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In the presence of established atherosclerosis, estrogens are potentially harmful. MMP-2 and MMP-9, their inhibitors (TIMP-2 and TIMP-1), RANK, RANKL, OPG, MCP-1, lysyl oxidase (LOX), PDGF-ß, and ADAMTS-4 play critical roles in plaque instability/rupture. We aimed to investigate (i) the effect of estradiol on the expression of the abovementioned molecules in endothelial cells, (ii) which type(s) of estrogen receptors mediate these effects, and (iii) the role of p21 in the estrogen-mediated regulation of the aforementioned factors. Human aortic endothelial cells (HAECs) were cultured with estradiol in the presence or absence of TNF-α. The expression of the aforementioned molecules was assessed by qRT-PCR and ELISA. Zymography was also performed. The experiments were repeated in either ERα- or ERß-transfected HAECs and after silencing p21. HAECs expressed only the GPR-30 estrogen receptor. Estradiol, at low concentrations, decreased MMP-2 activity by 15-fold, increased LOX expression by 2-fold via GPR-30, and reduced MCP-1 expression by 3.5-fold via ERß. The overexpression of ERα increased MCP-1 mRNA expression by 2.5-fold. In a low-grade inflammation state, lower concentrations of estradiol induced the mRNA expression of MCP-1 (3.4-fold) and MMP-9 (7.5-fold) and increased the activity of MMP-2 (1.7-fold) via GPR-30. Moreover, p21 silencing resulted in equivocal effects on the expression of the abovementioned molecules. Estradiol induced different effects regarding atherogenic plaque instability through different ERs. The balance of the expression of the various ER subtypes may play an important role in the paradoxical characterization of estrogens as both beneficial and harmful.
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Aterosclerose , Placa Aterosclerótica , Células Endoteliais/metabolismo , Estradiol/farmacologia , Receptor alfa de Estrogênio/genética , Receptor alfa de Estrogênio/metabolismo , Receptor beta de Estrogênio/genética , Estrogênios/farmacologia , Humanos , Metaloproteinase 2 da Matriz/genética , Metaloproteinase 2 da Matriz/metabolismo , Metaloproteinase 9 da Matriz/genética , Metaloproteinase 9 da Matriz/metabolismo , Placa Aterosclerótica/genética , Proteína-Lisina 6-Oxidase/metabolismo , RNA Mensageiro/metabolismo , Receptores de Estrogênio/metabolismo , Inibidor Tecidual de Metaloproteinase-1/metabolismo , Inibidor Tecidual de Metaloproteinase-2/metabolismo , Transcriptoma , Fator de Necrose Tumoral alfa/genéticaRESUMO
[This corrects the article DOI: 10.1371/journal.pone.0057458.].
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A critical mechanism that has been proposed for transcription regulation by estrogen receptor α (ER) is the tethering of ER to DNA via other transcription factors, such as AP-1. However, genome-wide assessment of the overlap in chromatin binding repertoires of these two transcription factors has not been reported. Here, we show that the AP-1 transcription factor c-Jun interacts with ER and that c-Jun chromatin binding shows extensive overlap with ER binding at the global level. Further, we show that c-Jun overexpression reprograms ER chromatin binding and modulates ER-mediated gene regulation. Our data are consistent with a mechanism where estrogen/ER-dependent crosstalk with AP-1 at the transcriptional level is mediated through the tethering of ER to DNA bound AP-1. Additionally, in our system c-Jun overexpression causes reduced sensitivity to tamoxifen in ER+ breast cancer cells. Integrated cistrome, transcriptome, and clinical data reveal TGFBI as a candidate gene which may confer tamoxifen resistance by ER and AP-1 crosstalk. Further, we show that TGFBI expression is elevated in breast cancer compared to normal breast. Together, our data provide a novel genome-wide footprint of ER and AP-1 crosstalk and suggest AP-1 and TGFBI signaling as potential therapeutic targets in AP-1-overexpressing ER-positive breast tumors.
