RESUMO
Family support plays an important role in promoting resilience and health among transgender and/or nonbinary youth (TNBY), but family members often experience barriers to supporting their TNBY, including minority-adjacent stress stemming from exposure to structural stigma and antitransgender legislation. TNBY and their families need effective family-level interventions developed using community-based participatory research (CBPR), which integrates community members (e.g., TNBY, family members, service providers for families with TNBY) into the intervention development process to ensure the resulting intervention is relevant and useful. Informed by findings from the Trans Teen and Family Narratives Project, we used CBPR to develop the Trans Teen and Family Narratives Conversation Toolkit, a family-level intervention designed to educate families about TNBY and facilitate conversations about gender. The toolkit was developed across 1.5 years (June 2019 to January 2021) using four integrated phases: (1) content development: digital storytelling workshop with TNBY; (2) content review: digital storyteller interviews and user focus groups; (3) content development: study team content synthesis and website development; and (4) content review: website review by TNBY, family members, and mental health providers, and intervention refinement. This article outlines the intervention development process, describes strategies employed to navigate challenges encountered along the way, and shares key learnings to inform future CBPR intervention development efforts. (PsycInfo Database Record (c) 2024 APA, all rights reserved).
RESUMO
There has been an increase in the number of youth in the United States who identify as transgender and/or as nonbinary, which generally refers to people whose gender identity does not align with the one associated with their assigned sex at birth. Unfortunately, transgender and gender diverse (TGD) youth have higher rates of depression, suicidality, self-harm, and disordered eating compared to cisgender peers.1 It is critical to emphasize, however, that such health disparities are reflective of TGD people having to simultaneously navigate transphobic, homophobic, and racist social contexts; manage gender dysphoria, which is the distress resulting from an incongruence between gender identity and assigned sex at birth; and endure an onslaught of anti-transgender legislation that attacks the rights and bodies of TGD people. Given that TGD youth have had alarmingly high rates of suicide for the past decade, it is imperative that health care providers identify and increase access to opportunities and activities that promote positive socioemotional development to mitigate such risk. One resource that has the potential to boost mental health outcomes for this community is physical activity and sports (PA/S).
Assuntos
Comportamento Autodestrutivo , Pessoas Transgênero , Recém-Nascido , Humanos , Masculino , Feminino , Adolescente , Estados Unidos , Identidade de Gênero , Saúde Mental , Pessoas Transgênero/psicologia , Ideação SuicidaRESUMO
Few studies have disentangled differences in victimization exposures and mental health symptoms among gender diverse subgroups, nor considered the role of potential protective factors in ameliorating the impact of victimization on gender diverse youths' mental health. Here we report findings from a secondary data analysis, in which we address this gap by analyzing cross-sectional survey data (N = 11,264 in the final analytic sample) from a population-based survey of youth in participating school districts in a large Midwestern U.S. county. Relative to cisgender youth with gender conforming expression, transgender youth and cisgender youth with nonconforming gender expression are more likely to experience victimization and severe mental health concerns. Additionally, school-connectedness moderates the association between bias-based harassment and depression for cisgender youth with gender nonconforming expression, and family support/monitoring buffers the association of peer victimization with suicide attempts among transgender youth. Findings highlight the need to better understand factors which may confer protection among gender diverse adolescents, so that in turn appropriate supports across key contexts can be implemented.
RESUMO
The peroxisome proliferator-activated receptors (PPARs) are transcriptional regulators of glucose, lipid, and cholesterol metabolism. We report the x-ray crystal structure of the ligand binding domain of PPAR alpha (NR1C1) as a complex with the agonist ligand GW409544 and a coactivator motif from the steroid receptor coactivator 1. Through comparison of the crystal structures of the ligand binding domains of the three human PPARs, we have identified molecular determinants of subtype selectivity. A single amino acid, which is tyrosine in PPAR alpha and histidine in PPAR gamma, imparts subtype selectivity for both thiazolidinedione and nonthiazolidinedione ligands. The availability of high-resolution cocrystal structures of the three PPAR subtypes will aid the design of drugs for the treatments of metabolic and cardiovascular diseases.
