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1.
Development ; 145(14)2018 07 26.
Artigo em Inglês | MEDLINE | ID: mdl-29986870

RESUMO

Erk5 belongs to the mitogen-activated protein kinase (MAPK) family. Following its phosphorylation by Mek5, Erk5 modulates several signaling pathways in a number of cell types. In this study, we demonstrated that Erk5 inactivation in mesenchymal cells causes abnormalities in skeletal development by inducing Sox9, an important transcription factor of skeletogenesis. We further demonstrate that Erk5 directly phosphorylates and activates Smurf2 (a ubiquitin E3 ligase) at Thr249, which promotes the proteasomal degradation of Smad proteins and phosphorylates Smad1 at Ser206 in the linker region known to trigger its proteasomal degradation by Smurf1. Smads transcriptionally activated the expression of Sox9 in mesenchymal cells. Accordingly, removal of one Sox9 allele in mesenchymal cells from Erk5-deficient mice rescued some abnormalities of skeletogenesis. These findings highlight the importance of the Mek5-Erk5-Smurf-Smad-Sox9 axis in mammalian skeletogenesis.


Assuntos
Proteína Quinase 7 Ativada por Mitógeno/metabolismo , Osteogênese , Fatores de Transcrição SOX9/metabolismo , Transdução de Sinais , Proteínas Smad/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Animais , Diferenciação Celular , Condrogênese , Humanos , Mesoderma/citologia , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fosforilação , Proteólise , Crânio/anormalidades , Ubiquitina/metabolismo , Ubiquitinação
2.
Stem Cell Reports ; 11(1): 228-241, 2018 07 10.
Artigo em Inglês | MEDLINE | ID: mdl-30008325

RESUMO

The mechanistic/mammalian target of rapamycin complex 1 (mTORC1) regulates cellular function in various cell types. Although the role of mTORC1 in skeletogenesis has been investigated previously, here we show a critical role of mTORC1/4E-BPs/SOX9 axis in regulating skeletogenesis through its expression in undifferentiated mesenchymal cells. Inactivation of Raptor, a component of mTORC1, in limb buds before mesenchymal condensations resulted in a marked loss of both cartilage and bone. Mechanistically, we demonstrated that mTORC1 selectively controls the RNA translation of Sox9, which harbors a 5' terminal oligopyrimidine tract motif, via inhibition of the 4E-BPs. Indeed, introduction of Sox9 or a knockdown of 4E-BP1/2 in undifferentiated mesenchymal cells markedly rescued the deficiency of the condensation observed in Raptor-deficient mice. Furthermore, introduction of the Sox9 transgene rescued phenotypes of deficient skeletal growth in Raptor-deficient mice. These findings highlight a critical role of mTORC1 in mammalian skeletogenesis, at least in part, through translational control of Sox9 RNA.


Assuntos
Alvo Mecanístico do Complexo 1 de Rapamicina/metabolismo , Osteogênese/genética , Biossíntese de Proteínas , Fatores de Transcrição SOX9/genética , Esqueleto/metabolismo , Animais , Diferenciação Celular/genética , Expressão Gênica , Camundongos , Camundongos Transgênicos , Fenótipo , Fatores de Transcrição SOX9/metabolismo , Esqueleto/embriologia
3.
Pharmacology ; 101(1-2): 64-71, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29065407

RESUMO

The availability of amino acid in the brown adipose tissue (BAT) has been shown to be altered under various conditions; however, little is known about the possible expression and pivotal role of amino acid transporters in BAT under physiological and pathological conditions. The present study comprehensively investigated whether amino acid transporters are regulated by obesogenic conditions in BAT in vivo. Moreover, we investigated the mechanism underlying the regulation of the expression of amino acid transporters by various stressors in brown adipocytes in vitro. The expression of solute carrier family 38 member 1 (Slc38a1; gene encoding sodium-coupled neutral amino acid transporter 1) was preferentially upregulated in the BAT of both genetic and acquired obesity mice in vivo. Moreover, the expression of Slc38a1 was induced by hypoxic stress through hypoxia-inducible factor-1α, which is a master transcription factor of the adaptive response to hypoxic stress, in brown adipocytes in vitro. These results indicate that Slc38a1 is an obesity-associated gene in BAT and a hypoxia-responsive gene in brown adipocytes.


