Estrogen-related Receptor Gamma Research Articles

Dependence of Hippocampal Function on ERRγ-Regulated Mitochondrial Metabolism

Neurons utilize mitochondrial oxidative phosphorylation (OxPhos) to generate energy essential for survival, function, and behavioral output. Unlike most cells that burn both fat and sugar, neurons only burn sugar. Despite its importance, how neurons meet the increased energy demands of complex behaviors such as learning and memory is poorly understood. Here we show that the estrogen-related receptor gamma (ERRγ) orchestrates the expression of a distinct neural gene network promoting mitochondrial oxidative metabolism that reflects the extraordinary neuronal dependence on glucose. ERRγ(-/-) neurons exhibit decreased metabolic capacity. Impairment of long-term potentiation (LTP) in ERRγ(-/-) hippocampal slices can be fully rescued by the mitochondrial OxPhos substrate pyruvate, functionally linking the ERRγ knockout metabolic phenotype and memory formation. Consistent with this notion, mice lacking neuronal ERRγ in cerebral cortex and hippocampus exhibit defects in spatial learning and memory. These findings implicate neuronal ERRγ in the metabolic adaptations required for memory formation.

The effects of PGC-1α on the proliferation and energy metabolism of malignant endometrial cancer cells.

BackgroundIt is well known that peroxisome proliferator-activated receptor gamma coactivator-1 alpha (PGC-1α) plays an important role in tissue energy metabolism. However, the roles of PGC-1α in malignant endometrial cancer remain unknown.MethodsForty cases of endometrial carcinoma, 15 cases with proliferative endometrial tissues, and 21 cases with normal endometrial tissues were collected. Real-time polymerase chain reaction was used to detect the mRNA levels of PGC-1α and estrogen-related receptor gamma (ERRγ). ELISA (enzyme-linked immunosorbent assay) was used to detect the concentrations of pyruvate kinase and isocitrate dehydrogenase. The results were analyzed using medical statistical methods.ResultsThe mRNA levels of PGC-1α and ERRγ in the endometrial carcinoma tissues and hyperplasic endometrial tissues were significantly greater than those in the normal endometria. The mRNA levels of PGC-1α and ERRγ in the endometrial carcinoma patients with type 2 diabetes were higher than those in patients without diabetes. The mRNA levels of PGC-1α and ERRγ in the endometrial adenocarcinomas increased with clinical staging, depth of myometrial invasion, and increases in the number of metastatic lymph nodes. The PGC-1α mRNA level was positively correlated with ERRγ in the endometrial carcinoma tissues. The mRNA levels of PGC-1α were positively correlated with the concentrations of pyruvate kinase and isocitrate dehydrogenase in the endometrial carcinoma tissues, and similar results were found for ERRγ.ConclusionOur results suggested that the upregulation of PGC-1α and ERRγ in endometrial cancer might be a requirement for cancer cell energy metabolism, which contributes to the development of endometrial cancer.

Estrogen-related receptor gamma regulates dopaminergic neuronal phenotype by activating GSK3β/NFAT signaling in SH-SY5Y cells.

