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1.
Cell ; 171(7): 1545-1558.e18, 2017 Dec 14.
Artigo em Inglês | MEDLINE | ID: mdl-29153836

RESUMO

mTORC1 is a signal integrator and master regulator of cellular anabolic processes linked to cell growth and survival. Here, we demonstrate that mTORC1 promotes lipid biogenesis via SRPK2, a key regulator of RNA-binding SR proteins. mTORC1-activated S6K1 phosphorylates SRPK2 at Ser494, which primes Ser497 phosphorylation by CK1. These phosphorylation events promote SRPK2 nuclear translocation and phosphorylation of SR proteins. Genome-wide transcriptome analysis reveals that lipid biosynthetic enzymes are among the downstream targets of mTORC1-SRPK2 signaling. Mechanistically, SRPK2 promotes SR protein binding to U1-70K to induce splicing of lipogenic pre-mRNAs. Inhibition of this signaling pathway leads to intron retention of lipogenic genes, which triggers nonsense-mediated mRNA decay. Genetic or pharmacological inhibition of SRPK2 blunts de novo lipid synthesis, thereby suppressing cell growth. These results thus reveal a novel role of mTORC1-SRPK2 signaling in post-transcriptional regulation of lipid metabolism and demonstrate that SRPK2 is a potential therapeutic target for mTORC1-driven metabolic disorders.


Assuntos
Regulação da Expressão Gênica , Lipogênese , Processamento Pós-Transcricional do RNA , Transdução de Sinais , Animais , Núcleo Celular/metabolismo , Colesterol/metabolismo , Ácidos Graxos/metabolismo , Feminino , Xenoenxertos , Humanos , Alvo Mecanístico do Complexo 1 de Rapamicina/metabolismo , Camundongos , Camundongos Nus , Transplante de Neoplasias , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Quinases S6 Ribossômicas 70-kDa/metabolismo
2.
Mol Cell ; 83(16): 3010-3026.e8, 2023 08 17.
Artigo em Inglês | MEDLINE | ID: mdl-37595559

RESUMO

The mechanistic target of rapamycin complex 1 (mTORC1) is a master regulator of cell growth that stimulates macromolecule synthesis through transcription, RNA processing, and post-translational modification of metabolic enzymes. However, the mechanisms of how mTORC1 orchestrates multiple steps of gene expression programs remain unclear. Here, we identify family with sequence similarity 120A (FAM120A) as a transcription co-activator that couples transcription and splicing of de novo lipid synthesis enzymes downstream of mTORC1-serine/arginine-rich protein kinase 2 (SRPK2) signaling. The mTORC1-activated SRPK2 phosphorylates splicing factor serine/arginine-rich splicing factor 1 (SRSF1), enhancing its binding to FAM120A. FAM120A directly interacts with a lipogenic transcription factor SREBP1 at active promoters, thereby bridging the newly transcribed lipogenic genes from RNA polymerase II to the SRSF1 and U1-70K-containing RNA-splicing machinery. This mTORC1-regulated, multi-protein complex promotes efficient splicing and stability of lipogenic transcripts, resulting in fatty acid synthesis and cancer cell proliferation. These results elucidate FAM120A as a critical transcription co-factor that connects mTORC1-dependent gene regulation programs for anabolic cell growth.


Assuntos
Arginina , Lipogênese , Proteína de Ligação a Elemento Regulador de Esterol 1 , Lipogênese/genética , Alvo Mecanístico do Complexo 1 de Rapamicina/genética , Fatores de Processamento de RNA , Proteína de Ligação a Elemento Regulador de Esterol 1/metabolismo , Humanos , Proteínas de Ligação a Elemento Regulador de Esterol/metabolismo
3.
Mol Cell ; 81(10): 2064-2075.e8, 2021 05 20.
Artigo em Inglês | MEDLINE | ID: mdl-33756105

RESUMO

Dysregulated mTORC1 signaling alters a wide range of cellular processes, contributing to metabolic disorders and cancer. Defining the molecular details of downstream effectors is thus critical for uncovering selective therapeutic targets. We report that mTORC1 and its downstream kinase S6K enhance eIF4A/4B-mediated translation of Wilms' tumor 1-associated protein (WTAP), an adaptor for the N6-methyladenosine (m6A) RNA methyltransferase complex. This regulation is mediated by 5' UTR of WTAP mRNA that is targeted by eIF4A/4B. Single-nucleotide-resolution m6A mapping revealed that MAX dimerization protein 2 (MXD2) mRNA contains m6A, and increased m6A modification enhances its degradation. WTAP induces cMyc-MAX association by suppressing MXD2 expression, which promotes cMyc transcriptional activity and proliferation of mTORC1-activated cancer cells. These results elucidate a mechanism whereby mTORC1 stimulates oncogenic signaling via m6A RNA modification and illuminates the WTAP-MXD2-cMyc axis as a potential therapeutic target for mTORC1-driven cancers.


