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1.
Pharmacol Rev ; 75(6): 1233-1318, 2023 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-37586884

RESUMO

The NR superfamily comprises 48 transcription factors in humans that control a plethora of gene network programs involved in a wide range of physiologic processes. This review will summarize and discuss recent progress in NR biology and drug development derived from integrating various approaches, including biophysical techniques, structural studies, and translational investigation. We also highlight how defective NR signaling results in various diseases and disorders and how NRs can be targeted for therapeutic intervention via modulation via binding to synthetic lipophilic ligands. Furthermore, we also review recent studies that improved our understanding of NR structure and signaling. SIGNIFICANCE STATEMENT: Nuclear receptors (NRs) are ligand-regulated transcription factors that are critical regulators of myriad physiological processes. NRs serve as receptors for an array of drugs, and in this review, we provide an update on recent research into the roles of these drug targets.


Assuntos
Farmacologia Clínica , Humanos , Receptores Citoplasmáticos e Nucleares/metabolismo , Fatores de Transcrição/metabolismo , Proteínas de Transporte , Ligantes
2.
J Neurosci ; 41(24): 5190-5205, 2021 06 16.
Artigo em Inglês | MEDLINE | ID: mdl-33941651

RESUMO

Hypertension susceptibility in women increases at the transition to menopause, termed perimenopause, a state characterized by erratic estrogen fluctuation and extended hormone cycles. Elucidating the role of estrogen signaling in the emergence of hypertension during perimenopause has been hindered by animal models that are confounded by abrupt estrogen cessation or effects of aging. In the present study, accelerated ovarian failure (AOF) in estrogen receptor ß (ERß) reporter mice was induced by 4-vinylcyclohexene diepoxide in young mice to model early-stage ovarian failure (peri-AOF) characteristic of peri-menopause. It was found that administering ERß agonists suppressed elevated blood pressure in a model of neurogenic hypertension induced by angiotensin II (AngII) in peri-AOF, but not in age-matched male mice. It was also found that ERß agonist administration in peri-AOF females, but not males, suppressed the heightened NMDAR signaling and reactive oxygen production in ERß neurons in the hypothalamic paraventricular nucleus (PVN), a critical neural regulator of blood pressure. It was further shown that deleting ERß in the PVN of gonadally intact females produced a phenotype marked by a sensitivity to AngII hypertension. These results suggest that ERß signaling in the PVN plays an important role in blood pressure regulation in female mice and contributes to hypertension susceptibility in females at an early stage of ovarian failure comparable to human perimenopause.


Assuntos
Receptor beta de Estrogênio/metabolismo , Hipertensão/metabolismo , Plasticidade Neuronal/fisiologia , Núcleo Hipotalâmico Paraventricular/metabolismo , Perimenopausa/metabolismo , Animais , Modelos Animais de Doenças , Feminino , Hipertensão/etiologia , Camundongos , Camundongos Endogâmicos C57BL
3.
FASEB J ; 35(5): e21563, 2021 05.
Artigo em Inglês | MEDLINE | ID: mdl-33818810

RESUMO

One of the endogenous estrogens, 17ß-estradiol (E2 ) is a female steroid hormone secreted from the ovary. It is well established that E2 causes biochemical and histological changes in the uterus. However, it is not completely understood how E2 regulates the oviductal environment in vivo. In this study, we assessed the effect of E2 on each oviductal cell type, using an ovariectomized-hormone-replacement mouse model, single-cell RNA-sequencing (scRNA-seq), in situ hybridization, and cell-type-specific deletion in mice. We found that each cell type in the oviduct responded to E2 distinctively, especially ciliated and secretory epithelial cells. The treatment of exogenous E2 did not drastically alter the transcriptomic profile from that of endogenous E2 produced during estrus. Moreover, we have identified and validated genes of interest in our datasets that may be used as cell- and region-specific markers in the oviduct. Insulin-like growth factor 1 (Igf1) was characterized as an E2 -target gene in the mouse oviduct and was also expressed in human fallopian tubes. Deletion of Igf1 in progesterone receptor (Pgr)-expressing cells resulted in female subfertility, partially due to an embryo developmental defect and embryo retention within the oviduct. In summary, we have shown that oviductal cell types, including epithelial, stromal, and muscle cells, are differentially regulated by E2 and support gene expression changes, such as growth factors that are required for normal embryo development and transport in mouse models. Furthermore, we have identified cell-specific and region-specific gene markers for targeted studies and functional analysis in vivo.