Assuntos
Oxazóis/química , Receptores Citoplasmáticos e Nucleares/química , Fatores de Transcrição/química , Tirosina/análogos & derivados , Tirosina/química , Sequência de Aminoácidos , Sítios de Ligação , Cristalografia por Raios X , Humanos , Ligantes , Modelos Moleculares , Dados de Sequência Molecular , Estrutura Molecular , Estrutura Terciária de Proteína , Receptores Citoplasmáticos e Nucleares/agonistas , Fatores de Transcrição/agonistasRESUMO
Cytochromes P450 (P450s) are involved in the oxidative metabolism of a plethora of structurally unrelated compounds, including therapeutic drugs. Two orphan members of the nuclear receptor superfamily, the pregnane X receptor (PXR; NR1I2) and constitutive androstane receptor (CAR; NR1I3) have been implicated in this phenomenon. In the present study, we examined the transcriptional regulation of the human CYP2B6 gene. In primary cultures of human hepatocytes, CYP2B6 was highly inducible by a number of compounds known to be human PXR ligands, including rifampicin and hyperforin. PXR was shown to be capable of activating the phenobarbital-responsive enhancer module (PBREM) region of the CYP2B6 gene, a 51-base-pair enhancer element that mediates induction of CYP2B6 expression by CAR. The two nuclear receptor-binding motifs within the PBREM effectively bound PXR as a heterodimer with the 9-cis retinoic acid receptor alpha (NR2B1). Taken together, these observations demonstrate that the CYP2B6 gene is directly regulated by PXR and further establish this receptor as a key regulator of drug-metabolizing P450s.
Assuntos
Hidrocarboneto de Aril Hidroxilases , Sistema Enzimático do Citocromo P-450/genética , Regulação Enzimológica da Expressão Gênica , Hepatócitos/enzimologia , Oxirredutases N-Desmetilantes/genética , Receptores Citoplasmáticos e Nucleares/fisiologia , Receptores de Esteroides/fisiologia , Motivos de Aminoácidos , Núcleo Celular/fisiologia , Células Cultivadas , Receptor Constitutivo de Androstano , Citocromo P-450 CYP2B6 , Sistema Enzimático do Citocromo P-450/biossíntese , Dimerização , Indução Enzimática , Humanos , Oxirredutases N-Desmetilantes/biossíntese , Receptor de Pregnano X , RNA Mensageiro/biossíntese , RNA Mensageiro/efeitos dos fármacos , Receptores Citoplasmáticos e Nucleares/química , Receptores Citoplasmáticos e Nucleares/genética , Receptores Citoplasmáticos e Nucleares/metabolismo , Receptores do Ácido Retinoico/genética , Receptores do Ácido Retinoico/metabolismo , Receptores de Esteroides/química , Receptores de Esteroides/genética , Receptores de Esteroides/metabolismo , Receptores X de Retinoides , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismoRESUMO
The human nuclear pregnane X receptor (hPXR) activates cytochrome P450-3A expression in response to a wide variety of xenobiotics and plays a critical role in mediating dangerous drug-drug interactions. We present the crystal structures of the ligand-binding domain of hPXR both alone and in complex with the cholesterol-lowering drug SR12813 at resolutions of 2.5 and 2.75 angstroms, respectively. The hydrophobic ligand-binding cavity of hPXR contains a small number of polar residues, permitting SR12813 to bind in three distinct orientations. The position and nature of these polar residues were found to be critical for establishing the precise pharmacologic activation profile of PXR. Our findings provide important insights into how hPXR detects xenobiotics and may prove useful in predicting and avoiding drug-drug interactions.