The orphan nuclear receptor estrogen-related receptor gamma (ERRγ) is highly expressed in the nervous system during embryogenesis and in adult brains, but its physiological role in neuronal development remains unknown. In this study, we evaluated the relevance of ERRγ in regulating dopaminergic (DAergic) phenotype and the corresponding signaling pathway. We used retinoic acid (RA) to differentiate human neuroblastoma SH-SY5Y cells. RA induced neurite outgrowth of SH-SY5Y cells with an increase in DAergic neuron-like properties, including up-regulation of tyrosine hydroxylase, dopamine transporter, and vesicular monoamine transporter 2. ERRγ, but not ERRα, was up-regulated by RA, and participated in RA effect on SH-SY5Y cells. ERRγ over-expression enhanced mature DAergic neuronal phenotype with neurite outgrowth as with RA treatment; and RA-induced increase in DAergic phenotype was attenuated by silencing ERRγ expression. ERRγ appears to have a crucial role in morphological and functional regulation of cells that is selective for DAergic neurons. Polo-like kinase 2 was up-regulated in ERRγ-over-expressing SH-SY5Y cells, which was involved in phosphorylation of glycogen synthase kinase 3β and resulting downstream activation of nuclear factor of activated T cells. The likely involvement of ERRγ in regulating the DAergic neuronal phenotype makes this orphan nuclear receptor a novel target for understanding DAergic neuronal differentiation. We propose the relevance of estrogen-related receptor gamma (ERRγ) in regulating dopaminergic neuronal phenotype: ERRγ is up-regulated by retinoic acid in SH-SY5Y cells, and enhances dopaminergic phenotypes and induces neurite outgrowth; Polo-like kinase 2 (PLK2) and glycogen synthase kinase 3 beta/nuclear factor of activated T cells (GSK3β/NFAT) signaling are responsible for the ERRγ effect. Our findings provide the first insights into the role of ERRγ in the brain, as a novel approach toward understanding dopaminergic differentiation.

Cytoplasmic PELP1 and ERRgamma protect human mammary epithelial cells from Tam-induced cell death.

Tamoxifen (Tam) is the only FDA-approved chemoprevention agent for pre-menopausal women at high risk for developing breast cancer. While Tam reduces a woman's risk of developing estrogen receptor positive (ER+) breast cancer, the molecular mechanisms associated with risk reduction are poorly understood. Prior studies have shown that cytoplasmic proline, glutamic acid and leucine rich protein 1 (PELP1) promotes Tam resistance in breast cancer cell lines. Herein, we tested for PELP1 localization in breast epithelial cells from women at high risk for developing breast cancer and found that PELP1 was localized to the cytoplasm in 36% of samples. In vitro, immortalized HMECs expressing a nuclear localization signal (NLS) mutant of PELP1 (PELP1-cyto) were resistant to Tam-induced death. Furthermore, PELP1-cyto signaling through estrogen-related receptor gamma (ERRγ) promoted cell survival in the presence of Tam. Overexpression of ERRγ in immortalized HMECs protected cells from Tam-induced death, while knockdown of ERRγ sensitized PELP1-cyto expressing HMECs to Tam. Moreover, Tam-induced HMEC cell death was independent of apoptosis and involved accumulation of the autophagy marker LC3-II. Expression of PELP1-cyto and ERRγ reduced Tam-induced LC3-II accumulation, and knockdown of ERRγ increased LC3-II levels in response to Tam. Additionally, PELP1-cyto expression led to the upregulation of MMP-3 and MAOB, known PELP1 and ERRγ target genes, respectively. Our data indicate that cytoplasmic PELP1 induces signaling pathways that converge on ERRγ to promote cell survival in the presence of Tam. These data suggest that PELP1 localization and/or ERRγ activation could be developed as tissue biomarkers for Tam responsiveness.

MicroRNA-320a sensitizes tamoxifen-resistant breast cancer cells to tamoxifen by targeting ARPP-19 and ERRγ.

Tamoxifen represents a major adjuvant therapy to those patients with estrogen receptor-alpha positive breast cancer. However, tamoxifen resistance occurs quite often, either de novo or acquired during treatment. To investigate the role of miR-320a in the development of resistance to tamoxifen, we established tamoxifen-resistant (TamR) models by continually exposing MCF-7 or T47D breast cancer cells to tamoxifen, and identified microRNA(miRNA)-320a as a down-regulated miRNA in tamoxifen resistant cells. Re-expression of miR-320a was sufficient to sensitize TamR cells to tamoxifen by targeting cAMP-regulated phosphoprotein (ARPP-19) and estrogen-related receptor gamma (ERRγ) as well as their downstream effectors, c-Myc and Cyclin D1. Furthermore, progesterone (P4) promoted the expression of miR-320a by repressing c-Myc expression, while estrogen (E2) exerted the opposite effect. These results suggest the potential therapeutic approach for tamoxifen-resistant breast cancer by restorating miR-320a expression or depleting ARPP-19/ERRγ expression.

