RESUMO
Schwann cells are the nerve ensheathing cells of the peripheral nervous system. Absence, loss and malfunction of Schwann cells or their myelin sheaths lead to peripheral neuropathies such as Charcot-Marie-Tooth disease in humans. During Schwann cell development and myelination chromatin is dramatically modified. However, impact and functional relevance of these modifications are poorly understood. Here, we analyzed histone H2B monoubiquitination as one such chromatin modification by conditionally deleting the Rnf40 subunit of the responsible E3 ligase in mice. Rnf40-deficient Schwann cells were arrested immediately before myelination or generated abnormally thin, unstable myelin, resulting in a peripheral neuropathy characterized by hypomyelination and progressive axonal degeneration. By combining sequencing techniques with functional studies we show that H2B monoubiquitination does not influence global gene expression patterns, but instead ensures selective high expression of myelin and lipid biosynthesis genes and proper repression of immaturity genes. This requires the specific recruitment of the Rnf40-containing E3 ligase by Egr2, the central transcriptional regulator of peripheral myelination, to its target genes. Our study identifies histone ubiquitination as essential for Schwann cell myelination and unravels new disease-relevant links between chromatin modifications and transcription factors in the underlying regulatory network.
Assuntos
Proteína 2 de Resposta de Crescimento Precoce/fisiologia , Neuropatia Hereditária Motora e Sensorial/metabolismo , Histonas/metabolismo , Sistema Nervoso Periférico/metabolismo , Células de Schwann/metabolismo , Animais , Linhagem Celular Tumoral , Células HEK293 , Humanos , Camundongos , Camundongos Transgênicos , Sistema Nervoso Periférico/patologia , Ratos , Células de Schwann/patologia , Ubiquitina-Proteína Ligases/genética , UbiquitinaçãoRESUMO
γδ T cells with distinct properties develop in the embryonic and adult thymus and have been identified as critical players in a broad range of infections, antitumor surveillance, autoimmune diseases, and tissue homeostasis. Despite their potential value for immunotherapy, differentiation of γδ T cells in the thymus is incompletely understood. Here, we establish a high-resolution map of γδ T-cell differentiation from the fetal and adult thymus using single-cell RNA sequencing. We reveal novel sub-types of immature and mature γδ T cells and identify an unpolarized thymic population which is expanded in the blood and lymph nodes. Our detailed comparative analysis reveals remarkable similarities between the gene networks active during fetal and adult γδ T-cell differentiation. By performing a combined single-cell analysis of Sox13, Maf, and Rorc knockout mice, we demonstrate sequential activation of these factors during IL-17-producing γδ T-cell (γδT17) differentiation. These findings substantially expand our understanding of γδ T-cell ontogeny in fetal and adult life. Our experimental and computational strategy provides a blueprint for comparing immune cell differentiation across developmental stages.
Assuntos
Diferenciação Celular/imunologia , Feto/imunologia , Receptores de Antígenos de Linfócitos T gama-delta/imunologia , Linfócitos T/imunologia , Animais , Autoantígenos/genética , Autoantígenos/imunologia , Diferenciação Celular/genética , Camundongos , Camundongos Knockout , Membro 3 do Grupo F da Subfamília 1 de Receptores Nucleares/genética , Membro 3 do Grupo F da Subfamília 1 de Receptores Nucleares/imunologia , Proteínas Proto-Oncogênicas c-maf/genética , Proteínas Proto-Oncogênicas c-maf/imunologia , Receptores de Antígenos de Linfócitos T gama-delta/genética , Linfócitos T/citologiaRESUMO
Development of oligodendrocytes and myelin formation in the vertebrate central nervous system is under control of several basic helix-loop-helix transcription factors such as Olig2, Ascl1, Hes5 and the Id proteins. The class I basic helix-loop-helix proteins Tcf3, Tcf4 and Tcf12 represent potential heterodimerization partners and functional modulators for all, but have not been investigated in oligodendrocytes so far. Using mouse mutants, organotypic slice and primary cell cultures we here show that Tcf4 is required in a cell-autonomous manner for proper terminal differentiation and myelination in vivo and ex vivo. Partial compensation is provided by the paralogous Tcf3, but not Tcf12. On the mechanistic level Tcf4 was identified as the preferred heterodimerization partner of the central regulator of oligodendrocyte development Olig2. Both genetic studies in the mouse as well as functional studies on enhancer regions of myelin genes confirmed the relevance of this physical interaction for oligodendrocyte differentiation. Considering that alterations in TCF4 are associated with syndromic and non-syndromic forms of intellectual disability, schizophrenia and autism in humans, our findings point to the possibility of an oligodendroglial contribution to these disorders.