Assuntos
Difosfonatos/metabolismo , Receptores Citoplasmáticos e Nucleares/química , Receptores Citoplasmáticos e Nucleares/metabolismo , Receptores de Esteroides/química , Receptores de Esteroides/metabolismo , Xenobióticos/metabolismo , Sequência de Aminoácidos , Sítios de Ligação , Cristalografia por Raios X , Difosfonatos/química , Humanos , Ligantes , Modelos Moleculares , Dados de Sequência Molecular , Receptor de Pregnano X , Conformação Proteica , Estrutura Secundária de Proteína , Estrutura Terciária de Proteína , Rifampina/metabolismoRESUMO
A cell-free assay was developed for the orphan nuclear receptor LXRalpha that measures the ligand-dependent recruitment of a peptide from the steroid receptor coactivator 1 (SRC1) to the nuclear receptor. Using this ligand-sensing assay (LiSA), the structural requirements for activation of the receptor by oxysterols and related compounds were studied. The minimal pharmacophore for receptor activation was shown to be a sterol with a hydrogen bond acceptor at C24. 24(S),25-Epoxycholesterol (1), which meets this criterion, is among the most efficacious of the oxysterols and is an attractive candidate as the LXRalpha natural hormone. Cholenic acid dimethylamide (14) showed increased efficacy compared to 1, whereas the unnatural oxysterol 22(S)-hydroxycholesterol (4) was shown to be an antagonist of 1 in the LiSA. The structural requirements for SRC1 recruitment in the LiSA correlated with the transcriptional activity of compounds in a cell-based reporter assay employing LXRalpha-GAL4 chimeric receptors. Site-directed mutagenesis identified Trp(443) as an amino acid critical for activation of LXRalpha by oxysterol ligands. This information was combined with the structure-activity relationship developed from the LiSA to develop a 3D homology model of LXRalpha. This model may aid the design of synthetic drugs targeted at this transcriptional regulator of cholesterol homeostasis.
Assuntos
Colesterol/análogos & derivados , Receptores Citoplasmáticos e Nucleares/agonistas , Receptores de Esteroides/agonistas , Esteróis/farmacologia , Sequência de Aminoácidos , Animais , Sítios de Ligação , Western Blotting , Linhagem Celular , Núcleo Celular/metabolismo , Sistema Livre de Células , Chlorocebus aethiops , Colesterol/síntese química , Colesterol/química , Colesterol/farmacologia , Ácidos Cólicos/síntese química , Ácidos Cólicos/química , Ácidos Cólicos/farmacologia , Proteínas de Ligação a DNA , Transferência de Energia , Fluorescência , Histona Acetiltransferases , Hidroxicolesteróis/síntese química , Hidroxicolesteróis/química , Hidroxicolesteróis/farmacologia , Cetocolesteróis/síntese química , Cetocolesteróis/química , Cetocolesteróis/farmacologia , Receptores X do Fígado , Modelos Moleculares , Dados de Sequência Molecular , Coativador 1 de Receptor Nuclear , Receptores Nucleares Órfãos , Receptores Citoplasmáticos e Nucleares/antagonistas & inibidores , Receptores de Esteroides/antagonistas & inibidores , Estereoisomerismo , Esteróis/síntese química , Esteróis/química , Relação Estrutura-Atividade , Fatores de Transcrição/metabolismo , Triptofano/químicaRESUMO
Bile acids repress the transcription of cytochrome P450 7A1 (CYP7A1), which catalyzes the rate-limiting step in bile acid biosynthesis. Although bile acids activate the farnesoid X receptor (FXR), the mechanism underlying bile acid-mediated repression of CYP7A1 remained unclear. We have used a potent, nonsteroidal FXR ligand to show that FXR induces expression of small heterodimer partner 1 (SHP-1), an atypical member of the nuclear receptor family that lacks a DNA-binding domain. SHP-1 represses expression of CYP7A1 by inhibiting the activity of liver receptor homolog 1 (LRH-1), an orphan nuclear receptor that is known to regulate CYP7A1 expression positively. This bile acid-activated regulatory cascade provides a molecular basis for the coordinate suppression of CYP7A1 and other genes involved in bile acid biosynthesis.