Mechanisms of pluripotency and epigenetic reprogramming in primordial germ cells: lessons for the conversion of other cell types into the stem cell lineage

Primordial germ cells (PGCs) provide an excellent tool to better understand ancestor?descendent relationships as well as the efficiency and molecular mechanisms governing pluripotency in the reprogramming of somatic cells, since the latter type of cells have a relatively lower efficiency of conversion to pluripotent cells. This kind of comparison has gained credence from the commonalities regarding the expression of key transcription factors such as octamer-binding transcriptionhttp://en.wikipedia.org/wiki/Transcription_factor factor-4 (Oct3/4), SRY-related HMG box (Sox2), myelocytomatosis (c-Myc), and Nanog, as well as redundancy in terms of Kruppel-like factor 2 (Klf2), Kruppel-like factor 5 (Klf5), estrogen-related receptor beta (Esrrb), and estrogen-related receptor gamma (Esrrg) compensating for the absence of Kruppel-like factor 4 (Klf4). However, the exogenous addition of any one of these factors was found to be important, thereby implying that the expression level is important. L-Myelocytomatosis (L-myc) was shown to improve reprogramming efficiency without affecting tumorigenic potential. Molecular aspects of epigenetic reprogramming during the acquisition of pluripotency, as well as tumorigenic potential, have also been discussed, thus providing an understanding of the factors that can improve the former without increasing the possibility of neoplastic transformation. An improved understanding of the molecular events would pave the way for the development and use of endogenous biomolecules as well as currently available chemical reprogrammers for improving the efficiency of conversion of PGCs into cells of the stem cell lineage. Such chemicals, when adequately tested, can possibly be an alternative to viral vectors, since the introduced transgenes can become oncogenic.

The miR-545/374a cluster encoded in the Ftx lncRNA is overexpressed in HBV-related hepatocellular carcinoma and promotes tumorigenesis and tumor progression.

Hepatitis B virus (HBV) infection is a major risk factor for hepatocellular carcinoma (HCC). Previous studies have shown several long noncoding RNAs (lncRNAs) play various roles in HCC progression, but no research has focused on the expression pattern of microRNA clusters encoded in lncRNAs. The Ftx gene encodes a lncRNA which harbors 2 clusters of microRNAs in its introns, the miR-374b/421 cluster and the miR-545/374a cluster. To date, no research has focused on the role of the miR-545/374a and miR-374b/421 clusters in HBV-related HCC. In this study, 66 pairs of HBV-related HCC tissue and matched non-cancerous liver tissue specimens were analyzed for the expression of the Ftx microRNA clusters. Our results showed that the miR-545/374a cluster was upregulated in HBV-HCC tissue and significantly correlated with prognosis-related clinical features, including histological grade, metastasis and tumor capsule. Transfection studies with microRNA mimics and inhibitors revealed that miR-545/374a expression promoted in vitro cell proliferation, cell migration and invasion. The wild-type HBV-genome-containing plasmid or full-length HBx protein encoding plasmid was transfected into the Bel-7402 cell line and observed for their influence on miR-545/374a expression. We found that transfection of the HBV genome or HBx alone resulted in an increase in miR-545/374a expression. Next, by monitoring the expression of sera miR-545/374a before and after surgical tumor excision, we found serum miR-545/374a was tumor-derived and exhibited a sharp decrease 25 days after tumor excision. We also examined the gender-based difference in miR-545/374a expression among HCC patients and utilized microRNA target prediction software to find the targets of miR-545/374a. One of these targets, namely estrogen-related receptor gamma (ESRRG) was inversely correlated with miR-545 expression. In conclusion, the overexpression of miR-545/374a cluster located in the Ftx lncRNA is partially responsible for a poor prognosis, and monitoring sera levels of miR-545/374a may be a useful diagnostic marker for HCC.