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Antineoplásicos Hormonais/farmacologia , Neoplasias da Mama/metabolismo , Receptor alfa de Estrogênio/metabolismo , Proteínas Proto-Oncogênicas c-fos/metabolismo , Tamoxifeno/farmacologia , Sítios de Ligação , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/genética , Linhagem Celular Tumoral , Resistencia a Medicamentos Antineoplásicos , Receptor alfa de Estrogênio/química , Proteínas da Matriz Extracelular/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Redes Reguladoras de Genes/efeitos dos fármacos , Humanos , Células MCF-7 , Proteínas Proto-Oncogênicas c-fos/química , Transdução de Sinais/efeitos dos fármacos , Fator de Crescimento Transformador beta/genética , Regulação para CimaRESUMO
Context: Insulin resistance (IR) is promoted by a chronic low-grade inflammation in white adipose tissue (WAT). The latter might be regulated through epigenetic mechanisms such as DNA methylation. The one carbon cycle (1CC) is a central metabolic process governing DNA methylation. Objective: To identify adipocyte-expressed 1CC genes linked to WAT inflammation, IR, and their causal role. Design: Cohort study. Setting: Outpatient academic clinic. Participants: Obese and nonobese subjects. Methods: Gene expression and DNA methylation arrays were performed in subcutaneous WAT and isolated adipocytes. In in vitro differentiated human adipocytes, gene knockdown was achieved by small interfering RNA, and analyses included microarray, quantitative polymerase chain reaction, DNA methylation by enzyme-linked immunosorbent assay and pyrosequencing, protein secretion by enzyme-linked immunosorbent assay, targeted metabolomics, and luciferase reporter and thermal shift assays. Main Outcome Measures: Effects on adipocyte inflammation. Results: In adipocytes from obese individuals, global DNA hypermethylation was associated positively with gene expression of proinflammatory pathways. Among the 1CC genes, IR in vivo and proinflammatory gene expression in WAT were most strongly and inversely associated with SLC19A1, a gene encoding a membrane folate carrier. SLC19A1 knockdown in human adipocytes perturbed intracellular 1CC metabolism, induced global DNA hypermethylation, and increased expression of proinflammatory genes. Several CpG loci linked SLC19A1 to inflammation; validation studies were focused on the chemokine C-C motif chemokine ligand 2 (CCL2) in which methylation in the promoter (cg12698626) regulated CCL2 expression and CCL2 secretion through altered transcriptional activity. Conclusions: Reduced SLC19A1 expression in human adipocytes induces DNA hypermethylation, resulting in increased expression of specific proinflammatory genes, including CCL2. This constitutes an epigenetic mechanism that might link dysfunctional adipocytes to WAT inflammation and IR.