Assuntos
Fator de Transcrição 2 de Oligodendrócitos/genética , Oligodendroglia/citologia , Fator de Transcrição 4/genética , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Diferenciação Celular , Linhagem Celular Tumoral , Células Cultivadas , Dimerização , Feminino , Deleção de Genes , Células HEK293 , Humanos , Masculino , Camundongos , Bainha de Mielina/fisiologia , Oligodendroglia/metabolismo , Ratos WistarRESUMO
The high-mobility-group (HMG)-domain protein Sox9 is one of few transcription factors implicated in gliogenesis in the vertebrate central nervous system. To further study the role of Sox9 in early spinal cord development, we generated a mouse that allows expression of Sox9 in a temporally and spatially controlled manner. Using this mouse, we show that premature Sox9 expression in neural precursor cells disrupted the neuroepithelium of the ventricular zone. Sox9 also compromised development and survival of neuronal precursors and neurons. Additionally, we observed in these mice substantial increases in oligodendroglial and astroglial cells. Reversing the normal order of appearance of essential transcriptional regulators during oligodendrogenesis, Sox10 preceded Olig2. Our study reinforces the notion that Sox9 has a strong gliogenic activity. It also argues that Sox9 expression has to be tightly controlled to prevent negative effects on early spinal cord structure and neuronal development.
Assuntos
Astrócitos/metabolismo , Oligodendroglia/metabolismo , Fatores de Transcrição SOX9/metabolismo , Medula Espinal/metabolismo , Animais , Linhagem Celular Tumoral , Camundongos , Camundongos Transgênicos , Fator de Transcrição 2 de Oligodendrócitos/genética , Fator de Transcrição 2 de Oligodendrócitos/metabolismo , Fatores de Transcrição SOX9/genética , Medula Espinal/crescimento & desenvolvimentoRESUMO
Mechanisms of glomerular crescent formation and podocyte repair processes are still unclear. Therefore, we investigated the expression of the transcription factor Sox9 as a potential marker of a subpopulation of parietal epithelial cells (PECs) with potential regenerative properties. Glomerular Sox9 expression was characterized in detail in a rat anti-glomerular basement membrane (GBM) nephritis model using immunofluorescence and confocal laser scanning microscopy. In healthy kidneys Sox9 is expressed in a subpopulation of PECs restricted to approximately 20% to 50% of PEC nuclei and was highly conserved in all investigated species. During rat anti-GBM nephritis the number of glomerular Sox9+ cells increased and was associated with proliferation activity. In nephritic glomeruli Sox9 expression was not restricted to Bowman's capsule lining but was also found on cells of the glomerular tuft. Nearly all Sox9+ cells also expressed the PEC marker Pax8, whereas endothelial cells, mesangial cells, macrophages, and T lymphocytes lacked Sox9 expression. At the margins of crescents Sox9+/Pax8+ cells additionally expressed podocyte markers. In contrast, in sclerotic lesions a minority of Sox9+/Pax8+ cells expressed the myofibroblast marker α-smooth muscle actin. In glomerular Sox9+ cells Jagged 1 was up-regulated. During anti-GBM nephritis Sox9+ PECs proliferate and migrate onto the glomerular tuft. Future studies are needed to confirm the origin of Sox9+ cells from PECs and differentiation in both podocytes and/or myofibroblasts.