The pERK of being a target: Kinase regulation of the orphan nuclear receptor ERRγ

Estrogen-related receptors (ERRs) are orphan members of the nuclear receptor superfamily that are important regulators of mitochondrial metabolism with emerging roles in cancer. In the absence of an endogenous ligand, ERRs are reliant upon other regulatory mechanisms that include protein/protein interactions and post-translational modification, though the cellular and clinical significance of this latter mechanism is unclear. We recently published a study in which we establish estrogen-related receptor gamma (ERRγ) as a target for extracellular signal-regulated kinase (ERK), and show that regulation of ERRγ by ERK has important consequences for the function of this receptor in cellular models of estrogen receptor-positive (ER+) breast cancer. In this Research Highlight, we discuss the implications of these findings from a molecular and clinical perspective.

ERK/MAPK regulates ERRγ expression, transcriptional activity and receptor-mediated tamoxifen resistance in ER+ breast cancer.

Selective estrogen receptor modulators such as tamoxifen (TAM) significantly improve breast cancer-specific survival for women with estrogen receptor-positive (ER+) disease. However, resistance to TAM remains a major clinical problem. The resistant phenotype is usually not driven by loss or mutation of the estrogen receptor; instead, changes in multiple proliferative and/or survival pathways over-ride the inhibitory effects of TAM. Estrogen-related receptorγ (ERRγ) is an orphan member of the nuclear receptor superfamily that promotes TAM resistance in ER+ breast cancer cells. This study sought to clarify the mechanism(s) by which this orphan nuclear receptor is regulated, and hence affects TAM resistance. mRNA and protein expression/phosphorylation were monitored by RT-PCR and western blotting, respectively. Site-directed mutagenesis was used to disrupt consensus extracellular signal-regulated kinase (ERK) target sites. Cell proliferation and cell-cycle progression were measured by flow cytometric methods. ERRγ transcriptional activity was assessed by dual-luciferase promoter-reporter assays. We show that ERRγ protein levels are affected by the activation state of ERK/mitogen-activated protein kinase, and mutation of consensus ERK target sites impairs ERRγ-driven transcriptional activity and TAM resistance. These findings shed new light on the functional significance of ERRγ in ER+ breast cancer, and are the first to demonstrate a role for kinase regulation of this orphan nuclear receptor.

Orphan Nuclear Receptor Errγ Induces C-Reactive Protein Gene Expression through Induction of ER-Bound Bzip Transmembrane Transcription Factor CREBH

The orphan nuclear receptor estrogen-related receptor-γ (ERRγ) is a constitutively active transcription factor regulating genes involved in several important cellular processes, including hepatic glucose metabolism, alcohol metabolism, and the endoplasmic reticulum (ER) stress response. cAMP responsive element-binding protein H (CREBH) is an ER-bound bZIP family transcription factor that is activated upon ER stress and regulates genes encoding acute-phase proteins whose expression is increased in response to inflammation. Here, we report that ERRγ directly regulates CREBH gene expression in response to ER stress. ERRγ bound to the ERRγ response element (ERRE) in the CREBH promoter. Overexpression of ERRγ by adenovirus significantly increased expression of CREBH as well as C-reactive protein (CRP), whereas either knockdown of ERRγ or inhibition of ERRγ by ERRγ specific inverse agonist, GSK5182, substantially inhibited ER stress-mediated induction of CREBH and CRP. The transcriptional coactivator PGC1α was required for ERRγ mediated induction of the CREBH gene as demonstrated by the chromatin immunoprecipitation (ChIP) assay showing binding of both ERRγ and PGC1α on the CREBH promoter. The ChIP assay also revealed that histone H3 and H4 acetylation occurred at the ERRγ and PGC1α binding site. Moreover, chronic alcoholic hepatosteatosis, as well as the diabetic obese condition significantly increased CRP gene expression, and this increase was significantly attenuated by GSK5182 treatment. We suggest that orphan nuclear receptor ERRγ directly regulates the ER-bound transcription factor CREBH in response to ER stress and other metabolic conditions.