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Adipócitos/metabolismo , Tecido Adiposo/patologia , Metilação de DNA/genética , Inflamação/genética , Resistência à Insulina/genética , Proteína Carregadora de Folato Reduzido/genética , Adipócitos/patologia , Tecido Adiposo/metabolismo , Adulto , Estudos de Casos e Controles , Estudos de Coortes , Epigênese Genética/fisiologia , Feminino , Perfilação da Expressão Gênica , Humanos , Inflamação/metabolismo , Análise em Microsséries , Pessoa de Meia-Idade , Obesidade/complicações , Obesidade/genética , Obesidade/metabolismo , Obesidade/patologia , Paniculite/genética , Paniculite/metabolismo , Paniculite/patologia , Proteína Carregadora de Folato Reduzido/metabolismo , Adulto JovemRESUMO
The two estrogen receptor (ER) subtypes, ERα and ERß, belong to the nuclear receptor superfamily. The human ERß variant ERß2 is proposed to be expressed at higher levels than ERß1 in many breast tumors and it has been suggested that ERß2, in contrast to ERß1, is associated with aggressive phenotypes of various cancers. However, the role of endogenous ERß2 in breast cancer cells remains elusive. In this study, we identified that triple negative breast cancer (TNBC) cell lines express endogenous ERß2, but not ERα or ERß1. This allows novel studies of endogenous ERß2 functions independent of ERα and ERß1. We show that overexpression of ERß2 in TNBC cells increased whereas knockdown of endogenous ERß2 decreased cell proliferation and cell invasion. To elucidate the molecular mechanism responsible for these cellular phenotypes, we assayed ERß2 dependent global gene expression profiles. We show that ERß2 decreases prolyl hydroxylase 3 (PHD3) gene expression and further show that this is associated with increased hypoxia inducible factor 1α (HIF-1α) protein levels, thus providing a possible mechanism for the invasive phenotype. These results are further supported by analysing the expression of ERß2 and PHD3 in breast tumor samples where a negative correlation between ERß2 and PHD3 expression was observed. Together, we demonstrate that ERß2 has an important role in enhancing cell proliferation and invasion, beyond modulation of ERß and ERß1 signalling which might contribute to the invasive characteristics of TNBC. The invasive phenotype could potentially be mediated through transcriptional repression of PHD3 and increased HIF-1α protein levels.
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Both functional ovaries and estrogen replacement therapy (ERT) reduce the risk of type 2 diabetes (T2D). Understanding the mechanisms underlying the antidiabetic effects of 17ß-estradiol (E2) may permit the development of a molecular targeting strategy for the treatment of metabolic disease. This study examines how the promotion of insulin sensitivity and weight loss by E2 treatment in high-fat-diet (HFD)-fed mice involve several anti-adipogenic processes in the visceral adipose tissue. Magnetic resonance imaging (MRI) revealed specific reductions in visceral adipose tissue volume in HFD+E2 mice, compared with HFD mice. This loss of adiposity was associated with diminished visceral adipocyte size and reductions in expression of lipogenic genes, adipokines and of the nuclear receptor nr2c2/tr4. Meanwhile, expression levels of adipose triglyceride lipase/pnpla2 and leptin receptor were increased. As mRNA levels of stat3, a transcription factor involved in brown adipose tissue differentiation, were also increased in visceral adipose, the expression of other brown adipose-specific markers was assessed. Both expression and immunohistochemical staining of ucp-1 were increased, and mRNA levels of dio-2, and of adrß3, a regulator of ucp-1 expression during the thermogenic response, were increased. Furthermore, expression of cpt-1b, a brown adipose-specific gene involved in fatty acid utilization, was also increased. Methylation studies demonstrated that the methylation status of both dio-2 and adrß3 was significantly reduced. These results show that improved glycemic control and weight loss due to E2 involve anti-adipogenic mechanisms which include suppressed lipogenesis and augmented fatty acid utilization, and in addition, the activation of brown adipose tissue-specific gene expression in association with E2-dependent epigenetic modifications in these genes.