Assuntos
Doença Antimembrana Basal Glomerular/patologia , Células Epiteliais/patologia , Membrana Basal Glomerular/patologia , Nefrite/patologia , Podócitos/patologia , Fatores de Transcrição SOX9/metabolismo , Animais , Doença Antimembrana Basal Glomerular/metabolismo , Diferenciação Celular , Células Cultivadas , Células Epiteliais/metabolismo , Membrana Basal Glomerular/metabolismo , Proteína Jagged-1 , Masculino , Nefrite/metabolismo , Podócitos/metabolismo , Ratos , Ratos Endogâmicos WKYRESUMO
We and others previously showed that in mouse embryos lacking the transcription factor Sox10, olfactory ensheathing cell (OEC) differentiation is disrupted, resulting in defective olfactory axon targeting and fewer gonadotropin-releasing hormone (GnRH) neurons entering the embryonic forebrain. The underlying mechanisms are unclear. Here, we report that OECs in the olfactory nerve layer express Frzb-encoding a secreted Wnt inhibitor with roles in axon targeting and basement membrane breakdown-from embryonic day (E)12.5, when GnRH neurons first enter the forebrain, until E16.5, the latest stage examined. The highest levels of Frzb expression are seen in OECs in the inner olfactory nerve layer, abutting the embryonic olfactory bulb. We find that Sox10 is required for Frzb expression in OECs, suggesting that loss of Frzb could explain the olfactory axon targeting and/or GnRH neuron migration defects seen in Sox10-null mice. At E16.5, Frzb-null embryos show significant reductions in both the volume of the olfactory nerve layer expressing the maturation marker Omp and the number of Omp-positive olfactory receptor neurons in the olfactory epithelium. As Omp upregulation correlates with synapse formation, this suggests that Frzb deletion indeed disrupts olfactory axon targeting. In contrast, GnRH neuron entry into the forebrain is not significantly affected. Hence, loss of Frzb may contribute to the olfactory axon targeting phenotype, but not the GnRH neuron phenotype, of Sox10-null mice. Overall, our results suggest that Frzb secreted from OECs in the olfactory nerve layer is important for olfactory axon targeting.
Assuntos
Axônios/metabolismo , Regulação da Expressão Gênica no Desenvolvimento/genética , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Neuroglia/metabolismo , Bulbo Olfatório , Neurônios Receptores Olfatórios/patologia , Animais , Antígenos de Neoplasias/metabolismo , Embrião de Mamíferos , Hormônio Liberador de Gonadotropina/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/genética , Camundongos , Camundongos Transgênicos , Neuropeptídeo Y/metabolismo , Bulbo Olfatório/citologia , Bulbo Olfatório/embriologia , Bulbo Olfatório/metabolismo , Proteína de Marcador Olfatório/genética , Proteína de Marcador Olfatório/metabolismo , Mucosa Olfatória/citologia , Mucosa Olfatória/metabolismo , Fatores de Transcrição SOXE/genética , Fatores de Transcrição SOXE/metabolismo , Tubulina (Proteína)/metabolismoRESUMO
Understanding transcription factor (TF) regulation of limbal epithelial stem/progenitor cells (LEPCs) may aid in using non-ocular cells to regenerate the corneal surface. This study aimed to identify and characterize TF genes expressed specifically in LEPCs isolated from human donor eyes by laser capture microdissection. Using a profiling approach, preferential limbal expression was found for SoxE and SoxF genes, particularly for Sox9, which showed predominantly cytoplasmic localization in basal LEPCs and nuclear localization in suprabasal and corneal epithelial cells, indicating nucleocytoplasmic translocation and activation during LEPC proliferation and differentiation. Increased nuclear localization of Sox9 was also observed in activated LEPCs following clonal expansion and corneal epithelial wound healing. Knockdown of SOX9 expression in cultured LEPCs by RNAi led to reduced expression of progenitor cell markers, e.g. keratin 15, and increased expression of differentiation markers, e.g. keratin 3. Furthermore, SOX9 silencing significantly suppressed the proliferative capacity of LEPCs and reduced levels of glycogen synthase kinase 3 beta (GSK-3ß), a negative regulator of Wnt/ß-catenin signaling. Sox9 expression, in turn, was significantly suppressed by treatment of LEPCs with exogenous GSK-3ß inhibitors and enhanced by small molecule inhibitors of Wnt signaling. Our results suggest that Sox9 and Wnt/ß-catenin signaling cooperate in mutually repressive interactions to achieve a balance between quiescence, proliferation and differentiation of LEPCs in the limbal niche. Future molecular dissection of Sox9-Wnt interaction and mechanisms of nucleocytoplasmic shuttling of Sox9 may aid in improving the regenerative potential of LEPCs and the reprogramming of non-ocular cells for corneal surface regeneration.
