receptors. Further regulatory network analysis indicated that oestrogen receptors were mainly involved in the expression regulation of oncogenes and tumour suppressor genes, such as <i>PRSS8</i> (protease serine 8), <i>CLDN7</i> (claudin 7), and <i>RAB25</i> (Ras-related protein Rab-25). <h3>Interpretation</h3> In this study, the identified oestrogen receptor target genes were closely related to tumour development, which was helpful for understanding the regulation mechanism of oestrogen receptors during tumour development and for promoting the discovery of predictive markers in renal cell carcinoma.