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Adipogenia/efeitos dos fármacos , Tecido Adiposo Marrom/metabolismo , Biomarcadores/metabolismo , Estradiol/farmacologia , Regulação da Expressão Gênica/efeitos dos fármacos , Gordura Intra-Abdominal/metabolismo , Obesidade/metabolismo , Tecido Adiposo Marrom/efeitos dos fármacos , Tecido Adiposo Marrom/patologia , Animais , Western Blotting , Metilação de DNA , Dieta Hiperlipídica/efeitos adversos , Feminino , Perfilação da Expressão Gênica , Resistência à Insulina , Gordura Intra-Abdominal/citologia , Gordura Intra-Abdominal/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos C57BL , Obesidade/etiologia , Obesidade/patologia , Análise de Sequência com Séries de Oligonucleotídeos , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
Anti-estrogen treatment, exemplified by tamoxifen, is a well-established adjuvant therapy for estrogen receptor alpha (ERα)-positive breast cancer. However, the effectiveness of this drug is limited due to the development of resistance. The Hedgehog (HH) signaling pathway is critical in embryonic development, and aberrant activation of this transduction cascade is linked to various malignancies. However, it remains unclear whether HH signaling is activated in human breast cancer and related to tamoxifen resistance. Deciphering how this pathway may be involved in breast cancer is a crucial step towards the establishment of targeted combinatorial treatments for this disease. Here, we show that the expression of the HH signaling effector protein GLI1 is higher in tamoxifen resistant compared to sensitive cells. Tamoxifen resistant cells have stronger ERα transcriptional activity relative to sensitive cells, even though the ERα expression is similar in both cell types. Knockdown of GLI1 attenuates cell proliferation and reduces ERα transcriptional activity in both sensitive and resistant cells, irrespective of estrogen stimulation. Combinatorial treatment of tamoxifen and the GLI antagonist GANT61 further suppresses the growth of sensitive and resistant cells relative to administration of only tamoxifen, and this was irrespective of estrogen stimulation. Moreover, a positive correlation between GLI1 and ERα expression was identified in breast cancer samples. Additionally, high GLI1 expression predicted worse distant metastasis-free survival in breast cancer patients. These data suggest that the HH pathway may be a new candidate for therapeutic targeting and prognosis in ERα-positive breast cancer.
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Neoplasias da Mama/patologia , Receptor alfa de Estrogênio/genética , Proteínas Hedgehog/fisiologia , Transdução de Sinais/fisiologia , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/mortalidade , Proliferação de Células , Regulação para Baixo , Resistencia a Medicamentos Antineoplásicos , Receptor alfa de Estrogênio/fisiologia , Estrogênios/farmacologia , Feminino , Humanos , Células MCF-7 , Piridinas/farmacologia , Pirimidinas/farmacologia , Elementos de Resposta , Tamoxifeno/farmacologia , Proteína GLI1 em Dedos de Zinco/fisiologiaRESUMO
Endocrine therapy is the first-line targeted adjuvant therapy for hormone-sensitive breast cancer. In view of the potential anticancer property of the omega-3 polyunsaturated fatty acid docosahexaenoic acid (DHA) together with chemotherapy in estrogen receptor alpha (ERα) positive mammary tumors, we have explored the regulation by estradiol of the fatty acid desaturation and elongation enzymes involved in DHA synthesis in the human breast cancer cell line MCF7, which expresses ERα but not ERß. We demonstrate a robust up-regulation in the expression of the fatty acid elongases Elovl2 and Elovl5 upon estradiol stimulation in MCF7 cells, which was sustained for more than 24 hours. Exposure with the ER inhibitor tamoxifen abolished specifically the Elovl2 but not the Elovl5 expression. Similarly, knock-down of ERα eliminated almost fully the Elovl2 but not the Elovl5 expression. Furthermore, ERα binds to one specific ERE within the Elovl2 enhancer in a ligand dependent manner. The involvement of ERα in the control of especially Elovl2, which plays a crucial role in DHA synthesis, may have potential implications in the treatment of breast cancer.
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Acetiltransferases/genética , Acetiltransferases/metabolismo , Receptor alfa de Estrogênio/metabolismo , Estrogênios/farmacologia , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Receptor alfa de Estrogênio/deficiência , Receptor alfa de Estrogênio/genética , Elongases de Ácidos Graxos , Ácidos Graxos Insaturados/biossíntese , Técnicas de Silenciamento de Genes , Células Hep G2 , Humanos , Células MCF-7 , Regiões Promotoras Genéticas/efeitos dos fármacos , Regiões Promotoras Genéticas/genética , Elementos de Resposta/efeitos dos fármacos , Elementos de Resposta/genética , Tamoxifeno/farmacologiaRESUMO
Estrogen receptor α (ERα) is a key regulator of breast growth and breast cancer development. Here, we report how ERα impacts these processes by reprogramming metabolism in malignant breast cells. We employed an integrated approach, combining genome-wide mapping of chromatin-bound ERα with estrogen-induced transcript and metabolic profiling, to demonstrate that ERα reprograms metabolism upon estrogen stimulation, including changes in aerobic glycolysis, nucleotide and amino acid synthesis, and choline (Cho) metabolism. Cho phosphotransferase CHPT1, identified as a direct ERα-regulated gene, was required for estrogen-induced effects on Cho metabolism, including increased phosphatidylcholine synthesis. CHPT1 silencing inhibited anchorage-independent growth and cell proliferation, also suppressing early-stage metastasis of tamoxifen-resistant breast cancer cells in a zebrafish xenograft model. Our results showed that ERα promotes metabolic alterations in breast cancer cells mediated by its target CHPT1, which this study implicates as a candidate therapeutic target. Cancer Res; 76(19); 5634-46. ©2016 AACR.
