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1.
Cancer Biomark ; 25(2): 151-160, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31045512

RESUMO

BACKGROUND: Amplified in breast cancer 1 (AIB1) is a candidate oncogene in human breast cancer, which has been identified to be amplified and overexpressed in several types of other human cancers. Abnormalities of AIB1 and its clinical/prognostic significance, however, in upper tract urothelial carcinoma (UTUC) remain unclear. OBJECTIVE: To explore what role AIB1 plays in upper tract urothelial carcinoma. METHODS: The expression of AIB1 was analyzed using immunohistochemical staining in 133 UTUC patients. Overall, cancer specific and recurrence-free survival rates (OS, CSS, and RFS) were estimated using the Kaplan-Meier method. Multivariable COX regression models containing relevant clinicopathological variables addressed the prediction of postoperative outcome. RESULTS: High AIB1 expression was observed to be associated with increased hazard ratios for 5-year CSS (80.6% vs. 55.8%, p= 0.008) and OS (78.1% vs. 54.8%, p= 0.006). Multivariable analysis revealed that elevated AIB1 expression was an independent prognostic predictor of OS, CSS and RFS. Additionally, pT, pN and hydronephrosis were independently associated with oncologic outcome of UTUC. Three proposed nomograms were proposed to provide an individualized risk estimate of postoperative outcome in patients with UTUC. CONCLUSIONS: AIB1 can be used as an independent molecular marker for the prognosis of clinical outcomes of UTUC.


Assuntos
Biomarcadores Tumorais , Expressão Gênica , Coativador 3 de Receptor Nuclear/genética , Neoplasias Urológicas/genética , Neoplasias Urológicas/mortalidade , Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Humanos , Imuno-Histoquímica , Estimativa de Kaplan-Meier , Masculino , Pessoa de Meia-Idade , Gradação de Tumores , Estadiamento de Neoplasias , Nefroureterectomia/métodos , Coativador 3 de Receptor Nuclear/metabolismo , Modelos de Riscos Proporcionais , Curva ROC , Resultado do Tratamento , Carga Tumoral , Neoplasias Urológicas/patologia , Neoplasias Urológicas/cirurgia
2.
J Chem Inf Model ; 59(2): 842-857, 2019 02 25.
Artigo em Inglês | MEDLINE | ID: mdl-30658039

RESUMO

Androgen receptor (AR), as a member of the nuclear receptor (NR) superfamily, regulates the gene transcription in response to the sequential binding of diverse agonists and coactivators. Great progress has been made in studies on the pharmacology and structure of AR, but the atomic level mechanism of the bidirectional communications between the ligand-binding pocket (LBP) and the activation function-2 (AF2) region of AR remains poorly understood. Therefore, in this study, molecular dynamics (MD) simulations and free energy calculations were carried out to explore the interactions among water, agonist (DHT) or antagonist (HFT), AR, and coactivator (SRC3). Upon the binding of an agonist (DHT) or antagonist (HFT), the LBP structure would transform to the agonistic or antagonistic state, and the conformational changes of the LBP would regulate the structure of the AF2 surface. As a result, the binding of the androgen DHT could promote the recruitment of the coactivator SRC3 to the AF2, and on the contrary, the binding of the antagonist HFT would induce a perturbation to the shape of the AF2 and then weaken its accommodating capability of the coactivators with the LXXLL motif. The simulation results illustrated that the DHT-AR binding affinity was enhanced by the association of the coactivator SRC3, which would reduce the conformational fluctuation of the AR-LBD and expand the size of the AR LBP. On the other hand, the coactivator-to-HFT allosteric pathway, which involves the SRC3, helix 3 (H3), helix 4 (H4), the loop (L1-3) between helix 1 (H1) and H3, and HFT, was characterized. The HFT's skewness and different interactions between HFT and the LBP were observed in the SRC3-present AR. The mutual communications between the AF2 surface and LBP, together with the processes involving the interplay of the ligand binding and coactivator recruitment events, would help in understanding the association of coactivators and rationally develop potent drugs to inhibit the activity of AR.


Assuntos
Simulação de Dinâmica Molecular , Receptores Androgênicos/química , Receptores Androgênicos/metabolismo , Motivos de Aminoácidos , Sequência de Aminoácidos , Sítios de Ligação , Ligantes , Coativador 3 de Receptor Nuclear/metabolismo , Ligação Proteica , Termodinâmica
3.
J Steroid Biochem Mol Biol ; 185: 57-70, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-30048685

RESUMO

Steroid receptor coactivator 2 (SRC-2) is a nuclear receptor coactivator, important for the regulation of estrogen receptor alpha (ERα)-mediated transcriptional activity in breast cancer cells. However, the transcriptional role of SRC-2 in breast cancer is still ambiguous. Here we aimed to unravel a more precise transcriptional role of SRC-2 and uncover unique target genes in MCF-7 breast cancer cells, as opposed to the known oncogene SRC-3. Gene expression analyses of cells depleted of either SRC-2 or SRC-3 showed that they transcriptionally regulate mostly separate gene sets. However, individual unique gene sets were implicated in some of the same major gene ontology biological processes, such as cellular structure and development. This finding was supported by three-dimensional cell cultures, demonstrating that depletion of SRC-2 and SRC-3 changed the morphology of the cells into epithelial-like hollow acinar structures, indicating that both SRC proteins are involved in maintaining the hybrid E/M phenotype. In clinical ER-positive, HER2-negative breast cancer samples the expression of SRC-2 was negatively correlated with the expression of MCF-7-related luminal, cell cycle and cellular morphogenesis genes. Finally, elucidating SRC-2 unique transcriptional effects, we identified Lyn kinase (an EMT biomarker) to be upregulated exclusively after SRC-2 depletion. In conclusion, we show that both SRC-2 and SRC-3 are essential for the EMT in breast cancer cells, controlling different transcriptional niches.