Review: Anti-Oxidant and Anti-Aging Properties of Equol in Prostate Health (BPH)

Benign prostatic hyperplasia (BPH) is the pathological cellular progression of glandular proliferation associated with aging. The primary changes in prostate disorders are mediated by the conversion of the principle androgen, testosterone, to its more potent metabolite, 5α-dihydrotestosterone (5α-DHT). However, recent evidence suggests that estrogen hormonal actions via estrogen receptor subtypes also play an important role in BPH. Current pharmaceutical options for BPH have advantages, limitations and adverse effects. Complementary and Alternative Medicine (CAM) treatments for BPH include botanicals such as polyphenols and isoflavones. Equol is a polyphenolic/isoflavonoid molecule derived from intestinal metabolism, dairy and dietary plant sources. Equol has potent anti-oxidant and anti-aging properties to decrease prostatic irritation and potentially neoplastic growth. It has the unique characteristic to bind specifically 5α-DHT by sequestering 5α-DHT from the androgen receptor (AR), thus decreasing androgen hormone actions to improve prostate health by acting as a selective androgen modulator (SAM). It also has affinity for estrogen related receptor gamma (ERR-γ) and estrogen receptor beta (ER-β) within the prostate that is known to improve male health via selective estrogen receptor modulatory (SERM) activities to decrease inflammation, cellular proliferation and carcinogenesis. The possible clinical efficacy of equol on the symptoms associated with BPH is presented and the reviewed findings suggest that equol may provide a well-tolerated and rapid beneficial therapy for BPH that can be used alone or in combination with current pharmaceutical therapies. The beneficial clinical efficacy of equol observed may be due to the multiple positive biological actions that are not present in current pharmaceutical treatments.

Cartilage-Specific Overexpression of ERRγ Results in Chondrodysplasia and Reduced Chondrocyte Proliferation

While the role of estrogen receptor-related receptor alpha (ERRα) in chondrogenesis has been investigated, the involvement of ERR gamma (ERRγ) has not been determined. To assess the effect of increased ERRγ activity on cartilage development in vivo, we generated two transgenic (Tg) lines overexpressing ERRγ2 via a chondrocyte-specific promoter; the two lines exhibited ∼3 and ∼5 fold increased ERRγ2 protein expression respectively in E14.5 Tg versus wild type (WT) limbs. On postnatal day seven (P7), we observed a 4–10% reduction in the size of the craniofacial, axial and appendicular skeletons in Tg versus WT mice. The reduction in bone length was already present at birth and did not appear to involve bones that are derived via intramembranous bone formation as the bones of the calvaria, clavicle, and the mandible developed normally. Histological analysis of P7 growth plates revealed a reduction in the length of the Tg versus WT growth plate, the majority of which was attributable to a reduced proliferative zone. The reduced proliferative zone paralleled a decrease in the number of Ki67-positive proliferating cells, with no significant change in apoptosis, and was accompanied by large cell-free swaths of cartilage matrix, which extended through multiple zones of the growth plate. Using a bioinformatics approach, we identified known chondrogenesis-associated genes with at least one predicted ERR binding site in their proximal promoters, as well as cell cycle regulators known to be regulated by ERRγ. Of the genes identified, Col2al, Agg, Pth1r, and Cdkn1b (p27) were significantly upregulated, suggesting that ERRγ2 negatively regulates chondrocyte proliferation and positively regulates matrix synthesis to coordinate growth plate height and organization.