Assuntos
Biomarcadores/metabolismo , Estradiol/farmacologia , Tubas Uterinas/fisiologia , Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Fator de Crescimento Insulin-Like I/fisiologia , Oviductos/fisiologia , Análise de Célula Única/métodos , Animais , Estrogênios/farmacologia , Tubas Uterinas/citologia , Tubas Uterinas/efeitos dos fármacos , Feminino , Perfilação da Expressão Gênica , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Oviductos/citologia , Oviductos/efeitos dos fármacos , Receptores de Progesterona/fisiologia
4.
J Biol Chem ; 295(25): 8387-8400, 2020 06 19.
Artigo em Inglês | MEDLINE | ID: mdl-32354741

RESUMO

Estrogen receptor α (ERα) modulates gene expression by interacting with chromatin regions that are frequently distal from the promoters of estrogen-regulated genes. Active chromatin-enriched "super-enhancer" (SE) regions, mainly observed in in vitro culture systems, often control production of key cell type-determining transcription factors. Here, we defined super-enhancers that bind to ERα in vivo within hormone-responsive uterine tissue in mice. We found that SEs are already formed prior to estrogen exposure at the onset of puberty. The genes at SEs encoded critical developmental factors, including retinoic acid receptor α (RARA) and homeobox D (HOXD). Using high-throughput chromosome conformation capture (Hi-C) along with DNA sequence analysis, we demonstrate that most SEs are located at a chromatin loop end and that most uterine genes in loop ends associated with these SEs are regulated by estrogen. Although the SEs were formed before puberty, SE-associated genes acquired optimal ERα-dependent expression after reproductive maturity, indicating that pubertal processes that occur after SE assembly and ERα binding are needed for gene responses. Genes associated with these SEs affected key estrogen-mediated uterine functions, including transforming growth factor ß (TGFß) and LIF interleukin-6 family cytokine (LIF) signaling pathways. To the best of our knowledge, this is the first identification of SE interactions that underlie hormonal regulation of genes in uterine tissue and optimal development of estrogen responses in this tissue.


Assuntos
Cromatina/metabolismo , Receptor alfa de Estrogênio/metabolismo , Útero/metabolismo , Animais , Sítios de Ligação , Cromatina/química , Estradiol/farmacologia , Receptor alfa de Estrogênio/deficiência , Receptor alfa de Estrogênio/genética , Feminino , Histonas/metabolismo , Proteínas de Homeodomínio/metabolismo , Fator Inibidor de Leucemia/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Ligação Proteica , Receptor alfa de Ácido Retinoico/metabolismo , Transdução de Sinais , Fator de Crescimento Transformador beta/metabolismo , Útero/efeitos dos fármacos
5.
EMBO J ; 36(9): 1199-1214, 2017 05 02.
Artigo em Inglês | MEDLINE | ID: mdl-28314781

RESUMO

Control of energy homeostasis and metabolism is achieved by integrating numerous pathways, and miRNAs are involved in this process by regulating expression of multiple target genes. However, relatively little is known about the posttranscriptional processing of miRNAs and a potential role for the precursors they derive from. Here, we demonstrate that mature miRNA-22 is more abundant in muscle from male mice relative to females and that this enables sex-specific regulation of muscular lipid metabolism and body weight by repressing estrogen receptor alpha (ERα) expression. We found that the ERα adjusts its own activity by preventing processing of miR-22 via direct binding to a conserved ERα-binding element within the primary miR-22 precursor. Mutation of the ERα binding site within the pri-miR-22 in vivo eliminates sex-specific differences in miR-22 expression. We reason that the resulting tissue selective negative feedback regulation is essential to establish sex-specific differences in muscle metabolism and body weight development.


Assuntos
Receptor alfa de Estrogênio/biossíntese , Regulação da Expressão Gênica , Metabolismo dos Lipídeos , MicroRNAs/metabolismo , Músculos/metabolismo , Animais , Masculino , Camundongos , Fatores Sexuais
6.
FASEB J ; 34(12): 16003-16021, 2020 12.
Artigo em Inglês | MEDLINE | ID: mdl-33064339

RESUMO

Estrogen receptor alpha (ERα) is a ligand-dependent transcription regulator, containing two transactivation functional domains, AF-1 and AF-2. The selective estrogen receptor modulators (SERMs), including 4-hydroxytamoxifen (4OHT), activate AF-1 preferentially rather than AF-2. However, it is unclear whether this specific function is related to the tissue-selective functionality of SERMs. Moreover, there is no information determining AF-1-dependent estrogenic-genes existing in tissues. We sought to identify AF-1-dependent estrogenic-genes using the AF-2 mutated knock-in (KI) mouse model, AF2ERKI. AF2ER is an AF-2 disrupted estradiol (E2)-insensitive mutant ERα, but AF-1-dependent transcription can be activated by the estrogen-antagonists, fulvestrant (ICI) and 4OHT. Gene profiling and ChIP-Seq analysis identified Klk1b21 as an ICI-inducible gene in AF2ERKI uterus. The regulatory activity was analyzed further using a cell-based reporter assay. The 5'-flanking 0.4k bp region of Klk1b21 gene responded as an ERα AF-1-dependent estrogen-responsive promoter. The 150 bp minimum ERα binding element (EBE) consists of three direct repeats. These three half-site sequences were essential for the ERα-dependent transactivation and were differentially recognized by E2 and 4OHT for the gene activation. This response was impaired when the minimum EBE was fused with a thymidine-kinase promoter but could be restored by fusion with the 100 bp minimum transcription initiation element (TIE) of Klk1b21, suggesting that the cooperative function of EBE and TIE is essential for mediating AF-1-dependent transactivation. These findings provide the first in vivo evidence that endogenous ERα AF-1 dominant estrogenic-genes exist in estrogen-responsive organs. Such findings will aid in understanding the mechanism of ERα-dependent tissue-selective activity of SERMs.