Assuntos
Neoplasias da Mama/patologia , Transição Epitelial-Mesenquimal/fisiologia , Coativador 2 de Receptor Nuclear/metabolismo , Coativador 3 de Receptor Nuclear/metabolismo , Feminino , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Células MCF-7 , Coativador 2 de Receptor Nuclear/genética , Coativador 3 de Receptor Nuclear/genética , Receptor ErbB-2/metabolismo , Receptores Estrogênicos/metabolismo , Esferoides Celulares/citologia , Transcrição Genética/genética , Células Tumorais Cultivadas , Quinases da Família src/biossíntese , Quinases da Família src/genética
4.
Acta Pharmacol Sin ; 40(5): 648-657, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30089865

RESUMO

Hyperglycemia is a major factor in vascular endothelial injury that finally leads to a cardiovascular event. Steroid receptor coactivators (SRCs) are a group of non-DNA binding proteins that induce structural changes in steroid receptors (nuclear receptors) critical for transcriptional activation. SRCs, namely, SRC-1, SRC-2, and SRC-3, are implicated in the regulation of vascular homeostasis. In this study we investigate the role of SRCs in hyperglycemia-induced endothelial injury. Aortic endothelial cells were prepared from normal and diabetic rats, respectively. Diabetic rats were prepared by injection of streptozotocin (50 mg/kg, i.p.). The expression levels of SRC-1 and SRC-3 were significantly decreased in endothelial cells from the diabetic rats. Similar phenomenon was also observed in aortic endothelial cells from the normal rats treated with a high glucose (25 mM) for 4 h or 8 h. The expression levels of SRC-2 were little affected by hyperglycemia. Overexpression of SRC-1 and SRC-3 in high glucose-treated endothelial cells significantly increased the cell viability, suspended cell senescence, and inhibited cell apoptosis compared with the control cells. We further showed that overexpression of SRC-1 and SRC-3 markedly suppressed endothelial injury through restoring nitric oxide production, upregulating the expression of antioxidant enzymes (SOD, GPX, and CAT), and activating the PI3K/Akt pathway. The beneficial effects of SRC-1 and SRC-3 overexpression were blocked by treatment with the PI3K inhibitor LY294002 (10 mM) or with the Akt inhibitor MK-2206 (100 nM). In conclusion, hyperglycemia decreased SRC-1 and SRC-3 expression levels in rat aortic endothelial cells. SRC-1 and SRC-3 overexpression might protect against endothelial injury via inhibition of oxidative stress and activation of PI3K/Akt pathway.


Assuntos
Diabetes Mellitus Experimental/fisiopatologia , Células Endoteliais/metabolismo , Coativador 1 de Receptor Nuclear/metabolismo , Coativador 3 de Receptor Nuclear/metabolismo , Transdução de Sinais/fisiologia , Animais , Aorta/metabolismo , Aorta/patologia , Apoptose/genética , Sobrevivência Celular/genética , Senescência Celular/genética , Cromonas/farmacologia , Regulação para Baixo , Células Endoteliais/patologia , Endotélio/metabolismo , Endotélio/patologia , Compostos Heterocíclicos com 3 Anéis/farmacologia , Masculino , Morfolinas/farmacologia , Coativador 1 de Receptor Nuclear/genética , Coativador 3 de Receptor Nuclear/genética , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-akt/metabolismo , Ratos Sprague-Dawley , Transdução de Sinais/efeitos dos fármacos
5.
Cancer Lett ; 442: 310-319, 2019 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-30423406

RESUMO

Pancreatic ductal adenocarcinoma (PDAC) is a highly malignant and lethal disease with few treatment options. Steroid receptor coactivator-3 (SRC-3, also known as NCOA3, AIB1, pCIP, ACTR, RAC3, TRAM1) sits at the nexus of many growth signaling pathways and has been pursued as a therapeutic target for breast, prostate and lung cancers. In this study, we find that SRC-3 is overexpressed in PDAC and inversely correlates with patient overall survival. Knockdown of SRC-3 reduces pancreatic cancer cell proliferation, migration and invasion in vitro. Additionally, inhibition of SRC-3 using either shRNA or a small molecule inhibitor can significantly inhibit tumor growth in orthotopic pancreatic cancer mouse models. Collectively, this study establishes SRC-3 as a promising therapeutic target for pancreatic cancer treatment.


Assuntos
Antineoplásicos/farmacologia , Carcinoma Ductal Pancreático/terapia , Proliferação de Células/efeitos dos fármacos , Coativador 3 de Receptor Nuclear/antagonistas & inibidores , Coativador 3 de Receptor Nuclear/genética , Neoplasias Pancreáticas/terapia , RNA Interferente Pequeno/genética , Terapêutica com RNAi , Animais , Carcinoma Ductal Pancreático/genética , Carcinoma Ductal Pancreático/metabolismo , Carcinoma Ductal Pancreático/patologia , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Camundongos Nus , Camundongos SCID , Invasividade Neoplásica , Coativador 3 de Receptor Nuclear/metabolismo , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patologia , RNA Interferente Pequeno/metabolismo , Transdução de Sinais , Carga Tumoral/efeitos dos fármacos , Ensaios Antitumorais Modelo de Xenoenxerto
6.
J Biol Chem ; 294(4): 1230-1239, 2019 01 25.
Artigo em Inglês | MEDLINE | ID: mdl-30514761