Polymorphic AAAG Repeat Length in Estrogen-Related Receptor Gamma (ERRγ) and Risk of Breast Cancer in Iranian Women

Estrogen-related receptors (ERRs) alpha, beta, and gamma are orphan nuclear receptors that modulate the estrogen signaling pathway and play roles in the regulation of breast cancer cell growth. To determine the association between breast cancer risk and alleles of the tetranucleotide repeat (AAAG)n in the intron of ERRγ gene, a case–control study of 200 breast cancer patients and 200 controls was performed in Iranian women. Our results demonstrate that women with short AAAG repeat are at higher risk of breast cancer (OR 7). This result suggests a possible involvement of polymorphic AAAG repeat of ERRγ gene in regulating its expression.

Estrogen-related receptor gamma induces cardiac hypertrophy by activating GATA4

Estrogen-related receptor gamma (ERRγ) is an orphan nuclear receptor that has biological roles mainly in metabolism and that controls metabolic switching in perinatal heart. In adult heart diseases, however, the functional roles of ERRγ have not yet been elucidated. In the present study, we aimed to characterize the role of ERRγ in cardiac hypertrophy. The functional roles of ERRγ in the development of cardiac hypertrophy were examined in primary cultured cardiomyocytes and in animal models. ERRγ expression was increased in hearts from human hypertrophic cardiomyopathy patients and in both cellular and animal models of cardiac hypertrophy. Transgenic overexpression in mouse heart as well as forced expression of ERRγ in cardiomyocytes induced hypertrophic phenotypes. Knock-down of ERRγ blocked agonist-induced hypertrophic phenotypes. ERRγ bound directly to the proximal ERR-responsive element in the GATA4 promoter in a sequence-specific manner and thereby induced transcription. ERRγ-induced hypertrophy was blocked by inhibition of GATA4. GSK-5182, an inverse agonist of ERRγ, completely blocked cardiac hypertrophy in cardiomyocytes. It also prevented aortic banding-induced cardiac hypertrophy and fibrosis in mouse heart. These findings demonstrate a novel ERRγ/GATA4 signal cascade in the development of cardiac hypertrophy and suggest GSK-5182 as a possible therapeutic.

Computer-aided identification of novel protein targets of bisphenol A

The xenoestrogen bisphenol A (2,2-bis-(p-hydroxyphenyl)-2-propane, BPA) is a known endocrine-disrupting chemical used in the fabrication of plastics, resins and flame retardants, that can be found throughout the environment and in numerous every day products. Human exposure to this chemical is extensive and generally occurs via oral route because it leaches from the food and beverage containers that contain it. Although most of the effects related to BPA exposure have been linked to the activation of the estrogen receptor (ER), the mechanisms of the interaction of BPA with protein targets different from ER are still unknown. Therefore, the objective of this work was to use a bioinformatics approach to identify possible new targets for BPA. Docking studies were performed between the optimized structure of BPA and 271 proteins related to different biochemical processes, as selected by text-mining. Refinement docking experiments and conformational analyses were carried out using LigandScout 3.0 for the proteins selected through the affinity ranking (lower than −8.0kcal/mol). Several proteins including ERR gamma (−9.9kcal/mol), and dual specificity protein kinases CLK-4 (−9.5kcal/mol), CLK-1 (−9.1kcal/mol) and CLK-2 (−9.0kcal/mol) presented great in silico binding affinities for BPA. The interactions between those proteins and BPA were mostly hydrophobic with the presence of some hydrogen bonds formed by leucine and asparagine residues. Therefore, this study suggests that this endocrine disruptor may have other targets different from the ER.

Kaempferol inhibits cancer cell growth by antagonizing estrogen‐related receptor α and γ activities

Kaempferol is a dietary flavonoid that can function as a selective estrogen receptor modulator (SERM). Estrogen-related receptors alpha and gamma (ERRα and ERRγ) are orphan nuclear receptors that play important roles in mitochondrial biogenesis and cancer development. We have shown that kaempferol can functionally antagonize the activities of ERRs based on both response element reporter systems and target gene analysis. Kaempferol modulation of mitochondrial function and suppression cancer cell growth has been confirmed. These findings suggest that kaempferol may exert their anti-cancer activities through antagonizing ERRs activities.