Assuntos
Adenosina/análogos & derivados , Alvo Mecanístico do Complexo 1 de Rapamicina/metabolismo , Estabilidade de RNA , Adenosina/metabolismo , Animais , Sequência de Bases , Proteínas de Ciclo Celular/metabolismo , Linhagem Celular Tumoral , Proliferação de Células , Fatores de Iniciação em Eucariotos/metabolismo , Células HEK293 , Humanos , Masculino , Camundongos , Modelos Biológicos , Biossíntese de Proteínas , Proteínas Proto-Oncogênicas c-myc/metabolismo , Fatores de Processamento de RNA/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Proteínas Quinases S6 Ribossômicas/metabolismo , Transdução de Sinais
4.
Mol Cell ; 70(5): 949-960.e4, 2018 06 07.
Artigo em Inglês | MEDLINE | ID: mdl-29861159

RESUMO

The mammalian Target of Rapamycin Complex 1 (mTORC1)-signaling system plays a critical role in the maintenance of cellular homeostasis by sensing and integrating multiple extracellular and intracellular cues. Therefore, uncovering the effectors of mTORC1 signaling is pivotal to understanding its pathophysiological effects. Here we report that the transcription factor forkhead/winged helix family k1 (Foxk1) is a mediator of mTORC1-regulated gene expression. Surprisingly, Foxk1 phosphorylation is increased upon mTORC1 suppression, which elicits a 14-3-3 interaction, a reduction of DNA binding, and nuclear exclusion. Mechanistically, this occurs by mTORC1-dependent suppression of nuclear signaling by the Foxk1 kinase, Gsk3. This pathway then regulates the expression of multiple genes associated with glycolysis and downstream anabolic pathways directly modulated by Foxk1 and/or by Foxk1-regulated expression of Hif-1α. Thus, Foxk1 mediates mTORC1-driven metabolic rewiring, and it is likely to be critical for metabolic diseases where improper mTORC1 signaling plays an important role.


Assuntos
Reprogramação Celular , Metabolismo Energético , Fatores de Transcrição Forkhead/metabolismo , Quinase 3 da Glicogênio Sintase/metabolismo , Alvo Mecanístico do Complexo 1 de Rapamicina/metabolismo , Proteínas 14-3-3/metabolismo , Transporte Ativo do Núcleo Celular , Animais , Sítios de Ligação , Proliferação de Células , Regulação para Baixo , Fatores de Transcrição Forkhead/genética , Quinase 3 da Glicogênio Sintase/genética , Células HEK293 , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Alvo Mecanístico do Complexo 1 de Rapamicina/genética , Camundongos , Fosforilação , Ligação Proteica , Transdução de Sinais
5.
Proc Natl Acad Sci U S A ; 118(10)2021 03 09.
Artigo em Inglês | MEDLINE | ID: mdl-33649236

RESUMO

Mechanistic Target of Rapamycin Complex 1 (mTORC1) is a central regulator of cell growth and metabolism that senses and integrates nutritional and environmental cues with cellular responses. Recent studies have revealed critical roles of mTORC1 in RNA biogenesis and processing. Here, we find that the m6A methyltransferase complex (MTC) is a downstream effector of mTORC1 during autophagy in Drosophila and human cells. Furthermore, we show that the Chaperonin Containing Tailless complex polypeptide 1 (CCT) complex, which facilitates protein folding, acts as a link between mTORC1 and MTC. The mTORC1 activates the chaperonin CCT complex to stabilize MTC, thereby increasing m6A levels on the messenger RNAs encoding autophagy-related genes, leading to their degradation and suppression of autophagy. Altogether, our study reveals an evolutionarily conserved mechanism linking mTORC1 signaling with m6A RNA methylation and demonstrates their roles in suppressing autophagy.


Assuntos
Autofagia , Proteínas de Drosophila/metabolismo , Alvo Mecanístico do Complexo 1 de Rapamicina/metabolismo , Metiltransferases/metabolismo , Receptores Citoplasmáticos e Nucleares/metabolismo , Proteínas Repressoras/metabolismo , Transdução de Sinais , Animais , Linhagem Celular , Proteínas de Drosophila/genética , Drosophila melanogaster , Humanos , Alvo Mecanístico do Complexo 1 de Rapamicina/genética , Metilação , Metiltransferases/genética , Receptores Nucleares Órfãos , Estabilidade de RNA , Receptores Citoplasmáticos e Nucleares/genética , Proteínas Repressoras/genética
6.
Arterioscler Thromb Vasc Biol ; 36(9): 1772-81, 2016 09.
Artigo em Inglês | MEDLINE | ID: mdl-27417587