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Neoplasias da Mama/etiologia , Colina/metabolismo , Receptor alfa de Estrogênio/fisiologia , Animais , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/metabolismo , Colina-Fosfato Citidililtransferase/fisiologia , Diacilglicerol Colinofosfotransferase/fisiologia , Resistencia a Medicamentos Antineoplásicos , Feminino , Humanos , Células MCF-7 , Metástase Neoplásica , Tamoxifeno/uso terapêutico , Peixe-ZebraRESUMO
Endurance and resistance exercise training induces specific and profound changes in the skeletal muscle transcriptome. Peroxisome proliferator-activated receptor γ coactivator-1 α (PGC-1α) coactivators are not only among the genes differentially induced by distinct training methods, but they also participate in the ensuing signaling cascades that allow skeletal muscle to adapt to each type of exercise. Although endurance training preferentially induces PGC-1α1 expression, resistance exercise activates the expression of PGC-1α2, -α3, and -α4. These three alternative PGC-1α isoforms lack the arginine/serine-rich (RS) and RNA recognition motifs characteristic of PGC-1α1. Discrete functions for PGC-1α1 and -α4 have been described, but the biological role of PGC-1α2 and -α3 remains elusive. Here we show that different PGC-1α variants can affect target gene splicing through diverse mechanisms, including alternative promoter usage. By analyzing the exon structure of the target transcripts for each PGC-1α isoform, we were able to identify a large number of previously unknown PGC-1α2 and -α3 target genes and pathways in skeletal muscle. In particular, PGC-1α2 seems to mediate a decrease in the levels of cholesterol synthesis genes. Our results suggest that the conservation of the N-terminal activation and repression domains (and not the RS/RNA recognition motif) is what determines the gene programs and splicing options modulated by each PGC-1α isoform. By using skeletal muscle-specific transgenic mice for PGC-1α1 and -α4, we could validate, in vivo, splicing events observed in in vitro studies. These results show that alternative PGC-1α variants can affect target gene expression both quantitatively and qualitatively and identify novel biological pathways under the control of this system of coactivators.
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Processamento Alternativo , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/metabolismo , Animais , Células Cultivadas , Sequência Conservada , Éxons , Regulação da Expressão Gênica , Redes Reguladoras de Genes , Camundongos , Camundongos Transgênicos , Fibras Musculares Esqueléticas/metabolismo , Músculo Esquelético/metabolismo , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/química , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/genética , Regiões Promotoras Genéticas , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Estabilidade Proteica , Receptores de Esteroides/química , Receptores de Esteroides/genética , Receptores de Esteroides/metabolismoRESUMO
OBJECTIVE: Epigenetic modifications contribute to the etiology of type 2 diabetes. METHOD: We performed genome-wide methylome and transcriptome analysis in liver from severely obese men with or without type 2 diabetes and non-obese men to discover aberrant pathways underlying the development of insulin resistance. Results were validated by pyrosequencing. RESULT: We identified hypomethylation of genes involved in hepatic glycolysis and insulin resistance, concomitant with increased mRNA expression and protein levels. Pyrosequencing revealed the CpG-site within ATF-motifs was hypomethylated in four of these genes in liver of severely obese non-diabetic and type 2 diabetic patients, suggesting epigenetic regulation of transcription by altered ATF-DNA binding. CONCLUSION: Severely obese non-diabetic and type 2 diabetic patients have distinct alterations in the hepatic methylome and transcriptome, with hypomethylation of several genes controlling glucose metabolism within the ATF-motif regulatory site. Obesity appears to shift the epigenetic program of the liver towards increased glycolysis and lipogenesis, which may exacerbate the development of insulin resistance.