Assuntos
Ácidos e Sais Biliares/biossíntese , Proteínas de Ligação a DNA/metabolismo , Proteínas Tirosina Fosfatases/metabolismo , Receptores Citoplasmáticos e Nucleares/metabolismo , Fatores de Transcrição/metabolismo , Animais , Northern Blotting , Células Cultivadas , Colesterol 7-alfa-Hidroxilase/genética , Colesterol 7-alfa-Hidroxilase/metabolismo , Proteínas de Ligação a DNA/genética , Regulação Enzimológica da Expressão Gênica/fisiologia , Hepatócitos/citologia , Hepatócitos/enzimologia , Humanos , Peptídeos e Proteínas de Sinalização Intracelular , Masculino , Regiões Promotoras Genéticas/fisiologia , Proteína Tirosina Fosfatase não Receptora Tipo 11 , Proteína Tirosina Fosfatase não Receptora Tipo 6 , Proteínas Tirosina Fosfatases/genética , RNA Mensageiro/análise , Ratos , Ratos Endogâmicos F344 , Receptores Citoplasmáticos e Nucleares/genética , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Fatores de Transcrição/genética , TransfecçãoAssuntos
Proteínas de Ligação a DNA/agonistas , Isoxazóis/síntese química , Receptores Citoplasmáticos e Nucleares/agonistas , Fatores de Transcrição/agonistas , Animais , Disponibilidade Biológica , Linhagem Celular , Sistema Livre de Células , Técnicas de Química Combinatória , Humanos , Isoxazóis/química , Isoxazóis/farmacocinética , Isoxazóis/farmacologia , Camundongos , Ratos , Ratos Endogâmicos F344 , Transfecção , Triglicerídeos/sangueRESUMO
St. John's wort (Hypericum perforatum) is an herbal remedy used widely for the treatment of depression. Recent clinical studies demonstrate that hypericum extracts increase the metabolism of various drugs, including combined oral contraceptives, cyclosporin, and indinavir. In this report, we show that hyperforin, a constituent of St. John's wort with antidepressant activity, is a potent ligand (K(i) = 27 nM) for the pregnane X receptor, an orphan nuclear receptor that regulates expression of the cytochrome P450 (CYP) 3A4 monooxygenase. Treatment of primary human hepatocytes with hypericum extracts or hyperforin results in a marked induction of CYP3A4 expression. Because CYP3A4 is involved in the oxidative metabolism of >50% of all drugs, our findings provide a molecular mechanism for the interaction of St. John's wort with drugs and suggest that hypericum extracts are likely to interact with many more drugs than previously had been realized.
Assuntos
Hypericum/metabolismo , Fígado/efeitos dos fármacos , Fígado/metabolismo , Plantas Medicinais , Receptores Citoplasmáticos e Nucleares/agonistas , Receptores de Esteroides/agonistas , Compostos Bicíclicos com Pontes , Células Cultivadas , Citocromo P-450 CYP3A , Sistema Enzimático do Citocromo P-450/metabolismo , Interações Medicamentosas , Humanos , Ligantes , Oxigenases de Função Mista/metabolismo , Floroglucinol/análogos & derivados , Receptor de Pregnano X , Receptores Citoplasmáticos e Nucleares/metabolismo , Receptores de Esteroides/metabolismo , Terpenos/metabolismo , Terpenos/farmacologiaRESUMO
Xenobiotics induce the transcription of cytochromes P450 (CYPs) 2B and 3A through the constitutive androstane receptor (CAR; NR1I3) and pregnane X receptor (PXR; NR1I2), respectively. In this report, we have systematically compared a series of xenobiotics and natural steroids for their effects on mouse and human CAR and PXR. Our results demonstrate dual regulation of PXR and CAR by a subset of compounds that affect CYP expression. Moreover, there are marked pharmacological differences between the mouse (m) and human (h) orthologs of both CAR and PXR. For example, the planar hydrocarbon 1, 4-bis[2-(3,5-dichloropyridyl-oxy)]benzene activates mCAR and hPXR but has little or no activity on hCAR and mPXR. In contrast, the CAR deactivator androstanol activates both mouse and human PXR. Similarly, the PXR activator clotrimazole is a potent deactivator of hCAR. Using radioligand binding and fluorescence resonance energy transfer assays, we demonstrate that several of the compounds that regulate mouse and human CAR, including natural steroids, bind directly to the receptors. Our results suggest that CAR, like PXR, is a steroid receptor that is capable of recognizing structurally diverse compounds. Moreover, our findings underscore the complexity in the physiologic response to xenobiotics.