Assuntos
Adipócitos Marrons/metabolismo , Sistema A de Transporte de Aminoácidos/genética , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Hipóxia/genética , Obesidade/genética , Animais , Linhagem Celular , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Obesos , RNA Interferente Pequeno/genética
4.
Biol Pharm Bull ; 40(7): 1116-1120, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28674255

RESUMO

ß-Cryptoxanthin, which is primarily obtained from citrus fruits such as Satsuma mandarins, is a major carotenoid routinely found in human serum. Recently, we demonstrated that daily oral intake of ß-cryptoxanthin prevented ovariectomy-induced bone loss and ameliorated neuropathic pain in mice. Although ß-cryptoxanthin exerts preventive effects on various lifestyle-related diseases, there have been no studies on the effect of ß-cryptoxanthin on the development of osteoarthritis, the most common degenerative joint disease, which frequently leads to loss of ability and stiffness in the elderly. Here we showed that daily oral administration of ß-cryptoxanthin significantly prevented the development of osteoarthritis developed by surgically inducing knee joint instability in mice in vivo. Furthermore, in vitro experiments revealed that ß-cryptoxanthin markedly inhibited the expression of inflammatory cytokines and enzymes critical for the degradation of the extracellular matrix in primary chondrocytes. Our results suggest that oral supplementation of ß-cryptoxanthin would be beneficial for the maintenance of joint health and as prophylaxis against osteoarthritis.


Assuntos
beta-Criptoxantina/uso terapêutico , Osteoartrite/prevenção & controle , Animais , beta-Criptoxantina/administração & dosagem , Condrócitos/metabolismo , Citocinas/metabolismo , Mediadores da Inflamação/metabolismo , Camundongos , Osteoartrite/tratamento farmacológico
5.
Biosci Biotechnol Biochem ; 81(5): 1014-1017, 2017 May.
Artigo em Inglês | MEDLINE | ID: mdl-28110620

RESUMO

ß-cryptoxanthin, a xanthophyll carotenoid, exerts preventive effects on various lifestyle-related diseases. Here, we found that daily oral administration of ß-cryptoxanthin significantly ameliorated the development of tactile allodynia following spinal nerve injury but was ineffective in mechanical allodynia in an inflammatory pain model in mice. Our results suggest that ß-cryptoxanthin supplementation would be beneficial for the prophylaxis of neuropathic pain.


Assuntos
beta-Criptoxantina/administração & dosagem , beta-Criptoxantina/farmacologia , Neuralgia/tratamento farmacológico , Administração Oral , Animais , beta-Criptoxantina/uso terapêutico , Suplementos Nutricionais , Masculino , Camundongos
6.
FEBS J ; 284(5): 784-795, 2017 03.
Artigo em Inglês | MEDLINE | ID: mdl-28107769

RESUMO

Sympathetic tone activates the function of classical brown adipocytes, which constitutively exist in the brown adipose tissue (BAT), and inducible brown adipocytes (so-called beige adipocytes), which sporadically reside within the white adipose tissue (WAT). Here we identified the transcriptional modulator interferon-related developmental regulator 1 (Ifrd1) as a negative regulator of thermogenic and mitochondrial gene expression in brown adipocytes. Ifrd1 expression was markedly induced by cold exposure and administration of CL-316243 (a ß3 adrenergic agonist) in interscapular brown adipose and inguinal subcutaneous WATs, but not in epididymal visceral WAT, in vivo. Adrenergic stimulation also induced Ifrd1 expression in brown adipocytes in a cAMP responsive element binding protein-dependent manner in vitro. CL-316243 injection markedly elevated thermogenic and mitochondrial gene expression, including peroxisome proliferator-activated receptor γ coactivator 1α (Pgc1a) in the subcutaneous WAT of Ifrd1 knockout mice compared with gene expression in wild-type mice. Pgc1a promoter activity enhanced by the transcription factor specificity protein 1 (Sp1) was markedly repressed by co-introduction of Ifrd1 in brown adipocytes, whereas the repression was markedly prevented by the addition of trichostatin A, a histone deacetylase inhibitor. Moreover, adrenergic stimulation induced complex formation between Ifrd1, Sp1 and mSIN3B, which is a component of the SIN complex containing histone deacetylase, in brown adipocytes. These findings, therefore, suggest that Ifrd1 could be a pivotal negative regulator of sympathetic regulation of thermogenic and mitochondrial gene expression in brown adipocytes by interacting with Sp1 and the mSIN3 complex.