RESUMO

Many intrinsically disordered proteins (IDPs) attain a well-defined structure in a coupled folding and binding reaction with another protein. Such reactions may involve early to late formation of different native structural regions along the reaction pathway. To obtain insights into the transition state for a coupled binding and folding reaction, we performed restrained molecular dynamics simulations using previously determined experimental binding Φb values of the interaction between two IDP domains: the activation domain from the p160 transcriptional co-activator for thyroid hormone and retinoid receptors (ACTR) and the nuclear co-activator binding domain (NCBD) of CREB-binding protein, each forming three well-defined α-helices upon binding. These simulations revealed that both proteins are largely disordered in the transition state for complex formation, except for two helices, one from each domain, that display a native-like structure. The overall transition state structure was extended and largely dynamic with many weakly populated contacts. To test the transition state model, we combined site-directed mutagenesis with kinetic experiments, yielding results consistent with overall diffuse interactions and formation of native intramolecular interactions in the third NCBD helix during the binding reaction. Our findings support the view that the transition state and, by inference, any encounter complex in coupled binding and folding reactions are structurally heterogeneous and largely independent of specific interactions. Furthermore, experimental Φb values and Brønsted plots suggested that the transition state is globally robust with respect to most mutations but can display more native-like features for some highly destabilizing mutations, possibly because of Hammond behavior or ground-state effects.


Assuntos
Proteína de Ligação a CREB/metabolismo , Proteínas Intrinsicamente Desordenadas/metabolismo , Coativador 3 de Receptor Nuclear/metabolismo , Dobramento de Proteína , Proteína de Ligação a CREB/química , Proteína de Ligação a CREB/genética , Cristalografia por Raios X , Humanos , Proteínas Intrinsicamente Desordenadas/química , Proteínas Intrinsicamente Desordenadas/genética , Modelos Moleculares , Mutação , Coativador 3 de Receptor Nuclear/química , Coativador 3 de Receptor Nuclear/genética , Ligação Proteica , Conformação Proteica , Transdução de Sinais
7.
Int J Biol Sci ; 14(14): 2051-2064, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30585268

RESUMO

Goblet cell loss, which leads to the reduction of mucin secretion, is a hallmark of ulcerative colitis (UC). We previously reported that steroid receptor coactivator 3 (SRC-3), a transcriptional coactivator, contributes to host defense against Citrobacter rodentium by recruiting neutrophils, suggesting a role of SRC-3 in intestine homeostasis. However, the biological role of SRC-3 in UC remains unclear. Here, we showed that SRC-3-/- mice were more susceptible to dextran sulfate sodium (DSS)-induced colitis compared with wild-type mice after oral administration of 2% DSS dissolved in drinking water. After oral administration of 2% DSS, SRC-3-/- mice displayed higher mortality rate, significant body weight loss, and higher clinical symptom scores compared to wild-type mice. SRC-3-/- mice suffered a severe loss of mature colonic goblet cells, leading to more severe histopathology and more proinflammatory cytokine production. Mechanistically, SRC-3-/- mice exhibited a decreased expression of transcription factor KLF4 in the colons, which is responsible for colonic goblet cell differentiation and maturation. At the molecular level, SRC-3 cooperated with c-Fos to promote KLF4 expression at the transcriptional level. These results demonstrate that SRC-3 can ameliorate DSS-induced colitis by inhibiting inflammation and promoting colonic goblet cell differentiation and maturation through enhancing the expression of transcriptional factor KLF4, which is responsible for colonic goblet cell differentiation and maturation.


Assuntos
Colite/metabolismo , Sulfato de Dextrana/toxicidade , Células Caliciformes/citologia , Células Caliciformes/metabolismo , Inflamação/metabolismo , Fatores de Transcrição Kruppel-Like/metabolismo , Coativador 3 de Receptor Nuclear/metabolismo , Animais , Diferenciação Celular/genética , Diferenciação Celular/fisiologia , Feminino , Humanos , Immunoblotting , Imunoprecipitação , Fatores de Transcrição Kruppel-Like/genética , Camundongos , Camundongos Knockout , Coativador 3 de Receptor Nuclear/genética , Proteínas Proto-Oncogênicas c-fos/genética , Proteínas Proto-Oncogênicas c-fos/metabolismo
8.
J Biol Chem ; 293(42): 16193-16205, 2018 10 19.
Artigo em Inglês | MEDLINE | ID: mdl-30166347

RESUMO

ERK3 is an atypical mitogen-activated protein kinase (MAPK) that has recently gained interest for its role in promoting cancer cell migration and invasion. However, the molecular regulation of ERK3 functions in cancer cells is largely unknown. ERK3 has a single phospho-acceptor site (Ser189) in its activation motif rather than the TXY conserved in conventional MAPKs such as ERK1/2. Although dual phosphorylation of the TXY motif is known to be critical for the activation of conventional MAPKs, the role of Ser189 phosphorylation in ERK3 activity and its function in cancer cells remain elusive. In this study, we revealed that activation loop phosphorylation is important for ERK3 in promoting cancer cell invasiveness, as the S189A mutation greatly decreased the ability of ERK3 to promote migration and invasion of lung cancer cells. Interestingly, a catalytically inactive ERK3 mutant was still capable of increasing migration and invasion, although to a lesser extent compared with WT ERK3, suggesting that ERK3 promotes cancer cell invasiveness by both kinase-dependent and kinase-independent mechanisms. To elucidate how the S189A mutation reduces the invasiveness-promoting ability of ERK3, we tested its effect on the kinase activity of ERK3 toward steroid receptor coactivator 3 (SRC3), a recently identified substrate of ERK3 critical for cancer cell invasiveness. Compared with ERK3, ERK3-S189A exhibited a dramatic decrease in kinase activity toward SRC3 and a concomitantly reduced ability to stimulate matrix metalloproteinase expression. Taken together, our study unravels the importance of Ser189 phosphorylation for intramolecular regulation of ERK3 kinase activity and invasiveness-promoting ability in lung cancer cells.


