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1.
Proc Natl Acad Sci U S A ; 113(5): 1393-8, 2016 Feb 02.
Artigo em Inglês | MEDLINE | ID: mdl-26764384

RESUMO

Oncogenic transcription factors are commonly activated in acute leukemias and subvert normal gene expression networks to reprogram hematopoietic progenitors into preleukemic stem cells, as exemplified by LIM-only 2 (LMO2) in T-cell acute lymphoblastic leukemia (T-ALL). Whether or not these oncoproteins interfere with other DNA-dependent processes is largely unexplored. Here, we show that LMO2 is recruited to DNA replication origins by interaction with three essential replication enzymes: DNA polymerase delta (POLD1), DNA primase (PRIM1), and minichromosome 6 (MCM6). Furthermore, tethering LMO2 to synthetic DNA sequences is sufficient to transform these sequences into origins of replication. We next addressed the importance of LMO2 in erythroid and thymocyte development, two lineages in which cell cycle and differentiation are tightly coordinated. Lowering LMO2 levels in erythroid progenitors delays G1-S progression and arrests erythropoietin-dependent cell growth while favoring terminal differentiation. Conversely, ectopic expression in thymocytes induces DNA replication and drives these cells into cell cycle, causing differentiation blockade. Our results define a novel role for LMO2 in directly promoting DNA synthesis and G1-S progression.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , Replicação do DNA/genética , Células-Tronco Hematopoéticas/metabolismo , Proteínas com Domínio LIM/genética , Animais , Células-Tronco Hematopoéticas/citologia , Camundongos , Origem de Replicação , Fase S
2.
Med Sci (Paris) ; 39(6-7): 530-536, 2023.
Artigo em Francês | MEDLINE | ID: mdl-37387661

RESUMO

Muscle regeneration in response to injury or exercise relies on the ability of muscle stem cells to proliferate and differentiate to repair the damage. In the absence of damage, muscle stem cells are quiescent: they do not proliferate and have a very low metabolism. Recent studies have linked the metabolic state of the adult muscle stem cell to its epigenetic regulation. This article synthesizes the known concepts about histone modifications and metabolic pathways found in quiescent muscle stem cells, as well as the metabolic and epigenetic changes leading to muscle stem cell activation in response to injury. Here, we discuss the heterogeneity in quiescent stem cell metabolism and compare the metabolism of quiescent and activated muscle stem cells, and describe the epigenetic changes related to their activation. We also discuss the involvement of SIRT1, an important effector of muscle stem cells metabolism, together with the effects of aging and caloric restriction.


Title: Des liens entre métabolisme et régulation épigénétique des cellules souches musculaires. Abstract: La régénération musculaire dépend de la capacité des cellules souches musculaires, aussi appelées cellules satellites, à proliférer et à se différencier pour réparer les muscles endommagés. En l'absence de dommage, ces cellules sont quiescentes : elles ne prolifèrent pas et présentent un métabolisme réduit. Des études récentes ont révélé l'existence de liens entre la régulation épigénétique et le métabolisme des cellules souches musculaires. Dans cette synthèse, nous discutons les modifications épigénétiques des histones et les voies métaboliques qui ont été observées dans les cellules souches musculaires quiescentes et qui sont à l'origine de leur activation en réponse à une blessure.


Assuntos
Células-Tronco Adultas , Epigênese Genética , Adulto , Humanos , Envelhecimento , Restrição Calórica , Músculos
3.
Nat Commun ; 13(1): 3961, 2022 07 08.
Artigo em Inglês | MEDLINE | ID: mdl-35803939

RESUMO

Satellite cells are required for the growth, maintenance, and regeneration of skeletal muscle. Quiescent satellite cells possess a primary cilium, a structure that regulates the processing of the GLI family of transcription factors. Here we find that GLI3 processing by the primary cilium plays a critical role for satellite cell function. GLI3 is required to maintain satellite cells in a G0 dormant state. Strikingly, satellite cells lacking GLI3 enter the GAlert state in the absence of injury. Furthermore, GLI3 depletion stimulates expansion of the stem cell pool. As a result, satellite cells lacking GLI3 display rapid cell-cycle entry, increased proliferation and augmented self-renewal, and markedly enhanced regenerative capacity. At the molecular level, we establish that the loss of GLI3 induces mTORC1 signaling activation. Therefore, our results provide a mechanism by which GLI3 controls mTORC1 signaling, consequently regulating muscle stem cell activation and fate.