Abstract 071: Estrogen-related Receptor Gamma Induces Cardiac Hypertrophy By Activating Gata4

Introduction— Estrogen-related receptor gamma (ERRγ) is an orphan nuclear receptor that has biological roles mainly in metabolism and it controls metabolic switching in perinatal heart. In adult heart diseases, however, the functional roles of ERRγ have not yet been elucidated. Hypothesis— In the present study, we aimed to characterize the role of ERRγ in cardiac hypertrophy. Here we show that ERRγ provokes cardiac hypertrophy by inducing GATA4 and that its inverse agonist, GSK-5182, prevents cardiac hypertrophy. Methods and Results— The functional roles of ERRγ in association with development of cardiac hypertrophy were examined in primarily cultured cardiomyocytes, in animal models, and in heart samples from human hypertrophic cardiomyopathy patients. ERRγ expression was increased in hearts obtained from human hypertrophic cardiomyopathy patients and in both agonist-induced cellular models and aortic banding-induced animal models of cardiac hypertrophy. Transgenic overexpression in mouse heart as well as forced expression of ERRγ in cardiomyocytes induced hypertrophic phenotypes. Knock-down of ERRγ blocked agonist-induced hypertrophic phenotypes. ERRγ directly bound to the proximal ERR-responsive element in the GATA4 promoter in a sequence-specific manner and thereby induced transcription. ERRγ-induced hypertrophy was blocked by inhibition of GATA4. GSK-5182 completely blocked cardiac hypertrophy in cardiomyocytes. It also prevented aortic banding-induced cardiac hypertrophy and fibrosis in mouse heart. Conclusion— These findings demonstrate a novel ERRγ/GATA4 signal cascade in the development of cardiac hypertrophy and suggest GSK-5182 as a possible therapeutic.

Muscle ERRγ mitigates Duchenne muscular dystrophyviametabolic and angiogenic reprogramming

Treatment of Duchenne muscular dystrophy (DMD) by replacing mutant dystrophin or restoring dystrophin-associated glycoprotein complex (DAG) has been clinically challenging. Instead, identifying and targeting muscle pathways deregulated in DMD will provide new therapeutic avenues. We report that the expression of nuclear receptor estrogen-related receptor-γ (ERRγ), and its metabolic and angiogenic targets are down-regulated (50-85%) in skeletal muscles of mdx mice (DMD model) vs. wild-type mice. Corelatively, oxidative myofibers, muscle vasculature, and exercise tolerance (33%) are decreased in mdx vs. wild-type mice. Overexpressing ERRγ selectively in the dystrophic muscles of the mdx mice restored metabolic and angiogenic gene expression compared with control mdx mice. Further, ERRγ enhanced muscle oxidative myofibers, vasculature, and blood flow (by 33-66%) and improved exercise tolerance (by 75%) in the dystrophic mice. Restoring muscle ERRγ pathway ameliorated muscle damage and also prevented DMD hallmarks of postexercise muscle damage, hypoxia, and fatigue in mdx mice. Notably, ERRγ did not restore sarcolemmal DAG complex, which is thus dispensable for antidystrophic effects of ERRγ. In summary, ERRγ-dependent metabolic and angiogenic gene program is defective in DMD, and we demonstrate that its restoration is a potential strategy for treating muscular dystrophy.

Sex-specific epigenetic disruption and behavioral changes following low-dose in utero bisphenol A exposure.