Assuntos
Proliferação de Células , Células Epiteliais/citologia , Limbo da Córnea/citologia , Fatores de Transcrição SOX9/metabolismo , Nicho de Células-Tronco/fisiologia , Células-Tronco/citologia , Fatores de Transcrição/metabolismo , Diferenciação Celular , Células Cultivadas , Células Epiteliais/metabolismo , Humanos , Microdissecção e Captura a Laser , Limbo da Córnea/metabolismo , Fatores de Transcrição SOX9/genética , Células-Tronco/metabolismo , Fatores de Transcrição/genéticaRESUMO
The high-mobility-group domain containing SoxC transcription factors Sox4 and Sox11 are expressed and required in the vertebrate central nervous system in neuronal precursors and neuroblasts. To identify genes that are widely regulated by SoxC proteins during vertebrate neurogenesis we generated expression profiles from developing mouse brain and chicken neural tube with reduced SoxC expression and found the transcription factor prospero homeobox protein 1 (Prox1) strongly down-regulated under both conditions. This led us to hypothesize that Prox1 expression depends on SoxC proteins in the developing central nervous system of mouse and chicken. By combining luciferase reporter assays and over-expression in the chicken neural tube with in vivo and in vitro binding studies, we identify the Prox1 gene promoter and two upstream enhancers at -44 kb and -40 kb relative to the transcription start as regulatory regions that are bound and activated by SoxC proteins. This argues that Prox1 is a direct target gene of SoxC proteins during neurogenesis. Electroporations in the chicken neural tube furthermore show that Prox1 activates a subset of SoxC target genes, whereas it has no effects on others. We propose that the transcriptional control of Prox1 by SoxC proteins may ensure coupling of two types of transcription factors that are both required during early neurogenesis, but have at least in part distinct functions. Open Data: Materials are available on https://cos.io/our-services/open-science-badges/ https://osf.io/93n6m/.
Assuntos
Regulação da Expressão Gênica no Desenvolvimento/fisiologia , Proteínas de Homeodomínio/metabolismo , Células-Tronco Neurais/fisiologia , Neurogênese/fisiologia , Prosencéfalo/citologia , Fatores de Transcrição SOXC/metabolismo , Proteínas Supressoras de Tumor/metabolismo , Animais , Embrião de Galinha , Imunoprecipitação da Cromatina , Biologia Computacional , Ensaio de Desvio de Mobilidade Eletroforética , Eletroporação , Embrião de Mamíferos , Ontologia Genética , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Proteínas de Homeodomínio/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Tubo Neural/citologia , Tubo Neural/metabolismo , Fatores do Domínio POU/genética , Fatores do Domínio POU/metabolismo , Prosencéfalo/embriologia , Prosencéfalo/crescimento & desenvolvimento , Prosencéfalo/metabolismo , Fatores de Transcrição SOXC/genética , Tubulina (Proteína)/metabolismo , Proteínas Supressoras de Tumor/genéticaRESUMO
INTRODUCTION: Congenital hypomyelinating neuropathy (CHN) is a rare congenital neuropathy that presents in the neonatal period and has been linked previously to mutations in several genes associated with myelination. A recent study has linked 4 homozygous frameshift mutations in the contactin-associated protein 1 (CNTNAP1) gene with this condition. METHODS: We report a neonate with CHN who was found to have absent sensory nerve and compound muscle action potentials and hypomyelination on nerve biopsy. RESULTS: On whole exome sequencing, we identified a novel CNTNAP1 homozygous missense mutation (p.Arg388Pro) in the proband, and both parents were carriers. Molecular modeling suggests that this variant disrupts a ß-strand to cause an unstable structure and likely significant changes in protein function. CONCLUSIONS: This report links a missense CNTNAP1 variant to the disease phenotype previously associated only with frameshift mutations. Muscle Nerve 55: 761-765, 2017.