Assuntos
Neoplasias Pulmonares/patologia , Proteína Quinase 6 Ativada por Mitógeno/metabolismo , Sítios de Ligação , Movimento Celular , Humanos , Invasividade Neoplásica , Coativador 3 de Receptor Nuclear/metabolismo , Fosforilação , Serina/metabolismo
9.
Biophys J ; 115(6): 996-1006, 2018 09 18.
Artigo em Inglês | MEDLINE | ID: mdl-30173887

RESUMO

Interactions between emerging nascent polypeptide chains and the ribosome can modulate cotranslational protein folding. However, it has remained unclear how such interactions can affect the binding of nascent chains to their cellular targets. We thus investigated on the ribosome the interaction between two intrinsically disordered proteins of opposite charge, ACTR and NCBD, which form a high-affinity complex in a coupled folding-and-binding reaction. Using fluorescence correlation spectroscopy and arrest-peptide-mediated force measurements in vitro and in vivo, we find that the ACTR-NCBD complex can form cotranslationally but only with ACTR as the nascent chain and NCBD free in solution, not vice versa. We show that this surprising asymmetry in behavior is caused by pronounced charge interactions: attraction of the positively charged nascent chain of NCBD to the negatively charged ribosomal surface competes with complex formation and prevents ACTR binding. In contrast, the negatively charged nascent ACTR is repelled by the ribosomal surface and thus remains available for productively binding its partner. Electrostatic interactions may thus be more important for cotranslational folding and binding than previously thought.


Assuntos
Coativador 3 de Receptor Nuclear/química , Coativador 3 de Receptor Nuclear/metabolismo , Dobramento de Proteína , Ribossomos/metabolismo , Modelos Moleculares , Domínios Proteicos
10.
Cell Physiol Biochem ; 49(6): 2396-2413, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30261507

RESUMO

BACKGROUND/AIMS: Osteoarthritis (OA) is the most common joint disease. Recently, a novel variant near the nuclear receptor coactivator 3 (NCOA3) has been identified in association with greater risk of developing OA. However, how NCOA3 is regulated in chondrocytes and involved in OA pathogenesis remain elusive. METHODS: The expression and DNA methylation of NCOA3 in knee OA cartilage and in vitro dedifferentiated chondrocytes with or without rs6094710 SNP were analyzed by qRT-PCR, immunoblotting, methylation-specific PCR and bisulfite sequencing. NCOA3 was depleted by siRNA or shRNA or inhibited by a chemical inhibitor to assess its role in chondrocyte dedifferentiation or OA pathogenesis in posttraumatic OA animal model established by cruciate ligament transection surgery. RESULTS: We found that compared with normal counterparts, samples with rs6094710 SNP failed to upregulate NCOA3. Further evidence associated this phenotype with DNMT1-mediated hypermethylation in gene promoter region. Moreover, we showed that NCOA3 maintained the molecular signature of chondrocytes dedifferentiating in vitro or exposed to IL-1ß, nevertheless, NCOA3 appeared dispensable for preventing OA initiation, since NCOA3 loss did not trigger OA in young mice. Instead, NCOA3 loss promoted posttraumatic OA progression, and in parallel, enhanced NF-κB activation. Finally, the promoted posttraumatic OA progression was significantly retarded when administrated with NF-κB pathway inhibitor, suggesting that NCOA3 lose promotes posttraumatic OA at least partially by enhancing NF-κB activation. CONCLUSION: Thus, our findings indicate a critical role of NCOA3 in chondrocytes, and imply that manipulating NCOA3 might present a potential therapeutic approach to interfere OA progression.


Assuntos
Coativador 3 de Receptor Nuclear/metabolismo , Animais , Cartilagem Articular/citologia , Desdiferenciação Celular/efeitos dos fármacos , Células Cultivadas , Condrócitos/citologia , Condrócitos/metabolismo , DNA (Citosina-5-)-Metiltransferase 1/metabolismo , Metilação de DNA , Genótipo , Humanos , Interleucina-1beta/farmacologia , Joelho/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , NF-kappa B/metabolismo , Coativador 3 de Receptor Nuclear/antagonistas & inibidores , Coativador 3 de Receptor Nuclear/genética , Osteoartrite/patologia , Polimorfismo de Nucleotídeo Único , Interferência de RNA , RNA Interferente Pequeno/metabolismo
11.
Cancer Commun (Lond) ; 38(1): 53, 2018 08 13.
Artigo em Inglês | MEDLINE | ID: mdl-30103827