Assuntos
Células Satélites de Músculo Esquelético , Diferenciação Celular/fisiologia , Proliferação de Células , Alvo Mecanístico do Complexo 1 de Rapamicina , Músculo Esquelético , Células-Tronco , Internalização do Vírus
4.
Nat Commun ; 12(1): 3253, 2021 05 31.
Artigo em Inglês | MEDLINE | ID: mdl-34059674

RESUMO

Muscle stem cell function has been suggested to be regulated by Acetyl-CoA and NAD+ availability, but the mechanisms remain unclear. Here we report the identification of two acetylation sites on PAX7 that positively regulate its transcriptional activity. Lack of PAX7 acetylation reduces DNA binding, specifically to the homeobox motif. The acetyltransferase MYST1 stimulated by Acetyl-CoA, and the deacetylase SIRT2 stimulated by NAD +, are identified as direct regulators of PAX7 acetylation and asymmetric division in muscle stem cells. Abolishing PAX7 acetylation in mice using CRISPR/Cas9 mutagenesis leads to an expansion of the satellite stem cell pool, reduced numbers of asymmetric stem cell divisions, and increased numbers of oxidative IIA myofibers. Gene expression analysis confirms that lack of PAX7 acetylation preferentially affects the expression of target genes regulated by homeodomain binding motifs. Therefore, PAX7 acetylation status regulates muscle stem cell function and differentiation potential to facilitate metabolic adaptation of muscle tissue.


Assuntos
Autorrenovação Celular/genética , Músculo Esquelético/lesões , Fator de Transcrição PAX7/metabolismo , Regeneração/genética , Células Satélites de Músculo Esquelético/fisiologia , Acetilação , Animais , Células COS , Sistemas CRISPR-Cas , Cardiotoxinas/administração & dosagem , Cardiotoxinas/toxicidade , Diferenciação Celular/genética , Chlorocebus aethiops , Modelos Animais de Doenças , Técnicas de Silenciamento de Genes , Histona Acetiltransferases/genética , Histona Acetiltransferases/metabolismo , Humanos , Camundongos , Camundongos Transgênicos , Músculo Esquelético/citologia , Músculo Esquelético/efeitos dos fármacos , Mutagênese , Cultura Primária de Células , Regiões Promotoras Genéticas , Células Sf9 , Sirtuína 2/genética , Sirtuína 2/metabolismo , Spodoptera , Ativação Transcricional
5.
Nat Commun ; 10(1): 4256, 2019 09 18.
Artigo em Inglês | MEDLINE | ID: mdl-31534153

RESUMO

PAX7 is a paired-homeobox transcription factor that specifies the myogenic identity of muscle stem cells and acts as a nodal factor by stimulating proliferation while inhibiting differentiation. We previously found that PAX7 recruits the H3K4 methyltransferases MLL1/2 to epigenetically activate target genes. Here we report that in the absence of Mll1, myoblasts exhibit reduced H3K4me3 at both Pax7 and Myf5 promoters and reduced Pax7 and Myf5 expression. Mll1-deficient myoblasts fail to proliferate but retain their differentiation potential, while deletion of Mll2 had no discernable effect. Re-expression of PAX7 in committed Mll1 cKO myoblasts restored H3K4me3 enrichment at the Myf5 promoter and Myf5 expression. Deletion of Mll1 in satellite cells reduced satellite cell proliferation and self-renewal, and significantly impaired skeletal muscle regeneration. Pax7 expression was unaffected in quiescent satellite cells but was markedly downregulated following satellite cell activation. Therefore, MLL1 is required for PAX7 expression and satellite cell function in vivo. Furthermore, PAX7, but not MLL1, is required for Myf5 transcriptional activation in committed myoblasts.