Assuntos
Receptor alfa de Estrogênio/genética , Ativação Transcricional/genética , Animais , Linhagem Celular Tumoral , Estradiol/genética , Antagonistas de Estrogênios/farmacologia , Estrogênios/genética , Feminino , Fulvestranto/farmacologia , Células Hep G2 , Humanos , Ligantes , Camundongos , Modelos Animais , Regiões Promotoras Genéticas/efeitos dos fármacos , Regiões Promotoras Genéticas/genética , Ligação Proteica/genética , Moduladores Seletivos de Receptor Estrogênico/farmacologia , Sítio de Iniciação de Transcrição/efeitos dos fármacos , Ativação Transcricional/efeitos dos fármacos
7.
Proc Natl Acad Sci U S A ; 115(18): E4189-E4198, 2018 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-29666266

RESUMO

Early transient developmental exposure to an endocrine active compound, diethylstilbestrol (DES), a synthetic estrogen, causes late-stage effects in the reproductive tract of adult mice. Estrogen receptor alpha (ERα) plays a role in mediating these developmental effects. However, the developmental mechanism is not well known in male tissues. Here, we present genome-wide transcriptome and DNA methylation profiling of the seminal vesicles (SVs) during normal development and after DES exposure. ERα mediates aberrations of the mRNA transcriptome in SVs of adult mice following neonatal DES exposure. This developmental exposure impacts differential diseases between male (SVs) and female (uterus) tissues when mice reach adulthood due to most DES-altered genes that appear to be tissue specific during mouse development. Certain estrogen-responsive gene changes in SVs are cell-type specific. DNA methylation dynamically changes during development in the SVs of wild-type (WT) and ERα-knockout (αERKO) mice, which increases both the loss and gain of differentially methylated regions (DMRs). There are more gains of DMRs in αERKO compared with WT. Interestingly, the methylation changes between the two genotypes are in different genomic loci. Additionally, the expression levels of a subset of DES-altered genes are associated with their DNA methylation status following developmental DES exposure. Taken together, these findings provide an important basis for understanding the molecular and cellular mechanism of endocrine-disrupting chemicals (EDCs), such as DES, during development in the male mouse tissues. This unique evidence contributes to our understanding of developmental actions of EDCs in human health.


Assuntos
Metilação de DNA/efeitos dos fármacos , Dietilestilbestrol/efeitos adversos , Receptor alfa de Estrogênio/metabolismo , Estrogênios não Esteroides/efeitos adversos , Regulação da Expressão Gênica/efeitos dos fármacos , Glândulas Seminais/metabolismo , Transcriptoma/efeitos dos fármacos , Animais , Metilação de DNA/genética , Dietilestilbestrol/farmacologia , Receptor alfa de Estrogênio/agonistas , Receptor alfa de Estrogênio/genética , Estrogênios não Esteroides/farmacologia , Loci Gênicos , Masculino , Camundongos , Camundongos Knockout
8.
J Biol Chem ; 294(25): 9746-9759, 2019 06 21.
Artigo em Inglês | MEDLINE | ID: mdl-31073032

RESUMO

Insulin-like growth factor 1 (IGF1) is primarily synthesized in and secreted from the liver; however, estrogen (E2), through E2 receptor α (ERα), increases uterine Igf1 mRNA levels. Previous ChIP-seq analyses of the murine uterus have revealed a potential enhancer region distal from the Igf1 transcription start site (TSS) with multiple E2-dependent ERα-binding regions. Here, we show E2-dependent super enhancer-associated characteristics and suggest contact between the distal enhancer and the Igf1 TSS. We hypothesized that this distal super-enhancer region controls E2-responsive induction of uterine Igf1 transcripts. We deleted 430 bp, encompassing one of the ERα-binding sites, thereby disrupting interactions of the enhancer with gene-regulatory factors. As a result, E2-mediated induction of mouse uterine Igf1 mRNA is completely eliminated, whereas hepatic Igf1 expression remains unaffected. This highlights the central role of a distal enhancer in the assembly of the factors necessary for E2-dependent interaction with the Igf1 TSS and induction of uterus-specific Igf1 transcription. Of note, loss of the enhancer did not affect fertility or uterine growth responses. Deletion of uterine Igf1 in a PgrCre;Igf1f/f model decreased female fertility but did not impact the E2-induced uterine growth response. Moreover, E2-dependent activation of uterine IGF1 signaling was not impaired by disrupting the distal enhancer or by deleting the coding transcript. This indicated a role for systemic IGF1, suggested that other growth mediators drive uterine response to E2, and suggested that uterine-derived IGF1 is essential for reproductive success. Our findings elucidate the role of a super enhancer in Igf1 regulation and uterine growth.