Assuntos
Adipócitos Marrons/metabolismo , Proteínas Imediatamente Precoces/biossíntese , Proteínas de Membrana/biossíntese , Proteínas Repressoras/metabolismo , Fator de Transcrição Sp1/metabolismo , Termogênese/genética , Tecido Adiposo Marrom/metabolismo , Tecido Adiposo Branco/metabolismo , Agonistas Adrenérgicos/administração & dosagem , Animais , Temperatura Baixa , Dioxóis/administração & dosagem , Regulação da Expressão Gênica/efeitos dos fármacos , Proteínas Imediatamente Precoces/genética , Proteínas Imediatamente Precoces/metabolismo , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Camundongos , Complexos Multiproteicos/genética , Complexos Multiproteicos/metabolismo , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/genética , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/metabolismo , Proteínas Repressoras/genética , Fator de Transcrição Sp1/genética
7.
Biochem Biophys Res Commun ; 482(2): 329-334, 2017 Jan 08.
Artigo em Inglês | MEDLINE | ID: mdl-27856249

RESUMO

We previously demonstrated that the transcriptional coactivator/repressor interferon-related developmental regulator 1 (Ifrd1) was expressed in osteoblasts and participated in the regulation of bone homeostasis. However, it remains unclear how Ifrd1 expression itself is regulated in osteoblasts. In the present study, we investigated the upstream regulatory mechanisms of Ifrd1 in osteoblasts during osteoblastogenesis. Ifrd1 protein expression and runt-related transcription factor 2, the master regulator of osteoblastogenesis, were markedly upregulated by bone morphogenetic protein 2 (BMP-2) stimulation in primary osteoblasts. Moreover, BMP-2 stimulation significantly induced Ifrd1 mRNA expression and promoter activity in osteoblasts. LDN193189, an inhibitor of activin-like kinase 2/3, almost completely inhibited the BMP-2-induced increase in Ifrd1 protein expression. There were at least two putative Smad-binding elements in the 5'-flanking region, which was highly conserved between mouse and human Ifrd1 genes. Co-introduction of both Smad4 and Smad1 significantly increased Ifrd1 promoter activity in osteoblasts. In addition, BMP-2 induced the recruitment of Smad1 to the Ifrd1 promoter in osteoblasts. Moreover, BMP-2-dependent osteoblastogenesis was further enhanced in Ifrd1 knocked-down osteoblasts, as determined by the intensity of Alizarin red stain and marker gene expression. These results suggest that BMP-2 directly induces Ifrd1 expression at the transcriptional level in osteoblasts via the Smad pathway, and Ifrd1 negatively regulates BMP-2-dependent osteoblastogenesis.


Assuntos
Proteína Morfogenética Óssea 2/metabolismo , Proteínas Imediatamente Precoces/metabolismo , Proteínas de Membrana/metabolismo , Osteoblastos/citologia , Osteoblastos/fisiologia , Osteogênese/fisiologia , Ativação Transcricional/fisiologia , Animais , Diferenciação Celular/fisiologia , Células Cultivadas , Regulação para Baixo/fisiologia , Regulação da Expressão Gênica no Desenvolvimento/fisiologia , Camundongos
8.
Sci Rep ; 6: 30918, 2016 08 02.
Artigo em Inglês | MEDLINE | ID: mdl-27480204