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Mastermind-like 1 (MAML1) is a transcriptional coregulator that has been associated with early development of many systems such as neuronal, muscular and urogenital. The present study aimed to explore the genome wide effects of MAML1 on DNA methylation and RNA expression in human embryonic kidney cells. Infinium HumanMethylation450 BeadChip Illumina array, methylation-sensitive high-resolution melt technique, Chip Analysis Methylation Pipeline and RNA profiling approaches were used to study MAML1 effects on the epigenome. We found that 11802 CpG sites were differentially methylated in MAML1-expressing cells while only 225 genes were differentially expressed. MAML1 overexpression induced more global differential hypermethylation than hypomethylation changes. In addition, the differentially methylated regions were mapped predominantly to 3'untranslated regions, intragenic regions and gene bodies and to a lesser extent to gene regulatory sequences. Gene ontology analysis revealed that the differentially changed genes (including HOXC11, HTATIP2, SLFN12 and SOX11) are involved in the regulation of urogenital system development, cell adhesion and embryogenesis. This study is the first report that shows the global effect of a single coregulator on DNA methylation and gene expression. Our results stress and support the effects of transcriptional coregulators on the cell methylome.
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Ilhas de CpG , Metilação de DNA , Proteínas de Ligação a DNA/genética , Rim/citologia , Fatores de Transcrição/genética , Transcriptoma , Acetiltransferases/genética , Acetiltransferases/metabolismo , Células HEK293 , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Humanos , Rim/metabolismo , Análise de Sequência com Séries de Oligonucleotídeos , RNA Mensageiro , Fatores de Transcrição SOXC/genética , Fatores de Transcrição SOXC/metabolismo , Fatores de Transcrição/metabolismo , Regulação para CimaRESUMO
Triple-negative breast cancer (TNBC) represents a highly aggressive form of breast cancer with limited treatment options. Proinflammatory cytokines such as TNFα can facilitate tumor progression and metastasis. However, the mechanistic aspects of inflammation mediated TNBC progression remain unclear. Using ChIP-seq, we demonstrate that the cistrome for the AP-1 transcription factor c-Jun is comprised of 13,800 binding regions in TNFα-stimulated TNBC cells. In addition, we show that c-Jun regulates nearly a third of the TNFα-regulated transcriptome. Interestingly, high expression level of the c-Jun-regulated pro-invasion gene program is associated with poor clinical outcome in TNBCs. We further demonstrate that c-Jun drives TNFα-mediated increase of malignant characteristics of TNBC cells by transcriptional regulation of Ninj1. As exemplified by the CXC chemokine genes clustered on chromosome 4, we demonstrate that NF-κB might be a pioneer factor required for the regulation of TNFα-inducible inflammatory genes, whereas c-Jun has little effect. Together, our results uncover AP-1 as an important determinant for inflammation-induced cancer progression, rather than inflammatory response.