Assuntos
Hidrocarboneto de Aril Hidroxilases , Receptores Citoplasmáticos e Nucleares/metabolismo , Receptores de Esteroides/metabolismo , Esteroides/farmacologia , Fatores de Transcrição/metabolismo , Xenobióticos/farmacologia , Animais , Linhagem Celular , Clotrimazol/farmacologia , Receptor Constitutivo de Androstano , Citocromo P-450 CYP2B6 , Citocromo P-450 CYP3A , Sistema Enzimático do Citocromo P-450/genética , Humanos , Cinética , Ligantes , Camundongos , Mifepristona/farmacologia , Oxirredutases N-Desmetilantes/genética , Receptor de Pregnano X , Conformação Proteica/efeitos dos fármacos , Receptores Citoplasmáticos e Nucleares/química , Receptores de Esteroides/química , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , Esteroides/metabolismo , Fatores de Transcrição/química , Transcrição Gênica/efeitos dos fármacos , Transfecção , Xenobióticos/farmacocinéticaRESUMO
Transcription of genes encoding cytochrome P450 3A (CYP3A) monooxygenases is induced by a variety of xenobiotics and natural steroids. There are marked differences in the compounds that induce CYP3A gene expression between species. Recently, the mouse and human pregnane X receptor (PXR) were shown to be activated by compounds that induce CYP3A expression. However, most studies of CYP3A regulation have been performed using rabbit and rat hepatocytes. Here, we report the cloning and characterization of PXR from these two species. PXR is remarkably divergent between species, with the rabbit, rat, and human receptors sharing only approximately 80% amino acid identity in their ligand-binding domains. This sequence divergence is reflected by marked pharmacological differences in PXR activation profiles. For example, the macrolide antibiotic rifampicin, the antidiabetic drug troglitazone, and the hypocholesterolemic drug SR12813 are efficacious activators of the human and rabbit PXR but have little activity on the rat and mouse PXR. Conversely, pregnane 16alpha-carbonitrile is a more potent activator of the rat and mouse PXR than the human and rabbit receptor. The activities of xenobiotics in PXR activation assays correlate well with their ability to induce CYP3A expression in primary hepatocytes. Through the use of a novel scintillation proximity binding assay, we demonstrate that many of the compounds that induce CYP3A expression bind directly to human PXR. These data establish PXR as a promiscuous xenobiotic receptor that has diverged during evolution.
Assuntos
Hidrocarboneto de Aril Hidroxilases , Receptores Citoplasmáticos e Nucleares/química , Receptores de Esteroides/química , Xenobióticos/metabolismo , Sequência de Aminoácidos , Animais , Anticolesterolemiantes/farmacologia , Northern Blotting , Clonagem Molecular , Citocromo P-450 CYP3A , Sistema Enzimático do Citocromo P-450/metabolismo , Difosfonatos/farmacologia , Relação Dose-Resposta a Droga , Evolução Molecular , Humanos , Ligantes , Fígado/metabolismo , Camundongos , Dados de Sequência Molecular , Oxirredutases N-Desmetilantes/metabolismo , Receptor de Pregnano X , Ligação Proteica , Coelhos , Ratos , Receptores Citoplasmáticos e Nucleares/genética , Receptores Citoplasmáticos e Nucleares/metabolismo , Receptores de Esteroides/genética , Receptores de Esteroides/metabolismo , Proteínas Recombinantes/metabolismo , Homologia de Sequência de Aminoácidos , TransfecçãoRESUMO
An important step in differentiating the unique physiological roles of the alpha and beta forms of estrogen receptor is to determine the precise expression pattern of each of these receptors. We report the generation and characterization of a murine IgG1 monoclonal antibody (MAb), ER15.64A that is ERbeta subtype-specific and capable of recognizing full-length human ERbeta as well as all of its known protein isoforms. ER15.64A, raised against a ERbeta peptide (aa2-18)-keyhole limpet hemocyanine conjugate, reacted to the immunizing peptide and the full-length E. coli expressed ERbeta in ELISA and BIAcore assays. It also immunostained nuclei of Sf9 insect cells that were infected with an ERbeta-baculovirus. In Western analysis, ER15.64A recognized ERbeta1 and ERbeta2 proteins from a reticulocyte in vitro transcription/translation preparation. This antibody did not cross-react with recombinant ERalpha in ELISA, BIAcore, immunocytochemistry, or Western blot analysis. The specificity of ER15.64A should make this antibody a useful tool for monitoring expression of ERbeta and its isoforms at the protein level and should aid in distinguishing the pattern of ERbeta receptor expression from that of ERalpha.