Assuntos
Elementos Facilitadores Genéticos , Receptor alfa de Estrogênio/metabolismo , Estrogênios/farmacologia , Regulação da Expressão Gênica/efeitos dos fármacos , Fator de Crescimento Insulin-Like I/fisiologia , Transcrição Gênica/efeitos dos fármacos , Útero/metabolismo , Animais , Feminino , Camundongos , Camundongos Knockout , Útero/efeitos dos fármacos
9.
FASEB J ; 33(6): 7375-7386, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-30866655

RESUMO

Polycystic ovary syndrome (PCOS) is a hypothalamic-pituitary-gonadal (HPG) axis disorder. PCOS symptoms most likely result from a disturbance in the complex feedback regulation system of the HPG axis, which involves gonadotrophic hormones and ovarian steroid hormones. However, the nature of this complex and interconnecting feedback regulation makes it difficult to dissect the molecular mechanisms responsible for PCOS phenotypes. Global estrogen receptor α (ERα) knockout (KO) mice exhibit a disruption of the HPG axis, resulting in hormonal dysregulation in which female ERα KO mice have elevated levels of serum estradiol (E2), testosterone, and LH. The ERα KO females are anovulatory and develop cystic hemorrhagic ovaries that are thought to be due to persistently high circulating levels of LH from the pituitary. However, the role of ERα in the pituitary is still controversial because of the varied phenotypes reported in pituitary-specific ERα KO mouse models. Therefore, we developed a mouse model where ERα is reintroduced to be exclusively expressed in the pituitary on the background of a global ERα-null (PitERtgKO) mouse. Serum E2 and LH levels were normalized in PitERtgKO females and were comparable to wild-type serum levels. However, the ovaries of PitERtgKO adult mice displayed a more overt cystic and hemorrhagic phenotype when compared with ERα KO littermates. We determined that anomalous sporadic LH secretion caused the severe ovarian phenotype of PitERtgKO females. Our observations suggest that pituitary ERα is involved in the estrogen negative feedback regulation, whereas hypothalamic ERα is necessary for the precise control of LH secretion. Uncontrolled, irregular LH secretion may be the root cause of the cystic ovarian phenotype with similarities to PCOS.-Arao, Y., Hamilton, K. J., Wu, S.-P., Tsai, M.-J., DeMayo, F. J., Korach, K. S. Dysregulation of hypothalamic-pituitary estrogen receptor α-mediated signaling causes episodic LH secretion and cystic ovary.


Assuntos
Receptor alfa de Estrogênio/fisiologia , Hipotálamo/fisiopatologia , Hormônio Luteinizante/metabolismo , Ovário/fisiopatologia , Adeno-Hipófise/fisiopatologia , Síndrome do Ovário Policístico/fisiopatologia , Animais , Modelos Animais de Doenças , Estradiol/fisiologia , Receptor alfa de Estrogênio/deficiência , Receptor alfa de Estrogênio/genética , Retroalimentação Fisiológica , Feminino , Hemorragia/etiologia , Humanos , Sistema Hipotálamo-Hipofisário/fisiopatologia , Camundongos , Camundongos da Linhagem 129 , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Especificidade de Órgãos , Ovário/patologia , Síndrome do Ovário Policístico/sangue , Síndrome do Ovário Policístico/patologia , Proteínas Recombinantes/metabolismo
10.
FASEB J ; 33(2): 3010-3023, 2019 02.
Artigo em Inglês | MEDLINE | ID: mdl-30332301

RESUMO

Pausing of RNA polymerase II (Pol II) during early transcription, mediated by the negative elongation factor (NELF) complex, allows cells to coordinate and appropriately respond to signals by modulating the rate of transcriptional pause release. Promoter proximal enrichment of Pol II occurs at uterine genes relevant to reproductive biology; thus, we hypothesized that pausing might impact endometrial response by coordinating hormonal signals involved in establishing and maintaining pregnancy. We deleted the NELF-B subunit in the mouse uterus using PgrCre (NELF-B UtcKO). Resulting females were infertile. Uterine response to the initial decidual stimulus of NELF-B UtcKO was similar to that of control mice; however, subsequent full decidual response was not observed. Cultured NELF-B UtcKO stromal cells exhibited perturbances in extracellular matrix components and also expressed elevated levels of the decidual prolactin Prl8a2, as well as altered levels of transcripts encoding enzymes involved in prostaglandin synthesis and metabolism. Because endometrial stromal cell decidualization is also critical to human reproductive health and fertility, we used small interfering to suppress NELF-B or NELF-E subunits in cultured human endometrial stromal cells, which inhibited decidualization, as reflected by the impaired induction of decidual markers PRL and IGFBP1. Overall, our study indicates NELF-mediated pausing is essential to coordinate endometrial responses and that disruption impairs uterine decidual development during pregnancy.-Hewitt, S. C., Li, R., Adams, N., Winuthayanon, W., Hamilton, K. J., Donoghue, L. J., Lierz, S. L., Garcia, M., Lydon, J. P., DeMayo, F. J., Adelman, K., Korach, K. S. Negative elongation factor is essential for endometrial function.