RESUMO

OBJECTIVE: Liver-enriched transcription factor cAMP-responsive element-binding protein H (CREBH) regulates plasma triglyceride clearance by inducing lipoprotein lipase cofactors, such as apolipoprotein A-IV (apoA-IV), apoA-V, and apoC-II. CREBH also regulates apoA-I transcription. This study aims to determine whether CREBH has a role in lipoprotein metabolism and development of atherosclerosis. APPROACH AND RESULTS: CREBH-deficient Creb3l3(-/-) mice were bred with Ldlr(-/-) mice creating Ldlr(-/-) Creb3l3(-/-) double knockout mice. Mice were fed on a high-fat and high-sucrose Western diet for 20 weeks. We showed that CREBH deletion in Ldlr(-/-) mice increased very low-density lipoprotein-associated triglyceride and cholesterol levels, consistent with the impairment of lipoprotein lipase-mediated triglyceride clearance in these mice. In contrast, high-density lipoprotein cholesterol levels were decreased in CREBH-deficient mice, which was associated with decreased production of apoA-I from the liver. The results indicate that CREBH directly activated Apoa1 gene transcription. Accompanied by the worsened atherogenic lipid profile, Ldlr(-/-) Creb3l3(-/-) mice developed significantly more atherosclerotic lesions in the aortas than Ldlr(-/-) mice. CONCLUSIONS: We identified CREBH as an important regulator of lipoprotein metabolism and suggest that increasing hepatic CREBH activity may be a novel strategy for prevention and treatment of atherosclerosis.


Assuntos
Aorta/metabolismo , Doenças da Aorta/metabolismo , Aterosclerose/metabolismo , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/deficiência , Receptores de LDL/deficiência , Animais , Aorta/patologia , Doenças da Aorta/genética , Doenças da Aorta/patologia , Apolipoproteína A-I/sangue , Apolipoproteína A-I/genética , Apolipoproteína B-100 , Apolipoproteínas B/sangue , Aterosclerose/genética , Aterosclerose/patologia , Biomarcadores/sangue , Linhagem Celular Tumoral , HDL-Colesterol/sangue , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/genética , Dieta Hiperlipídica , Sacarose Alimentar , Modelos Animais de Doenças , Predisposição Genética para Doença , Humanos , Hiperlipidemias/genética , Hiperlipidemias/metabolismo , Lipase Lipoproteica/metabolismo , Lipoproteínas VLDL/sangue , Fígado/metabolismo , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fenótipo , Placa Aterosclerótica , Receptores de LDL/genética , Índice de Gravidade de Doença , Transcrição Gênica , Ativação Transcricional , Triglicerídeos/sangue
7.
Proc Natl Acad Sci U S A ; 111(45): 15987-92, 2014 Nov 11.
Artigo em Inglês | MEDLINE | ID: mdl-25349418

RESUMO

Intrinsically disordered protein regions are widely distributed in the cytoplasmic domains of many transmembrane receptors. The cytoplasmic domain of a disintegrin and metalloprotease (ADAM)10, a transmembrane metalloprotease mediating ectodomain shedding of diverse membrane proteins, was recently suggested to mediate the homodimerization of ADAM10. Here we show that a recombinant cytoplasmic domain of ADAM10 (A10Cp) is unstructured as judged by its susceptibility to limited trypsin digestion and its circular dichroism spectrum. In comparison, recombinant transmembrane-cytoplasmic domain of ADAM10 (A10TmCp) reconstituted in dodecylphosphocholine (DPC) micelles exhibits much greater resistance to trypsin digestion, with its cytoplasmic domain taking on a significant ordered structure. FRET analysis demonstrates that, although A10Cp remains monomeric, A10TmCp forms a tight homodimer (K(d) ∼ 7 nM) in DPC micelles. Phospholipid-conjugated A10Cp dose-dependently inhibits formation of A10TmCp homodimer, whereas A10Cp achieves only limited inhibition. Placing the transmembrane and cytoplasmic domains of ADAM10, but not the transmembrane domain alone, in their native orientation in the inner membrane of Escherichia coli produces specific and strong dimerization signal in the AraC-based transcriptional reporter assay. A chimeric construct containing the otherwise monomeric transmembrane domain of L-selectin and the cytoplasmic domain of ADAM10 produces a similar dimerization signal. Overall, these results demonstrate that a transmembrane domain imparts a stable structure to the adjacent and intrinsically disordered cytoplasmic domain of ADAM10 to form a homodimer in the membrane. This finding advances our understanding of the regulatory mechanism of ADAMs and has general implications for membrane-protein interactions in the process of transmembrane signaling.