Assuntos
Anticorpos Monoclonais/biossíntese , Receptores de Estrogênio/imunologia , Animais , Anticorpos Monoclonais/imunologia , Anticorpos Monoclonais/isolamento & purificação , Especificidade de Anticorpos , Reações Cruzadas , Receptor beta de Estrogênio , Feminino , Humanos , Hibridomas , Imunoglobulina G/biossíntese , Imunoglobulina G/imunologia , Imunoglobulina G/isolamento & purificação , Imuno-Histoquímica , Camundongos , Isoformas de Proteínas/imunologia , Receptores de Estrogênio/metabolismo , Transdução Genética , Células Tumorais CultivadasRESUMO
It has been shown in previous studies that a variety of estrogen receptor (ER) beta mRNA transcripts are expressed in human breast cancer cell lines and tumors. To complement the RNA expression studies, we have developed ER-beta-specific antibodies to characterize ER-beta protein expression in breast cancer cell lines and tumors. Monoclonal antibodies were made against a peptide representing the first 18 amino acids of the longest ER-beta open reading frame reported to date, and polyclonal antibodies were made against a peptide within the ER-beta B domain. By Western blot analysis, we show that ER-beta protein is expressed in all cancer cell lines tested and in three of five breast tumor samples. The breast cancer cell lines showed variation in the size of the expressed ER-beta protein. The longest form detected was consistent with the 530-amino acid, full-length ER-beta sequence. Shorter ER-beta isoforms were detected in the ER-alpha-negative MDA-MB-231 and MDA-MB-435 breast cancer cell lines, likely corresponding to previously described COOH-terminal RNA variant isoforms.
Assuntos
Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Receptores de Estrogênio/genética , Receptores de Estrogênio/metabolismo , Western Blotting , DNA Complementar/análise , Receptor beta de Estrogênio , Regulação Neoplásica da Expressão Gênica , Humanos , Osteossarcoma/metabolismo , Fenótipo , Isoformas de Proteínas , RNA Mensageiro/metabolismo , Células Tumorais CultivadasAssuntos
Butiratos/farmacologia , Proteínas de Ligação a DNA/agonistas , Hipolipemiantes/farmacologia , Metabolismo dos Lipídeos , Proteínas Nucleares/agonistas , Compostos de Fenilureia/farmacologia , Receptores Citoplasmáticos e Nucleares/agonistas , Fatores de Transcrição/agonistas , Dedos de Zinco , Animais , Fenofibrato/farmacologia , Humanos , Masculino , Camundongos , Ratos , Ratos Sprague-DawleyRESUMO
The peroxisome proliferator-activated receptors (PPARs) are nuclear hormone receptors that regulate glucose and lipid homeostasis. The PPARgamma subtype plays a central role in the regulation of adipogenesis and is the molecular target for the 2, 4-thiazolidinedione class of antidiabetic drugs. Structural studies have revealed that agonist ligands activate the PPARs through direct interactions with the C-terminal region of the ligand-binding domain, which includes the activation function 2 helix. GW0072 was identified as a high-affinity PPARgamma ligand that was a weak partial agonist of PPARgamma transactivation. X-ray crystallography revealed that GW0072 occupied the ligand-binding pocket by using different epitopes than the known PPAR agonists and did not interact with the activation function 2 helix. In cell culture, GW0072 was a potent antagonist of adipocyte differentiation. These results establish an approach to the design of PPAR ligands with modified biological activities.