RESUMO

BACKGROUND: We previously found that overexpression of the gene known as amplified in breast cancer 1 (AIB1) was associated with lymph node metastasis and poor prognosis in patients with lung adenocarcinoma. However, the role of AIB1 in that malignancy remains unknown. The present study aimed to investigate the function of AIB1 in the process of lung adenocarcinoma cell metastasis. METHODS: A series of in vivo and in vitro assays were performed to elucidate the function of AIB1, while real-time PCR and Western blotting were utilized to identify the potential downstream targets of AIB1 in the process of lung adenocarcinoma metastasis. Rescue experiments and in vitro assays were performed to investigate whether the invasiveness of AIB1-induced lung adenocarcinoma was mediated by C-X-C motif chemokine receptor 4 (CXCR4). RESULTS: The ectopic overexpression of AIB1 in lung adenocarcinoma cells substantially enhanced cell migration and invasive abilities in vitro and tumor metastasis in vivo, whereas the depletion of AIB1 expression substantially inhibited lung adenocarcinoma cell migration and invasion. CXCR4 was identified as a potential downstream target of AIB1 in lung adenocarcinoma. The knockdown of AIB1 greatly reduced CXCR4 gene expression at both the transcription and protein levels, whereas the knockdown of CXCR4 in cells with AIB1 ectopic overexpression diminished AIB1-induced migration and invasion in vitro and tumor metastasis in vivo. Furthermore, we found a significant positive association between the expression of AIB1 and CXCR4 in lung adenocarcinoma patients (183 cases), and the co-overexpression of AIB1 and CXCR4 predicted the poorest prognosis. CONCLUSIONS: These findings suggest that AIB1 promotes the aggressiveness of lung adenocarcinoma in vitro and in vivo by upregulating CXCR4 and that it might be usable as a novel prognostic marker and/or therapeutic target for this disease.


Assuntos
Adenocarcinoma/genética , Regulação Neoplásica da Expressão Gênica , Neoplasias Pulmonares/genética , Coativador 3 de Receptor Nuclear/genética , Receptores CXCR4/genética , Células A549 , Adenocarcinoma/metabolismo , Adenocarcinoma/terapia , Animais , Linhagem Celular Tumoral , Feminino , Humanos , Estimativa de Kaplan-Meier , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/terapia , Masculino , Camundongos Endogâmicos BALB C , Camundongos Nus , Pessoa de Meia-Idade , Coativador 3 de Receptor Nuclear/metabolismo , Interferência de RNA , Terapêutica com RNAi/métodos , Receptores CXCR4/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto/métodos
12.
Crit Rev Immunol ; 38(3): 245-252, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30004860

RESUMO

Steroid receptor coactivator 3 (SRC-3) is a transcriptional coactivator that interacts with nuclear receptors such as the estrogen receptor and the androgen receptor and several other transcription factors to enhance their effects on target gene expression. SRC-3 plays important roles in many developmental, physiological, and pathologic events, including body growth, mammary gland development, energy homeostasis, inflammatory regulation, and cancer initiation and progression. SRC-3 has been suggested to be involved in host defense against bacterial pathogens. In this review, we summarize the roles of SRC-3 in host defense against peritoneal and enteric bacterial infection and discuss the potential clinical implications.


Assuntos
Infecções Bacterianas/imunologia , Inflamação/metabolismo , Glândulas Mamárias Humanas/fisiologia , Neoplasias/metabolismo , Coativador 3 de Receptor Nuclear/metabolismo , Animais , Metabolismo Energético , Homeostase , Humanos , Coativador 3 de Receptor Nuclear/genética , Receptores Estrogênicos/metabolismo
13.
Blood ; 132(9): 911-923, 2018 08 30.
Artigo em Inglês | MEDLINE | ID: mdl-29959189

RESUMO

Quiescence maintenance is an important property of hematopoietic stem cells (HSCs), whereas the regulatory factors and underlying mechanisms involved in HSC quiescence maintenance are not fully uncovered. Here, we show that steroid receptor coactivator 3 (SRC-3) is highly expressed in HSCs, and SRC-3-deficient HSCs are less quiescent and more proliferative, resulting in increased sensitivity to chemotherapy and irradiation. Moreover, the long-term reconstituting ability of HSCs is markedly impaired in the absence of SRC-3, and SRC-3 knockout (SRC-3-/-) mice exhibit a significant disruption of hematopoietic stem and progenitor cell homeostasis. Further investigations show that SRC-3 deficiency leads to enhanced mitochondrial metabolism, accompanied by overproduction of reactive oxygen species (ROS) in HSCs. Notably, the downstream target genes of peroxisome proliferator-activated receptor-coactivators 1α (PGC-1α) involved in the regulation of mitochondrial metabolism are significantly upregulated in SRC-3-deficient HSCs. Meanwhile, a significant decrease in the expression of histone acetyltransferase GCN5 accompanied by downregulation of PGC-1α acetylation is observed in SRC-3-null HSCs. Conversely, overexpression of GCN5 can inhibit SRC-3 deficiency-induced mitochondrial metabolism enhancement and ROS overproduction, thereby evidently rescuing the impairment of HSCs in SRC-3-/- mice. Collectively, our findings demonstrate that SRC-3 plays an important role in HSC quiescence maintenance by regulating mitochondrial metabolism.


Assuntos
Células-Tronco Hematopoéticas/metabolismo , Homeostase/fisiologia , Mitocôndrias/metabolismo , Coativador 3 de Receptor Nuclear/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Animais , Células-Tronco Hematopoéticas/citologia , Camundongos , Camundongos Knockout , Mitocôndrias/genética , Coativador 3 de Receptor Nuclear/genética , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/genética , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/metabolismo , Fatores de Transcrição de p300-CBP/genética , Fatores de Transcrição de p300-CBP/metabolismo
14.
Cell Rep ; 23(8): 2318-2329, 2018 05 22.
Artigo em Inglês | MEDLINE | ID: mdl-29791844

RESUMO

T helper 17 (Th17) cell development is programmed by the orphan nuclear receptor RORγt, but the underlying mechanism is not well understood. Nuclear receptor-mediated transcriptional activation depends on coactivators. Here, we show that steroid receptor coactivator-3 (SRC-3) critically regulates Th17 cell differentiation. Reduced incidence of experimental autoimmune encephalitis (EAE) associated with decreased Th17 cell generation in vivo was observed in mice with SRC-3 deletion specifically in T cells. In vitro, SRC-3 deficiency did not affect TGF-ß/IL-6-induced Th17 cell generation but severely impaired pathogenic Th17 differentiation induced by IL-1/IL-6/IL-23. Microarray analysis revealed that SRC-3 not only regulates IL-17A but also IL-1R1 expression. SRC-3 bound to Il17a and Il1r1 loci in a RORγt-dependent manner and was required for recruitment of the p300 acetyltransferase. Thus, SRC-3 is critical for RORγt-dependent gene expression in Th17 cell-driven autoimmune diseases.