Assuntos
Células-Tronco Embrionárias/fisiologia , Endométrio/fisiologia , Infertilidade Feminina/fisiopatologia , Células Estromais/fisiologia , Fatores de Transcrição/fisiologia , Animais , Decídua/citologia , Decídua/fisiologia , Células-Tronco Embrionárias/citologia , Endométrio/citologia , Feminino , Voluntários Saudáveis , Humanos , Camundongos , Camundongos Knockout , Gravidez , Células Estromais/citologia , Útero/citologia , Útero/fisiologia
11.
Am J Respir Crit Care Med ; 200(10): 1246-1257, 2019 11 15.
Artigo em Inglês | MEDLINE | ID: mdl-31291549

RESUMO

Rationale: The relevance of hormones in idiopathic pulmonary fibrosis (IPF), a predominantly male lung disease, is unknown.Objectives: To determine whether the ER (estrogen receptor) facilitates the development of pulmonary fibrosis and is mediated in part through microRNA regulation of ERα and ERα-activated profibrotic pathways.Methods: ER expression in male lung tissue and myofibroblasts from control subjects (n = 6) and patients with IPF (n = 6), aging bleomycin (BLM)-treated mice (n = 7), and BLM-treated AF2ERKI mice (n = 7) was determined. MicroRNAs that regulate ER and fibrotic pathways were assessed. Transfections with a reporter plasmid containing the 3' untranslated region of the gene encoding ERα (ESR1) with and without miRNA let-7 mimics or inhibitors or an estrogen response element-driven reporter construct (ERE) construct were conducted.Measurements and Main Results: ERα expression increased in IPF lung tissue, myofibroblasts, or BLM mice. In vitro treatment with let-7 mimic transfections in human myofibroblasts reduced ERα expression and associated fibrotic pathways. AF2ERKI mice developed BLM-induced lung fibrosis, suggesting a role for growth factors in stimulating ER and fibrosis. IGF-1 (insulin-like growth factor 1) expression was increased and induced a fourfold increase of an ERE construct.Conclusions: Our data show 1) a critical role for ER and let-7 in lung fibrosis, and 2) that IGF may stimulate ER in an E2-independent manner. These results underscore the role of sex steroid hormones and their receptors in diseases that demonstrate a sex prevalence, such as IPF.


Assuntos
Fibrose Pulmonar Idiopática/etiologia , Fibrose Pulmonar Idiopática/metabolismo , MicroRNAs/fisiologia , Receptores de Estrogênio/metabolismo , Animais , Estudos de Casos e Controles , Humanos , Fibrose Pulmonar Idiopática/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Miofibroblastos/metabolismo , Técnicas de Cultura de Tecidos
12.
Yale J Biol Med ; 93(2): 291-305, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32607090

RESUMO

Essential oils (EOs) have risen in popularity over the past decade. These oils function in society as holistic integrative modalities to traditional medicinal treatments, where many Americans substitute EOs in place of other prescribed medications. EOs are found in a multitude of products including food flavoring, soaps, lotions, shampoos, hair styling products, cologne, laundry detergents, and even insect repellents. EOs are complex substances comprised of hundreds of components that can vary greatly in their composition depending upon the extraction process by the producer or the origin of the plant. Thus, making it difficult to determine which pathways in the body are affected. Here, we review the published research that shows the health benefits of EOs as well as some of their adverse effects. In doing so, we show that EOs, as well as some of their individual components, possess antimicrobial, antiviral, antibiotic, anti-inflammatory, and antioxidant properties as well as purported psychogenic effects such as relieving stress, treating depression, and aiding with insomnia. Not only do we show the health benefits of using EOs, but we also indicate risks associated with their use such as their endocrine disrupting properties leading to the induction of premature breast growth in young adolescents. Taken together, there are many positive and potentially negative risks to human health associated with EOs, which make it important to bring awareness to all their known effects on the human body.


Assuntos
Aromaterapia/métodos , Óleos Voláteis , Humanos , Medicina Tradicional/métodos , Óleos Voláteis/efeitos adversos , Óleos Voláteis/farmacologia , Medição de Risco
13.
J Biol Chem ; 293(22): 8495-8507, 2018 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-29632071

RESUMO

Estrogen receptor α (ERα) is a major transducer of estrogen-mediated physiological signals. ERα is a member of the nuclear receptor superfamily, which encompasses ligand-dependent transcription factors. The C terminus of nuclear receptors, termed the F domain, is the least homologous region among the members of this family. The ERα F domain possesses 45 amino acids; however, its function remains unclear. We noticed that the homology of the F domains between mouse and human ERαs is remarkably lower (75.6% similarity) than that between the entire proteins (94.7% similarity). To assess the functionality of the ERα F domains, here we generated chimeric ERα expression constructs with mouse-human-exchanged F domains. Using cell-based in vitro assays, we analyzed the transcriptional coactivator interaction and ligand-binding domain dimerization activities of these mouse-human F domain-swapped ERαs. We found that the transcriptional activity of the mouse WT ERα is more potent than that of the human WT ERα in the human hepatoma cell line HepG2. 4-Hydroxytamoxifen (4OHT)-mediated transcriptional activity of mouse-human F domain-swapped ERαs was the inverse of the WT ERα activities but not estradiol-mediated transcriptional activities. Further experiments with constructs containing deletion or point mutations of a predicted ß-strand region within the F domain suggested that this region governs the species-specific 4OHT-mediated transcriptional activity of ERα. We conclude that the ERα F domain has a species-specific function in 4OHT-mediated receptor transactivation and that mouse-human F domain-swapped ERα mutants enable key insights into ERα F domain structure and function.