Assuntos
Adipócitos/citologia , Receptores Citoplasmáticos e Nucleares/antagonistas & inibidores , Receptores Citoplasmáticos e Nucleares/química , Tiazóis/farmacologia , Tiazolidinedionas , Fatores de Transcrição/antagonistas & inibidores , Fatores de Transcrição/química , Animais , Sítios de Ligação , Diferenciação Celular/efeitos dos fármacos , Linhagem Celular , Chlorocebus aethiops , Cristalografia por Raios X , Humanos , Cinética , Ligantes , Camundongos , Modelos Moleculares , Conformação Molecular , Dados de Sequência Molecular , Proteínas Nucleares/metabolismo , Estrutura Secundária de Proteína , Receptores Citoplasmáticos e Nucleares/genética , Proteínas Recombinantes/antagonistas & inibidores , Proteínas Recombinantes/química , Rosiglitazona , Tiazóis/síntese química , Tiazóis/química , Tiazolidinas , Fatores de Transcrição/genética , TransfecçãoRESUMO
Multiple transcripts which arise from the human estrogen receptor beta (ER beta) gene have been characterized. Three full length isoforms of the hER beta gene, designated hER beta 1-3, were identified in a testis cDNA library. An additional two isoforms, designated hER beta 4 and hER beta 5, were identified by PCR amplification from testis cDNA and from the MDA-MB 435 cell line. hER beta 1 corresponds to the previously described hER beta. All five isoforms diverge at a common position within the predicted helix 10 of the ligand binding domain of hER beta, with nucleotide sequences consistent with differential exon usage. The hER beta isoform mRNAs displayed a differential pattern of expression in human tissues and in tumor cell lines when analyzed by RT-PCR. Further characterization of the three full length isoforms, hER beta 1-3, by in vitro band shift studies indicated that the isoforms were able to form DNA-binding homodimers and heterodimers with each other and with the ER alpha subtype.
Assuntos
Receptores de Estrogênio/química , Receptores de Estrogênio/genética , Processamento Alternativo , Sequência de Aminoácidos , Sequência de Bases , Clonagem Molecular , Proteínas de Ligação a DNA/metabolismo , Dimerização , Receptor beta de Estrogênio , Estrogênios/genética , Expressão Gênica , Humanos , Isomerismo , Masculino , Dados de Sequência Molecular , Especificidade de Órgãos/genética , RNA Mensageiro/biossíntese , Receptores de Estrogênio/biossíntese , Sequências Reguladoras de Ácido Nucleico , TestículoRESUMO
Accumulation of cholesterol causes both repression of genes controlling cholesterol biosynthesis and cellular uptake and induction of cholesterol 7alpha-hydroxylase, which leads to the removal of cholesterol by increased metabolism to bile acids. Here, we report that LXRalpha and LXRbeta, two orphan members of the nuclear receptor superfamily, are activated by 24(S), 25-epoxycholesterol and 24(S)-hydroxycholesterol at physiologic concentrations. In addition, we have identified an LXR response element in the promoter region of the rat cholesterol 7alpha-hydroxylase gene. Our data provide evidence for a new hormonal signaling pathway that activates transcription in response to oxysterols and suggest that LXRs play a critical role in the regulation of cholesterol homeostasis.
Assuntos
Colesterol/análogos & derivados , Hidroxicolesteróis/farmacologia , Receptores Citoplasmáticos e Nucleares/metabolismo , Animais , Sítios de Ligação , Colesterol/farmacologia , Colesterol 7-alfa-Hidroxilase/genética , Proteínas de Ligação a DNA , Relação Dose-Resposta a Droga , Receptores X do Fígado , Receptores Nucleares Órfãos , Regiões Promotoras Genéticas , RatosRESUMO
Thiazolidinedione derivatives are antidiabetic agents that increase the insulin sensitivity of target tissues in animal models of non-insulin-dependent diabetes mellitus. In vitro, thiazolidinediones promote adipocyte differentiation of preadipocyte and mesenchymal stem cell lines; however, the molecular basis for this adipogenic effect has remained unclear. Here, we report that thiazolidinediones are potent and selective activators of peroxisome proliferator-activated receptor gamma (PPAR gamma), a member of the nuclear receptor superfamily recently shown to function in adipogenesis. The most potent of these agents, BRL49653, binds to PPAR gamma with a Kd of approximately 40 nM. Treatment of pluripotent C3H10T1/2 stem cells with BRL49653 results in efficient differentiation to adipocytes. These data are the first demonstration of a high affinity PPAR ligand and provide strong evidence that PPAR gamma is a molecular target for the adipogenic effects of thiazolidinediones. Furthermore, these data raise the intriguing possibility that PPAR gamma is a target for the therapeutic actions of this class of compounds.