Assuntos
Proteínas ADAM/metabolismo , Secretases da Proteína Precursora do Amiloide/metabolismo , Membrana Celular/metabolismo , Proteínas de Membrana/metabolismo , Multimerização Proteica/fisiologia , Proteínas ADAM/química , Proteínas ADAM/genética , Proteína ADAM10 , Secretases da Proteína Precursora do Amiloide/química , Secretases da Proteína Precursora do Amiloide/genética , Membrana Celular/química , Membrana Celular/genética , Escherichia coli/química , Escherichia coli/genética , Escherichia coli/metabolismo , Humanos , Proteínas de Membrana/química , Proteínas de Membrana/genética , Estrutura Terciária de Proteína , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Transdução de Sinais/fisiologia
8.
Blood ; 123(21): 3344-53, 2014 May 22.
Artigo em Inglês | MEDLINE | ID: mdl-24713928

RESUMO

Thrombotic thrombocytopenic purpura (TTP) is a life-threatening disease that presents with thrombocytopenia, disseminated thrombosis, hemolytic anemia, and organ dysfunction. The etiology of TTP has revealed that patients share a deficiency in plasma protease a disintegrin and metalloproteinase with a thrombospondin type 1 motif, member 13 (ADAMTS13), the enzyme responsible for cleaving ultra-large von Willebrand factor (VWF) multimers into nonthrombogenic fragments. Therefore, existing TTP mouse models were developed by targeted disruption of the ADAMTS13 gene. ADAMTS13(-/-) mice are mostly asymptomatic in the absence of a trigger, as redundant proteases appear to take on VWF processing. As an alternative approach to creating one such model, we devised a strategy based on the expression of a cleavage-resistant VWF mutant in mice. The creation of a disulfide bond within the A2 domain of VWF was found to render VWF multimers resistant to proteolysis by plasma proteases under flow. Furthermore, mice expressing the murine VWF/p.S1494C-p.A1534C mutant present with symptoms characteristics of acute TTP such as thrombocytopenia, red cell shredding, accumulation of VWF-rich thrombi in the microvasculature, and advanced TTP symptoms such as renal dysfunction and splenomegaly. Because this model appears to faithfully emulate the pathophysiology of TTP, it should prove most useful in the study of microangiopathic diseases and their treatment.


Assuntos
Púrpura Trombocitopênica Trombótica/genética , Púrpura Trombocitopênica Trombótica/metabolismo , Fator de von Willebrand/química , Fator de von Willebrand/genética , Sequência de Aminoácidos , Animais , Dissulfetos/química , Dissulfetos/metabolismo , Células HEK293 , Humanos , Hidrólise , Rim/metabolismo , Rim/fisiopatologia , Camundongos , Camundongos Endogâmicos C57BL , Modelos Moleculares , Dados de Sequência Molecular , Mutação , Peptídeo Hidrolases/metabolismo , Estrutura Terciária de Proteína , Púrpura Trombocitopênica Trombótica/patologia , Fator de von Willebrand/metabolismo
9.
Bioorg Med Chem Lett ; 24(11): 2560-4, 2014 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-24768446

RESUMO

Among many signals to regulate hypoxia inducible factor 1α (HIF-1α), sphingosine kinase 1 (SPHK1) is also involved in various biological activities such as cell growth, survival, invasion, angiogenesis, and carcinogenesis. Thus, in the present study, molecular mechanisms of coumestrol were investigated on the SPHK1 and HIF-1α signaling pathway in hypoxic PC-3 prostate cancer cells. Coumestrol significantly suppressed SPHK1 activity and accumulation of HIF-1α in a time- and concentration-dependent manner in hypoxic PC-3 cells. In addition, coumestrol inhibited the phosphorylation status of AKT and glycogen synthase kinase-3ß (GSK 3ß) signaling involved in cancer metabolism. Furthermore, SPHK1 siRNA transfection, sphigosine kinase inhibitor (SKI), reactive oxygen species (ROS) enhanced the inhibitory effect of coumestrol on the accumulation of HIF-1α and the expression of pAKT and pGSK 3ß in hypoxic PC-3 cells by combination index. Overall, our findings suggest that coumestrol suppresses the accumulation of HIF-1α via suppression of SPHK1 pathway in hypoxic PC-3 cells.


Assuntos
Cumestrol/farmacologia , Subunidade alfa do Fator 1 Induzível por Hipóxia/antagonistas & inibidores , Fosfotransferases (Aceptor do Grupo Álcool)/antagonistas & inibidores , Espécies Reativas de Oxigênio/antagonistas & inibidores , Linhagem Celular Tumoral , Cumestrol/química , Relação Dose-Resposta a Droga , Humanos , Estrutura Molecular , Fosfotransferases (Aceptor do Grupo Álcool)/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Relação Estrutura-Atividade
10.
BMC Complement Altern Med ; 13: 189, 2013 Jul 24.
Artigo em Inglês | MEDLINE | ID: mdl-23883349