Assuntos
Diferenciação Celular , Coativador 3 de Receptor Nuclear/metabolismo , Células Th17/citologia , Células Th17/imunologia , Animais , Polaridade Celular , Cromatina/metabolismo , Encefalomielite Autoimune Experimental/imunologia , Encefalomielite Autoimune Experimental/patologia , Loci Gênicos , Células HEK293 , Humanos , Interleucinas/metabolismo , Camundongos Transgênicos , Coativador 3 de Receptor Nuclear/deficiência , Membro 3 do Grupo F da Subfamília 1 de Receptores Nucleares/metabolismo , Ligação Proteica , Receptores de Interleucina-1/metabolismo
15.
Mol Cell ; 70(4): 679-694.e7, 2018 05 17.
Artigo em Inglês | MEDLINE | ID: mdl-29775582

RESUMO

Enhancers are thought to activate transcription by physically contacting promoters via looping. However, direct assays demonstrating these contacts are required to mechanistically verify such cellular determinants of enhancer function. Here, we present versatile cell-free assays to further determine the role of enhancer-promoter contacts (EPCs). We demonstrate that EPC is linked to mutually stimulatory transcription at the enhancer and promoter in vitro. SRC-3 was identified as a critical looping determinant for the estradiol-(E2)-regulated GREB1 locus. Surprisingly, the GREB1 enhancer and promoter contact two internal gene body SRC-3 binding sites, GBS1 and GBS2, which stimulate their transcription. Utilizing time-course 3C assays, we uncovered SRC-3-dependent dynamic chromatin interactions involving the enhancer, promoter, GBS1, and GBS2. Collectively, these data suggest that the enhancer and promoter remain "poised" for transcription via their contacts with GBS1 and GBS2. Upon E2 induction, GBS1 and GBS2 disengage from the enhancer, allowing direct EPC for active transcription.


Assuntos
Neoplasias da Mama/genética , Cromatina/metabolismo , Estrogênios/farmacologia , Regulação Neoplásica da Expressão Gênica , Coativador 3 de Receptor Nuclear/metabolismo , Transcrição Genética , Sítios de Ligação , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Cromatina/genética , Elementos Facilitadores Genéticos , Receptor alfa de Estrogênio/genética , Receptor alfa de Estrogênio/metabolismo , Feminino , Humanos , Coativador 3 de Receptor Nuclear/genética , Regiões Promotoras Genéticas , Ligação Proteica , Células Tumorais Cultivadas
16.
Nature ; 556(7700): 249-254, 2018 04.
Artigo em Inglês | MEDLINE | ID: mdl-29615789

RESUMO

Alterations in both cell metabolism and transcriptional programs are hallmarks of cancer that sustain rapid proliferation and metastasis 1 . However, the mechanisms that control the interaction between metabolic reprogramming and transcriptional regulation remain unclear. Here we show that the metabolic enzyme 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase 4 (PFKFB4) regulates transcriptional reprogramming by activating the oncogenic steroid receptor coactivator-3 (SRC-3). We used a kinome-wide RNA interference-based screening method to identify potential kinases that modulate the intrinsic SRC-3 transcriptional response. PFKFB4, a regulatory enzyme that synthesizes a potent stimulator of glycolysis 2 , is found to be a robust stimulator of SRC-3 that coregulates oestrogen receptor. PFKFB4 phosphorylates SRC-3 at serine 857 and enhances its transcriptional activity, whereas either suppression of PFKFB4 or ectopic expression of a phosphorylation-deficient Ser857Ala mutant SRC-3 abolishes the SRC-3-mediated transcriptional output. Functionally, PFKFB4-driven SRC-3 activation drives glucose flux towards the pentose phosphate pathway and enables purine synthesis by transcriptionally upregulating the expression of the enzyme transketolase. In addition, the two enzymes adenosine monophosphate deaminase-1 (AMPD1) and xanthine dehydrogenase (XDH), which are involved in purine metabolism, were identified as SRC-3 targets that may or may not be directly involved in purine synthesis. Mechanistically, phosphorylation of SRC-3 at Ser857 increases its interaction with the transcription factor ATF4 by stabilizing the recruitment of SRC-3 and ATF4 to target gene promoters. Ablation of SRC-3 or PFKFB4 suppresses breast tumour growth in mice and prevents metastasis to the lung from an orthotopic setting, as does Ser857Ala-mutant SRC-3. PFKFB4 and phosphorylated SRC-3 levels are increased and correlate in oestrogen receptor-positive tumours, whereas, in patients with the basal subtype, PFKFB4 and SRC-3 drive a common protein signature that correlates with the poor survival of patients with breast cancer. These findings suggest that the Warburg pathway enzyme PFKFB4 acts as a molecular fulcrum that couples sugar metabolism to transcriptional activation by stimulating SRC-3 to promote aggressive metastatic tumours.