Assuntos
Histona-Lisina N-Metiltransferase/genética , Proteína de Leucina Linfoide-Mieloide/genética , Mioblastos/metabolismo , Fator Regulador Miogênico 5/metabolismo , Fator de Transcrição PAX7/metabolismo , Células Satélites de Músculo Esquelético/metabolismo , Animais , Diferenciação Celular/genética , Proliferação de Células/genética , Células Cultivadas , Feminino , Masculino , Metilação , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fator Regulador Miogênico 5/genética , Fator de Transcrição PAX7/genética , Regiões Promotoras Genéticas/genética
7.
Cell Stem Cell ; 22(5): 755-768.e6, 2018 05 03.
Artigo em Inglês | MEDLINE | ID: mdl-29681515

RESUMO

Asymmetrically dividing muscle stem cells in skeletal muscle give rise to committed cells, where the myogenic determination factor Myf5 is transcriptionally activated by Pax7. This activation is dependent on Carm1, which methylates Pax7 on multiple arginine residues, to recruit the ASH2L:MLL1/2:WDR5:RBBP5 histone methyltransferase complex to the proximal promoter of Myf5. Here, we found that Carm1 is a specific substrate of p38γ/MAPK12 and that phosphorylation of Carm1 prevents its nuclear translocation. Basal localization of the p38γ/p-Carm1 complex in muscle stem cells occurs via binding to the dystrophin-glycoprotein complex (DGC) through ß1-syntrophin. In dystrophin-deficient muscle stem cells undergoing asymmetric division, p38γ/ß1-syntrophin interactions are abrogated, resulting in enhanced Carm1 phosphorylation. The resulting progenitors exhibit reduced Carm1 binding to Pax7, reduced H3K4-methylation of chromatin, and reduced transcription of Myf5 and other Pax7 target genes. Therefore, our experiments suggest that dysregulation of p38γ/Carm1 results in altered epigenetic gene regulation in Duchenne muscular dystrophy.


Assuntos
Epigênese Genética , Músculo Esquelético/citologia , Fator Regulador Miogênico 5/metabolismo , Fator de Transcrição PAX7/metabolismo , Proteína-Arginina N-Metiltransferases/metabolismo , Células-Tronco/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Animais , Células Cultivadas , Feminino , Masculino , Camundongos , Camundongos Endogâmicos , Músculo Esquelético/metabolismo , Fator Regulador Miogênico 5/genética , Fator de Transcrição PAX7/genética , Proteínas Quinases p38 Ativadas por Mitógeno/genética
9.
Methods Mol Biol ; 1556: 41-50, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28247344

RESUMO

Primary myoblasts can be isolated from mouse muscle cell extracts and cultured in vitro. Muscle cells are usually dissociated manually by mincing with razor blades or scissors in a collagenase/dispase solution. Primary myoblasts are then gradually enriched by pre-plating on collagen-coated plates, based on the observation that mouse fibroblasts attach quickly to collagen-coated plates, and are less adherent. Here, we describe an automated muscle dissociation protocol. We also propose an alternative to pre-plating using magnetic bead separation of primary myoblasts, which improve myoblast purity by minimizing fibroblast contamination.


Assuntos
Separação Imunomagnética/métodos , Mioblastos/citologia , Animais , Biomarcadores , Citometria de Fluxo/métodos , Camundongos , Músculo Esquelético/citologia , Mioblastos/metabolismo , Células Satélites de Músculo Esquelético/citologia , Células Satélites de Músculo Esquelético/metabolismo
10.
Cell Rep ; 18(10): 2320-2330, 2017 03 07.
Artigo em Inglês | MEDLINE | ID: mdl-28273449

RESUMO

Wnt-mediated signals are involved in many important steps in mammalian regeneration. In multiple cell types, the R-spondin (Rspo) family of secreted proteins potently activates the canonical Wnt/ß-catenin pathway. Here, we identify Rspo1 as a mediator of skeletal muscle tissue repair. First, we show that deletion of Rspo1 results in global alteration of muscle regeneration kinetics following acute injury. We find that muscle progenitor cells lacking Rspo1 show delayed differentiation due to reduced activation of Wnt/ß-catenin target genes. Furthermore, muscle cells lacking Rspo1 have a fusion phenotype leading to larger myotubes containing supernumerary nuclei both in vitro and in vivo. The increase in muscle fusion was dependent on downregulation of Wnt/ß-catenin and upregulation of non-canonical Wnt7a/Fzd7/Rac1 signaling. We conclude that reciprocal control of antagonistic Wnt signaling pathways by Rspo1 in muscle stem cell progeny is a key step ensuring normal tissue architecture restoration following acute damage.