RESUMO

BACKGROUND: Though Mica, a thin and sheet like mineral, has been used as a mineral medicine for treatment of bleeding, dysentery and inflammation in traditional medicine including Ayurveda, the biological evidences of Mica were not clearly elucidated so far. Thus, in the present study, the antitumor mechanism of particled Mica (STB-HO) was examined in colorectal cancers. METHODS: Athymic nude mice were inoculated with HCT116 colon cancer cells and orally administered STB-HO daily for 41 days, and HCT116 and human umbilical vein endothelial cells (HUVECs) were treated with STB-HO for 0 ~ 24 hours to perform immunoblotting, cytotoxicity assay, FACs analysis and measurement of matrix metalloproteinase 9 (MMP-9) secretion and other experiments. Significant differences of all date were evaluated using Student's t-test and a Turkey-Kramer multiple-comparison post test. RESULTS: STB-HO significantly suppressed the tumor volume and weight in athymic nude mice inoculated with HCT116 cells at a dose of 100 mg/kg. Thus, the in vivo antitumor mechanism of STB-HO was to elucidated in vitro as well. STB-HO exerted cytotoxicity in HCT116, SW620 and HCT15 colorectal cancer cells. Also, STB-HO increased G1 cell population in a time and concentration dependent manner, enhanced the expression of p21, p27, p53 as cyclin dependent kinase (CDK) inhibitors, attenuated the expression of proliferating cell nuclear antigen (PCNA) and cyclin D1 and also reduced the production of vascular endothelial growth factor (VEGF) and matrix metalloproteinase 9 (MMP-9) in HCT116 cells. Consistently, STB-HO suppressed the phosphorylation of VEGFR2 in HCT116, SW620 and HCT15 cells. Also, STB-HO inhibited the VEGF mediated proliferation and also attenuated the phosphorylation of VEGFR2 and Akt in human umbilical vein endothelial cells (HUVECs). CONCLUSIONS: Collectively, these findings suggest that STB-HO has chemopreventive potential via G1 arrest and inhibition of proliferation and VEGFR2 in HCT116 colorectal cancer cells.


Assuntos
Silicatos de Alumínio/farmacologia , Silicatos de Alumínio/uso terapêutico , Antineoplásicos/uso terapêutico , Neoplasias Colorretais/tratamento farmacológico , Pontos de Checagem da Fase G1 do Ciclo Celular/efeitos dos fármacos , Fator A de Crescimento do Endotélio Vascular/metabolismo , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/metabolismo , Animais , Antineoplásicos/farmacologia , Proliferação de Células , Neoplasias Colorretais/metabolismo , Feminino , Células HCT116 , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Proteínas Proto-Oncogênicas c-akt/metabolismo , Veias Umbilicais
11.
Aging Cell ; 22(2): e13760, 2023 02.
Artigo em Inglês | MEDLINE | ID: mdl-36567449

RESUMO

Aging-associated microbial dysbiosis exacerbates various disorders and dysfunctions, and is a major contributor to morbidity and mortality in the elderly, but the underlying cause of this aging-related syndrome is confusing. SIRT6 knockout (SIRT6 KO) mice undergo premature aging and succumb to death by 4 weeks, and are therefore useful as a premature aging research model. Here, fecal microbiota transplantation from SIRT6 KO mice into wild-type (WT) mice phenocopies the gut dysbiosis and premature aging observed in SIRT6 KO mice. Conversely, an expanded lifespan was observed in SIRT6 KO mice when transplanted with microbiota from WT mice. Antibiotic cocktail treatment attenuated inflammation and cell senescence in KO mice, directly suggesting that gut dysbiosis contributes to the premature aging of SIRT6 KO mice. Increased Enterobacteriaceae translocation, driven by the overgrowth of Escherichia coli, is the likely mechanism for the premature aging effects of microbiome dysregulation, which could be reversed by a high-fat diet. Our results provide a mechanism for the causal link between gut dysbiosis and aging, and support a beneficial effect of a high-fat diet for correcting gut dysbiosis and alleviating premature aging. This study provides a rationale for the integration of microbiome-based high-fat diets into therapeutic interventions against aging-associated diseases.


Assuntos
Senilidade Prematura , Microbioma Gastrointestinal , Sirtuínas , Animais , Camundongos , Senilidade Prematura/genética , Dieta Hiperlipídica , Disbiose/etiologia , Enterobacteriaceae , Camundongos Endogâmicos C57BL
12.
Sci Adv ; 8(20): eabm8786, 2022 05 20.
Artigo em Inglês | MEDLINE | ID: mdl-35594343

RESUMO

Serine/one-carbon metabolism provides critical resources for nucleotide biosynthesis and epigenetic maintenance and is thus necessary in cancer cell growth, although the detailed regulatory mechanisms remain unclear. We uncover a critical role of glycogen synthase kinase 3 (GSK3) in regulating the expression of serine/one-carbon metabolic enzymes. Nuclear enrichment of GSK3 significantly suppresses genes that mediate de novo serine synthesis, including PHGDH, PSAT1, PSPH, and one-carbon metabolism, including SHMT2 and MTHFD2. FRAT1 promotes nuclear exclusion of GSK3, enhances serine/one-carbon metabolism, and, as a result, confers cell vulnerability to inhibitors that target this metabolic process such as SHIN1, a specific SHMT1/2 inhibitor. Furthermore, pharmacological or genetic suppression of GSK3 promotes serine/one-carbon metabolism and exhibits a significant synergistic effect in combination with SHIN1 in suppressing cancer cell proliferation in cultured cells and in vivo. Our observations indicate that inhibition of nuclear GSK3 signaling creates a vulnerability, which results in enhanced efficacy of serine/one-carbon metabolism inhibitors for the treatment of cancer.