Assuntos
Carcinoma Hepatocelular/genética , Antagonistas de Estrogênios/farmacologia , Receptor alfa de Estrogênio/metabolismo , Neoplasias Hepáticas/genética , Tamoxifeno/farmacologia , Ativação Transcricional/efeitos dos fármacos , Sequência de Aminoácidos , Animais , Sítios de Ligação , Carcinoma Hepatocelular/tratamento farmacológico , Carcinoma Hepatocelular/patologia , Receptor alfa de Estrogênio/química , Receptor alfa de Estrogênio/genética , Regulação Neoplásica da Expressão Gênica , Humanos , Neoplasias Hepáticas/tratamento farmacológico , Neoplasias Hepáticas/patologia , Camundongos , Ligação Proteica , Conformação Proteica , Domínios Proteicos , Homologia de Sequência , Especificidade da Espécie , Células Tumorais Cultivadas
14.
J Biol Chem ; 293(13): 4735-4751, 2018 03 30.
Artigo em Inglês | MEDLINE | ID: mdl-29378845

RESUMO

Estrogen receptor α (ERα) action plays an important role in pancreatic ß-cell function and survival; thus, it is considered a potential therapeutic target for the treatment of type 2 diabetes in women. However, the mechanisms underlying the protective effects of ERα remain unclear. Because ERα regulates mitochondrial metabolism in other cell types, we hypothesized that ERα may act to preserve insulin secretion and promote ß-cell survival by regulating mitochondrial-endoplasmic reticulum (EndoRetic) function. We tested this hypothesis using pancreatic islet-specific ERα knockout (PERαKO) mice and Min6 ß-cells in culture with Esr1 knockdown (KD). We found that Esr1-KD promoted reactive oxygen species production that associated with reduced fission/fusion dynamics and impaired mitophagy. Electron microscopy showed mitochondrial enlargement and a pro-fusion phenotype. Mitochondrial cristae and endoplasmic reticulum were dilated in Esr1-KD compared with ERα replete Min6 ß-cells. Increased expression of Oma1 and Chop was paralleled by increased oxygen consumption and apoptosis susceptibility in ERα-KD cells. In contrast, ERα overexpression and ligand activation reduced both Chop and Oma1 expression, likely by ERα binding to consensus estrogen-response element sites in the Oma1 and Chop promoters. Together, our findings suggest that ERα promotes ß-cell survival and insulin secretion through maintenance of mitochondrial fission/fusion-mitophagy dynamics and EndoRetic function, in part by Oma1 and Chop repression.


Assuntos
Apoptose , Estresse do Retículo Endoplasmático , Receptor alfa de Estrogênio/metabolismo , Células Secretoras de Insulina/metabolismo , Mitocôndrias/metabolismo , Mitofagia , Animais , Sobrevivência Celular , Receptor alfa de Estrogênio/genética , Feminino , Insulina/genética , Insulina/metabolismo , Metaloproteases/biossíntese , Metaloproteases/genética , Camundongos , Camundongos Knockout , Mitocôndrias/genética , Proteínas Mitocondriais/biossíntese , Proteínas Mitocondriais/genética , Espécies Reativas de Oxigênio/metabolismo , Fator de Transcrição CHOP/biossíntese , Fator de Transcrição CHOP/genética
15.
Int J Mol Sci ; 20(15)2019 Jul 30.
Artigo em Inglês | MEDLINE | ID: mdl-31366023

RESUMO

The isolation of estrogen receptor alpha (ERα) cDNA was successful around 30 years ago. The characteristics of ERα protein have been examined from various aspects, primarily through in vitro cell culture studies, but more recently using in vivo experimental models. There remains, however, some uncharacterized ERα functionalities. In particular, the mechanism of partial agonist activity of selective estrogen receptor modulators (SERMs) that involves control of the N-terminal transcription function of ERα, termed AF-1, is still an unsolved ERα functionality. We review the possible mechanism of SERM-dependent regulation of ERα AF-1-mediated transcriptional activity, which includes the role of helix 12 of ERα ligand binding domain (LBD) for SERM-dependent AF-1 regulation. In addition, we describe a specific portion of the LBD that associates with blocking AF-1 activity with an additional role of the F-domain in mediating SERM activity.


Assuntos
Receptor alfa de Estrogênio/metabolismo , Multimerização Proteica , Moduladores Seletivos de Receptor Estrogênico/farmacologia , Animais , Sítios de Ligação , Receptor alfa de Estrogênio/agonistas , Receptor alfa de Estrogênio/antagonistas & inibidores , Receptor alfa de Estrogênio/química , Humanos , Ativação Transcricional
16.
FASEB J ; 31(4): 1595-1607, 2017 04.
Artigo em Inglês | MEDLINE | ID: mdl-28082352

RESUMO

Newly fertilized embryos spend the first few days within the oviduct and are transported to the uterus, where they implant onto the uterine wall. An implantation of the embryo before reaching the uterus could result in ectopic pregnancy and lead to maternal death. Estrogen is necessary for embryo transport in mammals; however, the mechanism involved in estrogen-mediated cellular function within the oviduct remains unclear. In this study, we show in mouse models that ciliary length and beat frequency of the oviductal epithelial cells are regulated through estrogen receptor α (ESR1) but not estrogen receptor ß (ESR2). Gene profiling indicated that transcripts in the WNT/ß-catenin (WNT/CTNNB1) signaling pathway were regulated by estrogen in mouse oviduct, and inhibition of this pathway in a whole oviduct culture system resulted in a decreased embryo transport distance. However, selective ablation of CTNNB1 from the oviductal ciliated cells did not affect embryo transport, possibly because of a compensatory mechanism via intact CTNNB1 in the adjacent secretory cells. In summary, we demonstrated that disruption of estrogen signaling in oviductal epithelial cells alters ciliary function and impairs embryo transport. Therefore, our findings may provide a better understanding of etiology of the ectopic pregnancy that is associated with alteration of estrogen signals.-Li, S., O'Neill, S. R. S., Zhang, Y., Holtzman, M. J., Takemaru, K.-I., Korach, K. S., Winuthayanon, W. Estrogen receptor α is required for oviductal transport of embryos.