Assuntos
Mioblastos/citologia , Mioblastos/metabolismo , Trombospondinas/metabolismo , Via de Sinalização Wnt , Animais , Diferenciação Celular , Fusão Celular , Proliferação de Células , Células Cultivadas , Camundongos Endogâmicos C57BL , Desenvolvimento Muscular , Fator de Transcrição PAX7/metabolismo , Regeneração , Células Satélites de Músculo Esquelético/citologia , Células Satélites de Músculo Esquelético/metabolismo , beta Catenina/metabolismo
11.
Stem Cells Transl Med ; 5(3): 282-90, 2016 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-26798058

RESUMO

Skeletal muscle regeneration is initiated by satellite cells, a population of adult stem cells that reside in the muscle tissue. The ability of satellite cells to self-renew and to differentiate into the muscle lineage is under transcriptional and epigenetic control. Satellite cells are characterized by an open and permissive chromatin state. The transcription factor Pax7 is necessary for satellite cell function. Pax7 is a nodal factor regulating the expression of genes associated with satellite cell growth and proliferation, while preventing differentiation. Pax7 recruits chromatin modifiers to DNA to induce expression of specific target genes involved in myogenic commitment following asymmetric division of muscle stem cells. Emerging evidence suggests that replacement of canonical histones with histone variants is an important regulatory mechanism controlling the ability of satellite cells and myoblasts to differentiate. Differentiation into the muscle lineage is associated with a global gene repression characterized by a decrease in histone acetylation with an increase in repressive histone marks. However, genes important for differentiation are upregulated by the specific action of histone acetyltransferases and other chromatin modifiers, in combination with several transcription factors, including MyoD and Mef2. Treatment with histone deacetylase (HDAC) inhibitors enhances muscle regeneration and is considered as a therapeutic approach in the treatment of muscular dystrophy. This review describes the recent findings on epigenetic regulation in satellite stem cells and committed myoblasts. The potential of epigenetic drugs, such as HDAC inhibitors, as well as their molecular mechanism of action in muscle cells, will be addressed.


Assuntos
Diferenciação Celular/genética , Epigênese Genética , Desenvolvimento Muscular/genética , Músculo Esquelético/crescimento & desenvolvimento , Linhagem da Célula , Proliferação de Células/genética , Regulação da Expressão Gênica no Desenvolvimento , Histona Desacetilases/genética , Músculo Esquelético/metabolismo , Especificidade de Órgãos , Fator de Transcrição PAX7/genética , Regeneração , Células Satélites de Músculo Esquelético/metabolismo
12.
Nat Neurosci ; 19(6): 798-806, 2016 06.
Artigo em Inglês | MEDLINE | ID: mdl-27110918

RESUMO

EGFRvIII-STAT3 signaling is important in glioblastoma pathogenesis. Here, we identified the cytokine receptor OSMR as a direct target gene of the transcription factor STAT3 in mouse astrocytes and human brain tumor stem cells (BTSCs). We found that OSMR functioned as an essential co-receptor for EGFRvIII. OSMR formed a physical complex with EGFRvIII, and depletion of OSMR impaired EGFRvIII-STAT3 signaling. Conversely, pharmacological inhibition of EGFRvIII phosphorylation inhibited the EGFRvIII-OSMR interaction and activation of STAT3. EGFRvIII-OSMR signaling in tumors operated constitutively, whereas EGFR-OSMR signaling in nontumor cells was synergistically activated by the ligands EGF and OSM. Finally, knockdown of OSMR strongly suppressed cell proliferation and tumor growth of mouse glioblastoma cells and human BTSC xenografts in mice, and prolonged the lifespan of these mice. Our findings identify OSMR as a critical regulator of glioblastoma tumor growth that orchestrates a feed-forward signaling mechanism with EGFRvIII and STAT3 to drive tumorigenesis.