Assuntos
Neoplasias Pulmonares , Serina , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Carbono/metabolismo , Quinase 3 da Glicogênio Sintase , Humanos , Neoplasias Pulmonares/genética , Proteínas Proto-Oncogênicas/metabolismo , Transdução de Sinais
13.
Nat Commun ; 13(1): 6239, 2022 10 20.
Artigo em Inglês | MEDLINE | ID: mdl-36266345

RESUMO

The systemic metabolic shifts that occur during aging and the local metabolic alterations of a tumor, its stroma and their communication cooperate to establish a unique tumor microenvironment (TME) fostering cancer progression. Here, we show that methylmalonic acid (MMA), an aging-increased oncometabolite also produced by aggressive cancer cells, activates fibroblasts in the TME, which reciprocally secrete IL-6 loaded extracellular vesicles (EVs) that drive cancer progression, drug resistance and metastasis. The cancer-associated fibroblast (CAF)-released EV cargo is modified as a result of reactive oxygen species (ROS) generation and activation of the canonical and noncanonical TGFß signaling pathways. EV-associated IL-6 functions as a stroma-tumor messenger, activating the JAK/STAT3 and TGFß signaling pathways in tumor cells and promoting pro-aggressive behaviors. Our findings define the role of MMA in CAF activation to drive metastatic reprogramming, unveiling potential therapeutic avenues to target MMA at the nexus of aging, the tumor microenvironment and metastasis.


Assuntos
Fibroblastos Associados a Câncer , Vesículas Extracelulares , Neoplasias , Humanos , Fibroblastos Associados a Câncer/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Ácido Metilmalônico/metabolismo , Interleucina-6/metabolismo , Microambiente Tumoral , Neoplasias/patologia , Vesículas Extracelulares/metabolismo , Fator de Crescimento Transformador beta/metabolismo
14.
J Pineal Res ; 51(1): 87-93, 2011 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-21392092

RESUMO

Sphingosine kinase 1 (SPHK1) is a newly discovered modulator of hypoxia inducible factor 1α (HIF-1α) with various biological activities such as cell growth, survival, invasion, angiogenesis, and carcinogenesis. Thus, in the present study, the biological mechanisms of melatonin were elucidated in association with SPHK1 pathway in PC-3 prostate cancer cells under hypoxia. Melatonin inhibited the stability of HIF-1α in a time- and concentration- dependent manners. Also, melatonin decreased SPHK1 activity in PC-3 cells during hypoxia. Furthermore, melatonin suppressed AKT/glycogen synthase kinase-3ß (GSK-3ß) signaling pathway, which stabilizes HIF-1α via inhibition of von Hippel-Lindau tumor suppressor protein. Consistently, siRNA-SPHK1 and sphingosine kinase inhibitor (SKI) effectively blocked the expression of HIF-1α, phospho-AKT and vascular endothelial growth factor (VEGF) production in PC-3 cells under hypoxia, suggesting the role of SPHK1 in melatonin-inhibited HIF-1α accumulation. Moreover, reactive oxygen species (ROS) scavenger N-acteylcysteine enhanced melatonin-inhibited HIF-1α expression and SPHK1 activity. Overall, our findings suggest that melatonin suppresses HIF-1α accumulation via inhibition of SPHK1 pathway and ROS generation in PC-3 cells under hypoxia.


Assuntos
Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Melatonina/farmacologia , Redes e Vias Metabólicas/efeitos dos fármacos , Fosfotransferases (Aceptor do Grupo Álcool)/metabolismo , Neoplasias da Próstata/metabolismo , Hipóxia Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Expressão Gênica/efeitos dos fármacos , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/antagonistas & inibidores , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Masculino , Neovascularização Patológica/enzimologia , Neovascularização Patológica/metabolismo , Neovascularização Patológica/patologia , Fosfotransferases (Aceptor do Grupo Álcool)/antagonistas & inibidores , Neoplasias da Próstata/irrigação sanguínea , Neoplasias da Próstata/patologia , Espécies Reativas de Oxigênio/metabolismo , Estatísticas não Paramétricas , Fator A de Crescimento do Endotélio Vascular/metabolismo
15.
Br J Nutr ; 104(2): 180-8, 2010 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-20487577