Assuntos
Implantação Tardia do Embrião , Células Epiteliais/metabolismo , Receptor alfa de Estrogênio/metabolismo , Oviductos/fisiologia , Gravidez Ectópica/metabolismo , Animais , Cílios/metabolismo , Cílios/fisiologia , Células Epiteliais/fisiologia , Receptor alfa de Estrogênio/genética , Receptor beta de Estrogênio/genética , Receptor beta de Estrogênio/metabolismo , Feminino , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Oviductos/citologia , Oviductos/metabolismo , Gravidez , Gravidez Ectópica/genética , Proteínas Wnt/genética , Proteínas Wnt/metabolismo , beta Catenina/genética , beta Catenina/metabolismo
17.
J Biol Chem ; 290(28): 17611-27, 2015 Jul 10.
Artigo em Inglês | MEDLINE | ID: mdl-26028650

RESUMO

ERα has a ligand-dependent transactivation function in the ligand binding domain of ERα C terminus (AF-2) and a ligand-independent activation function in the N terminus (AF-1). It is still not fully understood how AF-1 and AF-2 activities are regulated cooperatively by ligands. To evaluate the AF-1 involvement in the estrogenic activities of various compounds, we analyzed these transactivation functions using AF-1-truncated and AF-2-mutated ERα mutants. AF-2 is composed of two domains with flexible and static regions. We used an AF-2 flexible region mutant and an AF-2 static region mutant. Both mutants have been reported as non-E2 responsive due to disruption of E2-mediated coactivator recruitment to the AF-2. The AF-2 mutants were not activated by agonists, but surprisingly antagonists and selective estrogen receptor modulators (SERMs) activated the AF-2 mutants. This antagonist reversal activity was derived from AF-1. Furthermore, we demonstrated that the AF-2 contains an AF-1 suppression function using C-terminal-truncated ERα mutants. From these findings we hypothesized that the mutation of AF-2 disrupted its ability to suppress AF-1, causing the antagonist reversal. To assess the AF-2-mediated AF-1 suppression, we analyzed the transcription activity of physically separated AF-1 and AF-2 using a novel hybrid reporter assay. We observed that the AF-1 activity was not suppressed by the physically separated AF-2. Furthermore, SERMs did not induce the AF-1-mediated activity from the separated mutant AF-2, which differed from the intact protein. These results imply that SERM activity is dependent on a conformational change of the full-length ERα molecule, which allows for AF-1 activation.


Assuntos
Receptor alfa de Estrogênio/química , Receptor alfa de Estrogênio/metabolismo , Animais , Disruptores Endócrinos/farmacologia , Receptor alfa de Estrogênio/genética , Células Hep G2 , Humanos , Ligantes , Camundongos , Mutagênese Sítio-Dirigida , Proteínas Mutantes/química , Proteínas Mutantes/genética , Proteínas Mutantes/metabolismo , Fitoestrógenos/farmacologia , Conformação Proteica , Domínios e Motivos de Interação entre Proteínas , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Moduladores Seletivos de Receptor Estrogênico/farmacologia , Deleção de Sequência , Transativadores/química , Transativadores/genética , Transativadores/metabolismo
18.
J Biol Chem ; 290(9): 5566-81, 2015 Feb 27.
Artigo em Inglês | MEDLINE | ID: mdl-25468909

RESUMO

Obesity is associated with increased breast cancer (BrCA) incidence. Considering that inactivation of estrogen receptor (ER)α promotes obesity and metabolic dysfunction in women and female mice, understanding the mechanisms and tissue-specific sites of ERα action to combat metabolic-related disease, including BrCA, is of clinical importance. To study the role of ERα in adipose tissue we generated fat-specific ERα knock-out (FERKO) mice. Herein we show that ERα deletion increased adipocyte size, fat pad weight, and tissue expression and circulating levels of the secreted glycoprotein, lipocalin 2 (Lcn2), an adipokine previously associated with BrCA development. Chromatin immunoprecipitation and luciferase reporter studies showed that ERα binds the Lcn2 promoter to repress its expression. Because adipocytes constitute an important cell type of the breast microenvironment, we examined the impact of adipocyte ERα deletion on cancer cell behavior. Conditioned medium from ERα-null adipocytes and medium containing pure Lcn2 increased proliferation and migration of a subset of BrCA cells in culture. The proliferative and promigratory effects of ERα-deficient adipocyte-conditioned medium on BrCA cells was reversed by Lcn2 deletion. BrCA cell responsiveness to exogenous Lcn2 was heightened in cell types where endogenous Lcn2 expression was minimal, but components of the Lcn2 signaling pathway were enriched, i.e. SLC22A17 and 3-hydroxybutyrate dehydrogenase (BDH2). In breast tumor biopsies from women diagnosed with BrCA we found that BDH2 expression was positively associated with adiposity and circulating Lcn2 levels. Collectively these data suggest that reduction of ERα expression in adipose tissue promotes adiposity and is linked with the progression and severity of BrCA via increased adipocyte-specific Lcn2 production and enhanced tumor cell Lcn2 sensitivity.