Assuntos
Neoplasias Encefálicas/metabolismo , Transformação Celular Neoplásica/metabolismo , Citocinas/metabolismo , Regulação Neoplásica da Expressão Gênica , Glioblastoma/metabolismo , Transdução de Sinais/fisiologia , Animais , Neoplasias Encefálicas/patologia , Linhagem Celular Tumoral , Transformação Celular Neoplásica/genética , Receptores ErbB/genética , Receptores ErbB/metabolismo , Glioblastoma/patologia , Humanos , Masculino , Camundongos Transgênicos , Transplante de Neoplasias/métodos , Fator de Transcrição STAT3/metabolismo
13.
Compr Physiol ; 5(3): 1027-59, 2015 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-26140708

RESUMO

Skeletal muscles are essential for vital functions such as movement, postural support, breathing, and thermogenesis. Muscle tissue is largely composed of long, postmitotic multinucleated fibers. The life-long maintenance of muscle tissue is mediated by satellite cells, lying in close proximity to the muscle fibers. Muscle satellite cells are a heterogeneous population with a small subset of muscle stem cells, termed satellite stem cells. Under homeostatic conditions all satellite cells are poised for activation by stimuli such as physical trauma or growth signals. After activation, satellite stem cells undergo symmetric divisions to expand their number or asymmetric divisions to give rise to cohorts of committed satellite cells and thus progenitors. Myogenic progenitors proliferate, and eventually differentiate through fusion with each other or to damaged fibers to reconstitute fiber integrity and function. In the recent years, research has begun to unravel the intrinsic and extrinsic mechanisms controlling satellite cell behavior. Nonetheless, an understanding of the complex cellular and molecular interactions of satellite cells with their dynamic microenvironment remains a major challenge, especially in pathological conditions. The goal of this review is to comprehensively summarize the current knowledge on satellite cell characteristics, functions, and behavior in muscle regeneration and in pathological conditions.


Assuntos
Diferenciação Celular , Músculo Esquelético/citologia , Regeneração , Células Satélites de Músculo Esquelético/citologia , Animais , Humanos , Músculo Esquelético/metabolismo , Músculo Esquelético/fisiologia , Células Satélites de Músculo Esquelético/metabolismo
14.
J Biol Chem ; 282(46): 33649-33658, 2007 Nov 16.
Artigo em Inglês | MEDLINE | ID: mdl-17878155

RESUMO

Gene expression programs are established by networks of interacting transcription factors. The basic helix-loop-helix factor SCL and the LIM-only protein LMO2 are components of transcription factor complexes that are essential for hematopoiesis. Here we show that LMO2 and SCL are predominant interaction partners in hematopoietic cells and that this interaction occurs through a conserved interface residing in the loop and helix 2 of SCL. This interaction nucleates the assembly of SCL complexes on DNA and is required for target gene induction and for the stimulation of erythroid and megakaryocytic differentiation. We also demonstrate that SCL determines LMO2 protein levels in hematopoietic cells and reveal that interaction with SCL prevents LMO2 degradation by the proteasome. We propose that the SCL-LMO2 interaction couples protein stabilization with higher order protein complex assembly, thus providing a powerful means of modulating the stoichiometry and spatiotemporal activity of SCL complexes. This interaction likely provides a rate-limiting step in the transcriptional control of hematopoiesis and leukemia, and similar mechanisms may operate to control the assembly of diverse protein modules.


Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Proteínas de Ligação a DNA/metabolismo , Metaloproteínas/metabolismo , Proteínas/química , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Adaptadoras de Transdução de Sinal , Sequência de Aminoácidos , Animais , Diferenciação Celular , Humanos , Proteínas com Domínio LIM , Megacariócitos/metabolismo , Camundongos , Modelos Moleculares , Conformação Molecular , Dados de Sequência Molecular , Células NIH 3T3 , Conformação Proteica , Homologia de Sequência de Aminoácidos , Proteína 1 de Leucemia Linfocítica Aguda de Células T
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