Assuntos
Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Regulação Neoplásica da Expressão Gênica , Glucose/metabolismo , Coativador 3 de Receptor Nuclear/metabolismo , Fosfofrutoquinase-2/metabolismo , Ativação Transcricional , Fator 4 Ativador da Transcrição/metabolismo , Animais , Neoplasias da Mama/enzimologia , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Feminino , Glicólise , Humanos , Neoplasias Pulmonares/prevenção & controle , Neoplasias Pulmonares/secundário , Camundongos , Metástase Neoplásica , Via de Pentose Fosfato , Fosforilação , Fosfosserina/metabolismo , Prognóstico , Purinas/biossíntese , Purinas/metabolismo , Interferência de RNA , Receptores Estrogênicos/metabolismo , Transcetolase/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto
17.
Oncogene ; 37(24): 3243-3259, 2018 06.
Artigo em Inglês | MEDLINE | ID: mdl-29545602

RESUMO

Liver receptor homolog-1 (LRH1) has been shown to promote tumor proliferation and development. However, the functions of LRH1 in mediating cancer cells chemoresistance are still not clear. Here, we found LRH1 levels were significantly elevated in primary breast cancer tissues in patients who developed early recurrence. Similarly, adriamycin (ADR)-resistant breast cancer cell lines also exerted high LRH1 expression. Indeed, overexpression of LRH1 attenuated cytotoxicity of chemotherapeutic drugs ADR and cisplatin (DDP) in breast cancer cells in vitro and in nude mice tumor model. Comet and BrdU assays showed overexpression of LRH1 blocked breast cancer cells DNA damage by chemotherapeutic drug, whereas depletion of LRH1 enhanced DNA damage. Remarkably, knockdown of LRH1 decreased the levels and foci of DNA damage marker γH2AX induced by ADR and DDP. Furthermore, plasmid end-joining assay indicated that knockdown of LRH1 significantly decreased non-homologous end-joining (NHEJ)-mediated double-strand break (DSB) repair efficiencies. Afterwards, we provided evidences that LRH1 promoted MDC1 transcription by directly activating MDC1 promoter and therefore increased γH2AX levels. Importantly, a LRH1-binding site mapped between -1812 and -1804 bp of the proximal MDC1 promoter was identified. Moreover, LRH1 and MDC1 mRNA levels were positively correlated in recurrent breast cancer samples. These results implied LRH1 enhanced breast cancer cell chemoresistance by upregulating MDC1 and attenuating DNA damage. Additionally, we elucidated the coactivator NCOA3 acted synergistically with LRH1 to promote MDC1 expression and chemoresistance. Altogether, LRH1-MDC1 signaling might be considered as a novel molecular target for designing novel therapeutic regimen in chemotherapy resistance breast cancer.


Assuntos
Neoplasias da Mama/tratamento farmacológico , Resistencia a Medicamentos Antineoplásicos/genética , Proteínas Nucleares/genética , Receptores Citoplasmáticos e Nucleares/genética , Transativadores/genética , Animais , Apoptose/efeitos dos fármacos , Apoptose/genética , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Dano ao DNA/efeitos dos fármacos , Doxorrubicina/farmacologia , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Feminino , Regulação Neoplásica da Expressão Gênica , Histonas/metabolismo , Humanos , Células MCF-7 , Camundongos Nus , Proteínas Nucleares/metabolismo , Coativador 3 de Receptor Nuclear/genética , Coativador 3 de Receptor Nuclear/metabolismo , Regiões Promotoras Genéticas , Receptores Citoplasmáticos e Nucleares/metabolismo , Transativadores/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto
18.
Mol Cancer Res ; 16(4): 707-719, 2018 04.
Artigo em Inglês | MEDLINE | ID: mdl-29348189

RESUMO

Proline, glutamic acid, leucine-rich protein 1 (PELP1) is overexpressed in approximately 80% of invasive breast tumors. PELP1 dynamically shuttles between the nucleus and cytoplasm, but is primarily nuclear in normal breast tissue. However, altered localization of PELP1 to the cytoplasm is an oncogenic event that promotes breast cancer initiation and progression. Herein, interacting partners unique to cytoplasmic PELP1 and the mechanisms by which these interactions promote oncogenic PELP1 signaling were sought. AIB1 (amplified in breast cancer 1; also known as SRC-3 or NCOA3) was identified as a novel binding partner of cytoplasmic PELP1 in both estrogen receptor-positive (ER+) and ER-negative cell lines. Cytoplasmic PELP1 expression elevated basal phosphorylation levels (i.e., activation) of AIB1 at Thr24, enhanced ALDH+ tumorsphere formation, and upregulated specific target genes independently of hormone stimulation. Direct manipulation of AIB1 levels using shRNA abrogated cytoplasmic PELP1-induced tumorsphere formation and downregulated cytoplasmic PELP1-specific target genes. SI-2, an AIB1 inhibitor, limited the PELP1/AIB1 interaction and decreased cytoplasmic PELP1-induced tumorsphere formation. Similar results were observed in a murine-derived MMTV-AIB1 tumor cell line. Furthermore, in vivo syngeneic tumor studies revealed that PELP1 knockdown resulted in increased survival of tumor-bearing mice as compared with mice injected with control cells.Implications: These data demonstrate that cytoplasmic PELP1/AIB1-containing complexes function to promote advanced cancer phenotypes, including outgrowth of stem-like cells, associated with estrogen-independent breast cancer progression. Mol Cancer Res; 16(4); 707-19. ©2018 AACR.