Assuntos
Proteínas de Fase Aguda/metabolismo , Tecido Adiposo/metabolismo , Receptor alfa de Estrogênio/metabolismo , Lipocalinas/metabolismo , Obesidade/metabolismo , Proteínas Oncogênicas/metabolismo , Células 3T3-L1 , Proteínas de Fase Aguda/genética , Adipócitos/citologia , Adipócitos/metabolismo , Tecido Adiposo/citologia , Animais , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Progressão da Doença , Receptor alfa de Estrogênio/genética , Feminino , Perfilação da Expressão Gênica , Células HEK293 , Humanos , Immunoblotting , Lipocalina-2 , Lipocalinas/sangue , Lipocalinas/genética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Obesos , Obesidade/genética , Proteínas Oncogênicas/sangue , Proteínas Oncogênicas/genética , Regiões Promotoras Genéticas/genética , Ligação Proteica , Reação em Cadeia da Polimerase Via Transcriptase Reversa
19.
FASEB J ; 28(12): 5042-54, 2014 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-25212221

RESUMO

The estrogens are female sex hormones that are involved in a variety of physiological processes, including reproductive development and function, wound healing, and bone growth. They are mainly known for their roles in reproductive tissues--specifically, 17ß-estradiol (E2), the primary estrogen, which is secreted by the ovaries and induces cellular proliferation and growth of the uterus and mammary glands. In addition to the role of estrogens in promoting tissue growth and development during normal physiological states, they have a well-established role in determining susceptibility to disease, particularly cancer, in reproductive tissues. The responsiveness of various tissues to estrogen is genetically controlled, with marked quantitative variation observed across multiple species, including humans. This variation presents both researchers and clinicians with a veritable physiological puzzle, the pieces of which--many of them unknown--are complex and difficult to fit together. Although genetics is known to play a major role in determining sensitivity to estrogens, there are other factors, including parent of origin and the maternal environment, that are intimately linked to heritable phenotypes but do not represent genotype, per se. The objectives of this review article were to summarize the current knowledge of the role of genotype, and uterine and neonatal environments, in phenotypic variation in the response to estrogens; to discuss recent findings and the potential mechanisms involved; and to highlight exciting research opportunities for the future.


Assuntos
Estrogênios/fisiologia , Animais , Feminino , Humanos , Glândulas Mamárias Humanas/fisiologia , Útero/fisiologia , Vagina/fisiologia
20.
Proc Natl Acad Sci U S A ; 109(51): 21140-5, 2012 Dec 18.
Artigo em Inglês | MEDLINE | ID: mdl-23213263

RESUMO

Estrogen receptor alpha (ERα) is a ligand-dependent transcription factor containing two transcriptional activation function (AF) domains. AF-1 is in the N terminus of the receptor protein, and AF-2 activity is dependent on helix 12 of the C-terminal ligand-binding domain. We recently showed that two point mutations converting leucines 543 and 544 to alanines in helix 12 (AF2ER) minimized estrogen-dependent AF-2 transcriptional activation. A characteristic feature of AF2ER is that the estrogen antagonists ICI182780 and tamoxifen (TAM) act as agonists through intact AF-1, but not through mutated AF-2. Here we report the reproductive phenotype of male AF2ER knock-in (AF2ERKI) mice and demonstrate the involvement of ERα in male fertility. The AF2ERKI male homozygotes are infertile because of seminiferous tubular dysmorphogenesis in the testis, similar to ERα KO males. Sperm counts and motility did not differ at age 6 wk in AF2ERKI and WT mice, but a significant testis defect was observed in adult AF2ERKI male mice. The expression of efferent ductal genes involved in fluid reabsorption was significantly lower in AF2ERKI males. TAM treatment for 3 wk beginning at age 21 d activated AF-2-mutated ERα (AF2ER) and restored expression of efferent ductule genes. At the same time, the TAM treatment reversed AF2ERKI male infertility compared with the vehicle-treated group. These results indicate that the ERα AF-2 mutation results in male infertility, suggesting that the AF-1 is regulated in an AF-2-dependent manner in the male reproductive tract. Activation of ERα AF-1 is capable of rescuing AF2ERKI male infertility.


Assuntos
Receptor alfa de Estrogênio/metabolismo , Animais , Estradiol/análogos & derivados , Estradiol/farmacologia , Receptor alfa de Estrogênio/química , Fertilidade , Fulvestranto , Homozigoto , Ligantes , Masculino , Camundongos , Camundongos Knockout , Mutação Puntual , Estrutura Terciária de Proteína , Reprodução , Contagem de Espermatozoides , Motilidade dos Espermatozoides , Tamoxifeno/farmacologia , Testículo/metabolismo , Ativação Transcricional
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