RESUMO

The antioxidant activity of lemon balm (Melissa officinalis) essential oil (LBEO) on 2,2-diphenyl-1-picrylhydrazyl (DPPH) radicals and its hypoglycaemic effect in db/db mice were investigated. LBEO scavenged 97 % of DPPH radicals at a 270-fold dilution. Mice administered LBEO (0.015 mg/d) for 6 weeks showed significantly reduced blood glucose (65 %; P < 0.05) and TAG concentrations, improved glucose tolerance, as assessed by an oral glucose tolerance test, and significantly higher serum insulin levels, compared with the control group. The hypoglycaemic mechanism of LBEO was further explored via gene and protein expression analyses using RT-PCR and Western blotting, respectively. Among all glucose metabolism-related genes studied, hepatic glucokinase and GLUT4, as well as adipocyte GLUT4, PPAR-gamma, PPAR-alpha and SREBP-1c expression, were significantly up-regulated, whereas glucose-6-phosphatase and phosphoenolpyruvate carboxykinase expression was down-regulated in the livers of the LBEO group. The results further suggest that LBEO administered at low concentrations is an efficient hypoglycaemic agent, probably due to enhanced glucose uptake and metabolism in the liver and adipose tissue and the inhibition of gluconeogenesis in the liver.


Assuntos
Glicemia/efeitos dos fármacos , Diabetes Mellitus Tipo 2/enzimologia , Metabolismo dos Lipídeos/efeitos dos fármacos , Melissa/química , Óleos Voláteis/farmacologia , Óleos de Plantas/farmacologia , Animais , Diabetes Mellitus Tipo 2/tratamento farmacológico , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Glucoquinase/efeitos dos fármacos , Glucoquinase/genética , Glucoquinase/metabolismo , Teste de Tolerância a Glucose , Proteínas Facilitadoras de Transporte de Glucose/efeitos dos fármacos , Proteínas Facilitadoras de Transporte de Glucose/genética , Proteínas Facilitadoras de Transporte de Glucose/metabolismo , Glucose-6-Fosfatase/efeitos dos fármacos , Glucose-6-Fosfatase/genética , Glucose-6-Fosfatase/metabolismo , Insulina/sangue , Camundongos , Óleos Voláteis/química , Óleos Voláteis/uso terapêutico , Receptores Ativados por Proliferador de Peroxissomo/efeitos dos fármacos , Receptores Ativados por Proliferador de Peroxissomo/genética , Receptores Ativados por Proliferador de Peroxissomo/metabolismo , Fosfoenolpiruvato Carboxiquinase (GTP)/efeitos dos fármacos , Fosfoenolpiruvato Carboxiquinase (GTP)/genética , Fosfoenolpiruvato Carboxiquinase (GTP)/metabolismo , Fitoterapia , Óleos de Plantas/química , Óleos de Plantas/uso terapêutico , Proteína de Ligação a Elemento Regulador de Esterol 1/efeitos dos fármacos , Proteína de Ligação a Elemento Regulador de Esterol 1/genética , Proteína de Ligação a Elemento Regulador de Esterol 1/metabolismo
17.
Sci Rep ; 6: 27938, 2016 06 15.
Artigo em Inglês | MEDLINE | ID: mdl-27301791

RESUMO

Adipose tissue lipolysis produces glycerol and nonesterified fatty acids (NEFA) that serve as energy sources during nutrient scarcity. Adipose tissue lipolysis is tightly regulated and excessive lipolysis causes hepatic steatosis, as NEFA released from adipose tissue constitutes a major source of TG in the liver of patients with nonalcoholic fatty liver diseases. Here we show that the liver-enriched transcription factor CREBH is activated by TG accumulation and induces FGF21, which suppresses adipose tissue lipolysis, ameliorating hepatic steatosis. CREBH-deficient mice developed severe hepatic steatosis due to increased adipose tissue lipolysis, when fasted or fed a high-fat low-carbohydrate ketogenic diet. FGF21 production was impaired in CREBH-deficient mice, and adenoviral overexpression of FGF21 suppressed adipose tissue lipolysis and improved hepatic steatosis in these mice. Thus, our results uncover a negative feedback loop in which CREBH regulates NEFA flux from adipose tissue to the liver via FGF21.


Assuntos
Tecido Adiposo/patologia , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Fígado Gorduroso/patologia , Fatores de Crescimento de Fibroblastos/metabolismo , Lipólise , Animais , Retroalimentação Fisiológica , Camundongos Endogâmicos C57BL , Camundongos Knockout
18.
Mol Cell Biol ; 35(16): 2761-70, 2015 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-26031337

RESUMO

The unfolded protein response (UPR) regulates endoplasmic reticulum (ER) homeostasis and protects cells from ER stress. IRE1α is a central regulator of the UPR that activates the transcription factor XBP1s through an unconventional splicing mechanism using its endoribonuclease activity. IRE1α also cleaves certain mRNAs containing XBP1-like secondary structures to promote the degradation of these mRNAs, a process known as regulated IRE1α-dependent decay (RIDD). We show here that the mRNA of CReP/Ppp1r15b, a regulatory subunit of eukaryotic translation initiation factor 2α (eIF2α) phosphatase, is a RIDD substrate. eIF2α plays a central role in the integrated stress response by mediating the translational attenuation to decrease the stress level in the cell. CReP expression was markedly suppressed in XBP1-deficient mice livers due to hyperactivated IRE1α. Decreased CReP expression caused the induction of eIF2α phosphorylation and the attenuation of protein synthesis in XBP1-deficient livers. ER stress also suppressed CReP expression in an IRE1α-dependent manner, which increased eIF2α phosphorylation and consequently attenuated protein synthesis. Taken together, the results of our study reveal a novel function of IRE1α in the regulation of eIF2α phosphorylation and the translational control.