Bisphenol A (BPA) is an estrogenic endocrine disruptor widely used in the production of plastics. Increasing evidence indicates that in utero BPA exposure affects sexual differentiation and behavior; however, the mechanisms underlying these effects are unknown. We hypothesized that BPA may disrupt epigenetic programming of gene expression in the brain. Here, we provide evidence that maternal exposure during pregnancy to environmentally relevant doses of BPA (2, 20, and 200 µg/kg/d) in mice induces sex-specific, dose-dependent (linear and curvilinear), and brain region-specific changes in expression of genes encoding estrogen receptors (ERs; ERα and ERβ) and estrogen-related receptor-γ in juvenile offspring. Concomitantly, BPA altered mRNA levels of epigenetic regulators DNA methyltransferase (DNMT) 1 and DNMT3A in the juvenile cortex and hypothalamus, paralleling changes in estrogen-related receptors. Importantly, changes in ERα and DNMT expression in the cortex (males) and hypothalamus (females) were associated with DNA methylation changes in the ERα gene. BPA exposure induced persistent, largely sex-specific effects on social and anxiety-like behavior, leading to disruption of sexually dimorphic behaviors. Although postnatal maternal care was altered in mothers treated with BPA during pregnancy, the effects of in utero BPA were not found to be mediated by maternal care. However, our data suggest that increased maternal care may partially attenuate the effects of in utero BPA on DNA methylation. Overall, we demonstrate that low-dose prenatal BPA exposure induces lasting epigenetic disruption in the brain that possibly underlie enduring effects of BPA on brain function and behavior, especially regarding sexually dimorphic phenotypes.

Abstract 3570: Phospho-dependent regulation of ERRγ expression, transcriptional activity, and Tamoxifen resistance in ER+ breast cancer.

Abstract Selective estrogen receptor modulators (SERMs) such as Tamoxifen (TAM) can significantly improve breast cancer-specific survival for women with ER-positive (ER+) disease. However, resistance to TAM remains a major clinical problem. Estrogen-related receptor gamma (ERRγ) is an orphan nuclear receptor with broad, structural similarities to classical ER that is widely implicated in the transcriptional regulation of energy homeostasis. We previously reported that ERRγ is upregulated during the acquisition of TAM resistance in ER+ breast cancer cell lines, exogenous expression of ERRγ is sufficient to induce TAM resistance, and ERRγ mRNA is significantly increased in tumor samples from women with ER+ breast cancer who relapse following TAM treatment. Because ERRγ has no known ligand, our recent studies have focused on understanding how the expression and activity of this orphan nuclear receptor is regulated, and how this contributes to the TAM resistant phenotype. We have found that TAM-resistant breast cancer cells in which endogenous ERRγ is upregulated show a concomitant hyper activation of p44/p42 ERK. Furthermore, ERK activity (but not that of JNK or p38 MAPK) directly enhances ERRγ protein stability via Serines 57, 81, and/or 219 of the receptor. Phospho-deficient ERRγ is impaired in its ability to induce TAM resistance, and this is associated with a significant reduction in transcriptional activity at the estrogen-related response element (ERRE) half-site, in particular. This led us to hypothesize that ERRγ action at ERREs is most relevant to the development of TAM resistance. We examined a meta-list of validated, ERRE-containing ERR target genes in association with distant metastasis-free survival (DMFS) within 5 years of primary diagnosis in each of 3 publicly available clinical datasets comprised of ER+ breast cancer patients treated with TAM, and obtained a list of 37 differentially expressed targets. The proximal promoter regions of these 37 genes are enriched for binding sites for ELK1, a well-known ERK substrate. Sub-network analysis reveals enrichment of genes whose protein products regulate the mitochondrial unfolded protein response (UPRmito). We also performed differential dependency network (DDN) analysis in an independent dataset using the full meta-list of ERRγ target genes and also identified genes associated with UPRmito. These data suggest a potentially novel role for ERK-mediated regulation of ERRγ in mitochondrial protein folding in TAM-resistant breast cancer. Citation Format: Rebecca B. Riggins, Mary M. Heckler, Hemang Thakor, Cara C. Schafer, Salendra Singh, Ye Tian, Yuriy Gusev, Subha Madhavan, Yue Wang. Phospho-dependent regulation of ERRγ expression, transcriptional activity, and Tamoxifen resistance in ER+ breast cancer. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3570. doi:10.1158/1538-7445.AM2013-3570