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Regulação Neoplásica da Expressão Gênica , Proteínas Proto-Oncogênicas c-jun/metabolismo , Fator de Transcrição AP-1/metabolismo , Neoplasias de Mama Triplo Negativas/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Moléculas de Adesão Celular Neuronais/genética , Moléculas de Adesão Celular Neuronais/metabolismo , Linhagem Celular Tumoral , Cromossomos Humanos Par 4/genética , Feminino , Humanos , NF-kappa B/genética , NF-kappa B/metabolismo , Fatores de Crescimento Neural/genética , Fatores de Crescimento Neural/metabolismo , Proteínas Proto-Oncogênicas c-jun/genética , Fator de Transcrição AP-1/genética , Transcriptoma , Neoplasias de Mama Triplo Negativas/genética , Neoplasias de Mama Triplo Negativas/patologia , Fator de Necrose Tumoral alfa/genéticaRESUMO
OBJECTIVE: Siblings born before (BMS) and after (AMS) maternal biliopancreatic diversion (BPD) show differences in the methylome. The objective was to use a sibling-pair design to examine the effects from interpregnancy weight loss as a consequence of maternal bariatric surgery, other than BPD, on the methylome comparing BMS and AMS. METHODS: Women with at least one child born before and one after bariatric surgery were identified in Swedish national registers. Whole blood samples from BMS (N = 31) and AMS (N = 31) siblings were collected for epigenetic methylation analysis while maternal information was collected from antenatal medical records. RESULTS: In total 3,074 genes, with corresponding 23,449 CpG methylation sites, were differently methylated and associated with an overrepresentation of differently methylated CpG sites in genes involved with insulin receptor signaling, type 2 diabetes signaling, and leptin signaling in obesity, while the most significant differently methylated genes were HLA-DQA1, HLA-DQB1, and TSPAN18, when comparing BMS and AMS siblings. CONCLUSIONS: These results suggest that maternal bariatric surgery, with subsequent weight loss between pregnancies, is associated with alterations in the methylome of genes involved in insulin receptor signaling, type 2 diabetes signaling, and leptin signaling in obesity in a comparison of BMS and AMS siblings.
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Cirurgia Bariátrica , Metilação de DNA/genética , Diabetes Mellitus Tipo 2/genética , Doenças em Gêmeos , Inflamação/genética , Adulto , Desvio Biliopancreático , Criança , Pré-Escolar , Diabetes Mellitus Tipo 2/complicações , Feminino , Cadeias alfa de HLA-DQ/genética , Cadeias beta de HLA-DQ/genética , Humanos , Lactente , Recém-Nascido , Inflamação/complicações , Leptina/sangue , Masculino , Obesidade/complicações , Gravidez , Suécia , Tetraspaninas/genéticaRESUMO
Estrogen receptor alpha (ERα) is highly expressed in most breast cancers. Consequently, ERα modulators, such as tamoxifen, are successful in breast cancer treatment, although tamoxifen resistance is commonly observed. While tamoxifen resistance may be caused by altered ERα signaling, the molecular mechanisms regulating ERα signaling and tamoxifen resistance are not entirely clear. Here, we found that PAK4 expression was consistently correlated to poor patient outcome in endocrine treated and tamoxifen-only treated breast cancer patients. Importantly, while PAK4 overexpression promoted tamoxifen resistance in MCF-7 human breast cancer cells, pharmacological treatment with a group II PAK (PAK4, 5, 6) inhibitor, GNE-2861, sensitized tamoxifen resistant MCF-7/LCC2 breast cancer cells to tamoxifen. Mechanistically, we identified a regulatory positive feedback loop, where ERα bound to the PAK4 gene, thereby promoting PAK4 expression, while PAK4 in turn stabilized the ERα protein, activated ERα transcriptional activity and ERα target gene expression. Further, PAK4 phosphorylated ERα-Ser305, a phosphorylation event needed for the PAK4 activation of ERα-dependent transcription. In conclusion, PAK4 may be a suitable target for perturbing ERα signaling and tamoxifen resistance in breast cancer patients.