Assuntos
Neoplasias da Mama/metabolismo , Proteínas Correpressoras/metabolismo , Citoplasma/metabolismo , Células-Tronco Neoplásicas/metabolismo , Coativador 3 de Receptor Nuclear/metabolismo , Receptores Estrogênicos/metabolismo , Fatores de Transcrição/metabolismo , Animais , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Proliferação de Células , Células Cultivadas , Proteínas Correpressoras/genética , Citoplasma/genética , Progressão da Doença , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Células MCF-7 , Camundongos , Modelos Biológicos , Transplante de Neoplasias , Células-Tronco Neoplásicas/patologia , Fosforilação , Fatores de Transcrição/genética
19.
Genome Med ; 10(1): 2, 2018 01 04.
Artigo em Inglês | MEDLINE | ID: mdl-29301589

RESUMO

BACKGROUND: The Ras pathway genes KRAS, BRAF, or ERBBs have somatic mutations in ~ 60% of human colorectal carcinomas. At present, it is unknown whether the remaining cases lack mutations activating the Ras pathway or whether they have acquired mutations in genes hitherto unknown to belong to the pathway. METHODS: To address the second possibility and extend the compendium of Ras pathway genes, we used genome-wide transposon mutagenesis of two human colorectal cancer cell systems deprived of their activating KRAS or BRAF allele to identify genes enabling growth in low glucose, a Ras pathway phenotype, when targeted. RESULTS: Of the 163 recurrently targeted genes in the two different genetic backgrounds, one-third were known cancer genes and one-fifth had links to the EGFR/Ras/MAPK pathway. When compared to cancer genome sequencing datasets, nine genes also mutated in human colorectal cancers were identified. Among these, stable knockdown of FOXO3, NCOA3, and TCF7L2 restored growth in low glucose but reduced MEK/MAPK phosphorylation, reduced anchorage-independent growth, and modulated expressions of GLUT1 and Ras pathway related proteins. Knockdown of NCOA3 and FOXO3 significantly decreased the sensitivity to cetuximab of KRAS mutant but not wild-type cells. CONCLUSIONS: This work establishes a proof-of-concept that human cell-based genome-wide forward genetic screens can assign genes to pathways with clinical importance in human colorectal cancer.


Assuntos
Neoplasias Colorretais/genética , Proteína Forkhead Box O3/genética , Testes Genéticos , Genoma Humano , Coativador 3 de Receptor Nuclear/genética , Transdução de Sinais/genética , Proteína 2 Semelhante ao Fator 7 de Transcrição/genética , Proteínas ras/metabolismo , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/genética , Cetuximab/farmacologia , Cetuximab/uso terapêutico , Neoplasias Colorretais/tratamento farmacológico , Neoplasias Colorretais/patologia , Elementos de DNA Transponíveis/genética , Resistencia a Medicamentos Antineoplásicos/genética , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Proteína Forkhead Box O3/metabolismo , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Técnicas de Silenciamento de Genes , Transportador de Glucose Tipo 1/genética , Transportador de Glucose Tipo 1/metabolismo , Humanos , Coativador 3 de Receptor Nuclear/metabolismo , Fenótipo , Fosforilação/efeitos dos fármacos , Proteômica , RNA Interferente Pequeno/metabolismo , Proteína 2 Semelhante ao Fator 7 de Transcrição/metabolismo
20.
Mol Oncol ; 12(3): 391-405, 2018 03.
Artigo em Inglês | MEDLINE | ID: mdl-29360267

RESUMO

Nuclear receptor coactivator 3 (NCOA3) is a transcriptional coactivator that has elevated expression in multiple tumor types, including colorectal cancer (CRC). However, the molecular mechanisms that regulate the tumorigenic functions of NCOA3 in CRC remain largely unknown. In this study, we aimed to discover and identify the novel regulatory proteins of NCOA3 and explore their mechanisms of action. Immunoprecipitation (IP) coupled with mass spectrometry (IP-MS) analysis was used to detect, identify, and verify the proteins that interacted with NCOA3 in CRC cells. The biological functions of the candidate proteins and the underlying molecular mechanism were investigated in CRC cells and mouse model in vitro and in vivo. The clinical significance of NCOA3 and its interaction partner protein in CRC patients was also studied. We identified mitotic arrest deficient 2-like protein 2 (MAD2L2, also known as MAD2B or REV7), with two signal peptide sequences of LIPLK and EVYPVGIFQK, to be an interaction partner of NCOA3. Overexpression of MAD2L2 suppressed the proliferation, migration, and clonogenicity of CRC cells by inducing the degradation of NCOA3. The mechanism study showed that increased MAD2L2 expression in CRC cells activated p38, which was required for the phosphorylation of NCOA3 that led to its ubiquitination and degradation by the proteasome. Moreover, we found that MAD2L2 predicted favorable prognosis in CRC patients. We have discovered a novel role of MAD2L2 in the regulation of NCOA3 degradation and proposed that MAD2L2 serves as a tumor suppressor in CRC.


Assuntos
Neoplasias Colorretais/patologia , Proteínas Mad2/metabolismo , Coativador 3 de Receptor Nuclear/metabolismo , Proteólise , Ubiquitinação , Animais , Linhagem Celular Tumoral , Neoplasias Colorretais/metabolismo , Feminino , Células HT29 , Humanos , Proteínas Mad2/genética , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Neoplasias Experimentais/metabolismo , Neoplasias Experimentais/patologia , Coativador 3 de Receptor Nuclear/genética , Prognóstico , Ensaios Antitumorais Modelo de Xenoenxerto
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