RESUMO

Bone homeostasis is maintained by the sophisticated coupled actions of bone-resorbing osteoclasts and bone-forming osteoblasts. Here we identify activating transcription factor 3 (ATF3) as a pivotal transcription factor for the regulation of bone resorption and bone remodeling under a pathological condition through modulating the proliferation of osteoclast precursors. The osteoclast precursor-specific deletion of ATF3 in mice led to the prevention of receptor activator of nuclear factor-κB (RANK) ligand (RANKL)-induced bone resorption and bone loss, although neither bone volume nor osteoclastic parameter were markedly altered in these knockout mice under the physiological condition. RANKL-dependent osteoclastogenesis was impaired in vitro in ATF3-deleted bone marrow macrophages (BMM). Mechanistically, the deficiency of ATF3 impaired the RANKL-induced transient increase in cell proliferation of osteoclast precursors in bone marrow in vivo as well as of BMM in vitro. Moreover, ATF3 regulated cyclin D1 mRNA expression though modulating activator protein-1-dependent transcription in the osteoclast precursor, and the introduction of cyclin D1 significantly rescued the impairment of osteoclastogenesis in ATF3-deleted BMM. Therefore, these findings suggest that ATF3 could have a pivotal role in osteoclastogenesis and bone homeostasis though modulating cell proliferation under pathological conditions, thereby providing a target for bone diseases.


Assuntos
Fator 3 Ativador da Transcrição/fisiologia , Remodelação Óssea , Reabsorção Óssea/prevenção & controle , Osteoclastos/citologia , Ligante RANK/efeitos adversos , Animais , Células da Medula Óssea/metabolismo , Reabsorção Óssea/etiologia , Reabsorção Óssea/metabolismo , Reabsorção Óssea/patologia , Diferenciação Celular , Proliferação de Células , Regulação da Expressão Gênica , Macrófagos/citologia , Macrófagos/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Osteoclastos/metabolismo
9.
Mol Cell Biol ; 36(19): 2451-63, 2016 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-27381458

RESUMO

Bone homeostasis is maintained by the synergistic actions of bone-resorbing osteoclasts and bone-forming osteoblasts. Here, we show that the transcriptional coactivator/repressor interferon-related developmental regulator 1 (Ifrd1) is expressed in osteoclast lineages and represents a component of the machinery that regulates bone homeostasis. Ifrd1 expression was transcriptionally regulated in preosteoclasts by receptor activator of nuclear factor κB (NF-κB) ligand (RANKL) through activator protein 1. Global deletion of murine Ifrd1 increased bone formation and decreased bone resorption, leading to a higher bone mass. Deletion of Ifrd1 in osteoclast precursors prevented RANKL-induced bone loss, although no bone loss was observed under normal physiological conditions. RANKL-dependent osteoclastogenesis was impaired in vitro in Ifrd1-deleted bone marrow macrophages (BMMs). Ifrd1 deficiency increased the acetylation of p65 at residues K122 and K123 via the inhibition of histone deacetylase-dependent deacetylation in BMMs. This repressed the NF-κB-dependent transcription of nuclear factor of activated T cells, cytoplasmic 1 (NFATc1), an essential regulator of osteoclastogenesis. These findings suggest that an Ifrd1/NF-κB/NFATc1 axis plays a pivotal role in bone remodeling in vivo and represents a therapeutic target for bone diseases.


Assuntos
Proteínas Imediatamente Precoces/genética , Proteínas Imediatamente Precoces/metabolismo , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , NF-kappa B/metabolismo , Fatores de Transcrição NFATC/metabolismo , Osteoclastos/citologia , Ligante RANK/farmacologia , Animais , Diferenciação Celular/efeitos dos fármacos , Células Cultivadas , Deleção de Genes , Regulação da Expressão Gênica/efeitos dos fármacos , Macrófagos/citologia , Macrófagos/efeitos dos fármacos , Camundongos , Osteoclastos/efeitos dos fármacos , Osteoclastos/metabolismo , Osteogênese/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Fator de Transcrição AP-1/metabolismo , Regulação para Cima
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