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Benzimidazóis/farmacologia , Neoplasias da Mama/metabolismo , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Inibidores Enzimáticos/farmacologia , Receptor alfa de Estrogênio/metabolismo , Pirimidinas/farmacologia , Quinases Ativadas por p21/metabolismo , Neoplasias da Mama/mortalidade , Neoplasias da Mama/patologia , Bases de Dados Genéticas , Resistencia a Medicamentos Antineoplásicos/fisiologia , Feminino , Citometria de Fluxo , Humanos , Immunoblotting , Estimativa de Kaplan-Meier , Células MCF-7 , Reação em Cadeia da Polimerase , RNA Interferente Pequeno , Moduladores Seletivos de Receptor Estrogênico/farmacologia , Transdução de Sinais/fisiologia , Tamoxifeno/farmacologia , TransfecçãoRESUMO
BACKGROUND: Obesity is associated with changes in fat cell gene expression and metabolism. What drives these changes is not well understood. We aimed to explore fat cell epigenetics, i.e., DNA methylation, as one mediator of gene regulation, in obese women. The global DNA methylome for abdominal subcutaneous fat cells was compared between 15 obese case (BMI 41.4 ± 4.4 kg/m(2), mean ± SD) and 14 never-obese control women (BMI 25.2 ± 2.5 kg/m(2)). Global array-based transcriptome analysis was analyzed for subcutaneous white adipose tissue (WAT) from 11 obese and 9 never-obese women. Limma was used for statistical analysis. RESULTS: We identified 5529 differentially methylated DNA sites (DMS) for 2223 differentially expressed genes between obese cases and never-obese controls (false discovery rate <5 %). The 5529 DMS displayed a median difference in beta value of 0.09 (range 0.01 to 0.40) between groups. DMS were under-represented in CpG islands and in promoter regions, and over-represented in open sea-regions and gene bodies. The 2223 differentially expressed genes with DMS were over-represented in key fat cell pathways: 31 of 130 (25 %) genes linked to "adipogenesis" (adjusted P = 1.66 × 10(-11)), 31 of 163 (19 %) genes linked to "insulin signaling" (adjusted P = 1.91 × 10(-9)), and 18 of 67 (27 %) of genes linked to "lipolysis" (P = 6.1 × 10(-5)). In most cases, gene expression and DMS displayed reciprocal changes in obese women. Furthermore, among 99 candidate genes in genetic loci associated with body fat distribution in genome-wide association studies (GWAS); 22 genes displayed differential expression accompanied by DMS in obese versus never-obese women (P = 0.0002), supporting the notion that a significant proportion of gene loci linked to fat distribution are epigenetically regulated. CONCLUSIONS: Subcutaneous WAT from obese women is characterized by congruent changes in DNA methylation and expression of genes linked to generation, distribution, and metabolic function of fat cells. These alterations may contribute to obesity-associated metabolic disturbances such as insulin resistance in women.
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Estrogen receptor alpha (ERα) is an important regulator of the estrous cycle and mice with global ERα deletion, as well as some conditional knockout mouse lines, have an interruption in the estrous cycle. In this study we observed that conditional ERα knockout mice where the Cre gene is regulated by the rat insulin promoter (RIP), RIP-Cre/ERα(KO) mice, have a 3.7-fold increase in serum 17ß-estradiol levels, blocked estrous cycle, and develop a fluid-filled uterus (hydrometra). Using a proteomics approach, we identified three proteins, lactoferrin, complement C3 and chitinase 3-like protein 1 (CHI3L1), as highly expressed proteins in hydrometra fluid. The mRNA levels of the corresponding genes were more than 50-fold higher in RIP-Cre/ERα(KO) uterus compared to controls. High expression of CHI3L1 in the uterine fluid was not reflected as elevated levels in the serum. The high expression of lactoferrin, complement C3 and CHI3L1 in the uterine fluid, in association with elevated estrogen levels, prompted us to address if the expression of these genes is related to reproduction. However, gonadotropin treatment of mice reduced the uterine expression of these genes in a model of in vitro fertilization. Our findings identify lactoferrin, complement C3 and CHI3L1 as highly expressed proteins in hydrometra fluid in association with chronically elevated serum estradiol levels.