Assuntos
Endorribonucleases/metabolismo , Fator de Iniciação 2 em Eucariotos/metabolismo , Biossíntese de Proteínas , Proteína Fosfatase 1/genética , Proteínas Serina-Treonina Quinases/metabolismo , Estabilidade de RNA , RNA Mensageiro/metabolismo , Animais , Proteínas de Ligação a DNA/genética , Regulação para Baixo , Estresse do Retículo Endoplasmático , Técnicas de Inativação de Genes , Fígado/metabolismo , Camundongos Endogâmicos C57BL , Fosforilação , RNA Mensageiro/química , RNA Mensageiro/genética , Fatores de Transcrição de Fator Regulador X , Fatores de Transcrição/genética , Proteína 1 de Ligação a X-Box
19.
J Mol Biol ; 425(18): 3549-62, 2013 Sep 23.
Artigo em Inglês | MEDLINE | ID: mdl-23796515

RESUMO

Moesin and calmodulin (CaM) jointly associate with the cytoplasmic domain of l-selectin in the cell to modulate the function and ectodomain shedding of l-selectin. Using fluorescence spectroscopy, we have examined the association of moesin FERM domain with the recombinant transmembrane and cytoplasmic domains of l-selectin (CLS) reconstituted in model phospholipid liposomes. The dissociation constant of moesin FERM domain to CLS in the phosphatidylcholine liposome is about 300nM. In contrast to disrupting the CaM association with CLS, inclusion of anionic phosphatidylserine lipids in the phosphatidylcholine liposome increased the apparent binding affinity of moesin FERM domain for CLS. Using the environmentally sensitive fluorescent probe attached to the cytoplasmic domain of CLS and the nitroxide quencher attached to the lipid bilayer, we showed that the association of moesin FERM domain induced the desorption of the basic-rich cytoplasmic domain of CLS from the anionic membrane surface, which enabled subsequent association of CaM to the cytoplasmic domain of CLS. These results have elucidated the molecular basis for the moesin/l-selectin/CaM ternary complex and suggested an important role of phospholipids in modulating l-selectin function and shedding.


Assuntos
Aminoácidos Básicos/química , Membrana Celular/metabolismo , Citoplasma/metabolismo , Selectina L/química , Selectina L/metabolismo , Proteínas dos Microfilamentos/química , Sequência de Aminoácidos , Ânions/química , Ânions/metabolismo , Calmodulina/química , Calmodulina/metabolismo , Membrana Celular/química , Humanos , Bicamadas Lipídicas/química , Bicamadas Lipídicas/metabolismo , Lipossomos/metabolismo , Proteínas dos Microfilamentos/metabolismo , Proteínas dos Microfilamentos/fisiologia , Modelos Biológicos , Modelos Moleculares , Dados de Sequência Molecular , Ligação Proteica , Estrutura Terciária de Proteína/fisiologia , Espectrometria de Fluorescência
20.
Artigo em Inglês | MEDLINE | ID: mdl-23589723

RESUMO

Here, antitumor mechanism of cinnamaldehyde derivative CB-PIC was elucidated in human SW620 colon cancer cells. CB-PIC significantly exerted cytotoxicity, increased sub-G1 accumulation, and cleaved PARP with apoptotic features, while it enhanced the phosphorylation of AMPK alpha and ACC as well as activated the ERK in hypoxic SW620 cells. Furthermore, CB-PIC suppressed the expression of HIF1 alpha, Akt, and mTOR and activated the AMPK phosphorylation in hypoxic SW620 cells. Conversely, silencing of AMPK α blocked PARP cleavage and ERK activation induced by CB-PIC, while ERK inhibitor PD 98059 attenuated the phosphorylation of AMPK α in hypoxic SW620 cells, implying cross-talk between ERK and AMPK α . Furthermore, cotreatment of CB-PIC and metformin enhanced the inhibition of HIF1 α and Akt/mTOR and the activation of AMPK α and pACC in hypoxic SW620 cells. In addition, CB-PIC suppressed the growth of SW620 cells inoculated in BALB/c athymic nude mice, and immunohistochemistry revealed that CB-PIC treatment attenuated the expression of Ki-67, CD34, and CAIX and increased the expression of pAMPK α in CB-PIC-treated group. Interestingly, CP-PIC showed better antitumor activity in SW620 colon cancer cells under hypoxia than under normoxia, since it may be applied to chemoresistance. Overall, our findings suggest that activation of AMPK α and ERK mediates CB-PIC-induced apoptosis in hypoxic SW620 colon cancer cells.

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