Assuntos
Moléculas de Adesão Celular Neuronais/genética , Doença de Charcot-Marie-Tooth/genética , Mutação de Sentido Incorreto , Potenciais de Ação/fisiologia , Doença de Charcot-Marie-Tooth/fisiopatologia , Eletromiografia , Evolução Fatal , Humanos , Recém-Nascido , Masculino , Neurônios Motores/fisiologia , Condução Nervosa/fisiologiaRESUMO
Differentiation of oligodendrocytes and myelin production in the vertebrate central nervous system require highly concerted changes in gene expression. The transcription factors Sox10 and Myrf are both central to this process and jointly regulate expression of myelin genes. Here we show that Sox10 and Myrf also cooperate in the activation of the gene coding for the dual specificity protein phosphatase Dusp15 (also known as VHY) during this process. Activation is mediated by the Dusp15 promoter, which is also sufficient to drive oligodendroglial gene expression in vivo. It contains both a functional Sox10 and a functional Myrf binding site. Whereas Sox10 binds as a monomer, Myrf binds as a trimer. Available data furthermore indicate that cooperative activation is not a function of facilitated binding, but occurs at a later step of the activation process. shRNA-mediated knockdown of Dusp15 reduced expression of early and late differentiation markers in CG4 and primary oligodendroglial cells, whereas Dusp15 overexpression increased it transiently. This argues that Dusp15 is not only a joint target of Sox10 and Myrf in oligodendrocytes but may also mediate some of their effects during oligodendrocyte differentiation and myelin formation. GLIA 2016;64:2120-2132.
Assuntos
Fosfatases de Especificidade Dupla/metabolismo , Bainha de Mielina/metabolismo , Oligodendroglia/metabolismo , Fatores de Transcrição SOXE/metabolismo , Fatores de Transcrição/metabolismo , Fatores Etários , Animais , Animais Recém-Nascidos , Encéfalo/citologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/fisiologia , Células Cultivadas , Fosfatases de Especificidade Dupla/genética , Regulação da Expressão Gênica no Desenvolvimento/genética , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Humanos , Camundongos , Proteína Básica da Mielina/genética , Proteína Básica da Mielina/metabolismo , Regiões Promotoras Genéticas/genética , RNA Mensageiro/metabolismo , Ratos , Fatores de Transcrição SOXE/genética , Fatores de Transcrição/genética , TransfecçãoRESUMO
Melanoma is the most fatal skin cancer, but the etiology of this devastating disease is still poorly understood. Recently, the transcription factor Sox10 has been shown to promote both melanoma initiation and progression. Reducing SOX10 expression levels in human melanoma cells and in a genetic melanoma mouse model, efficiently abolishes tumorigenesis by inducing cell cycle exit and apoptosis. Here, we show that this anti-tumorigenic effect functionally involves SOX9, a factor related to SOX10 and upregulated in melanoma cells upon loss of SOX10. Unlike SOX10, SOX9 is not required for normal melanocyte stem cell function, the formation of hyperplastic lesions, and melanoma initiation. To the contrary, SOX9 overexpression results in cell cycle arrest, apoptosis, and a gene expression profile shared by melanoma cells with reduced SOX10 expression. Moreover, SOX9 binds to the SOX10 promoter and induces downregulation of SOX10 expression, revealing a feedback loop reinforcing the SOX10 low/SOX9 high ant,m/ii-tumorigenic program. Finally, SOX9 is required in vitro and in vivo for the anti-tumorigenic effect achieved by reducing SOX10 expression. Thus, SOX10 and SOX9 are functionally antagonistic regulators of melanoma development.
Assuntos
Carcinogênese/genética , Melanoma/genética , Fatores de Transcrição SOX9/genética , Fatores de Transcrição SOXE/genética , Animais , Apoptose/genética , Linhagem Celular Tumoral , Regulação Neoplásica da Expressão Gênica , Folículo Piloso , Humanos , Melanócitos/patologia , Melanoma/patologia , Camundongos , RNA Interferente Pequeno , Fatores de Transcrição SOX9/biossíntese , Fatores de Transcrição SOXE/biossínteseRESUMO
BACKGROUND: SOX9 mutations cause the skeletal malformation syndrome campomelic dysplasia in combination with XY sex reversal. Studies in mice indicate that SOX9 acts as a testis-inducing transcription factor downstream of SRY, triggering Sertoli cell and testis differentiation. An SRY-dependent testis-specific enhancer for Sox9 has been identified only in mice. A previous study has implicated copy number variations (CNVs) of a 78 kb region 517-595 kb upstream of SOX9 in the aetiology of both 46,XY and 46,XX disorders of sex development (DSD). We wanted to better define this region for both disorders. RESULTS: By CNV analysis, we identified SOX9 upstream duplications in three cases of SRY-negative 46,XX DSD, which together with previously reported duplications define a 68 kb region, 516-584 kb upstream of SOX9, designated XXSR (XX sex reversal region). More importantly, we identified heterozygous deletions in four families with SRY-positive 46,XY DSD without skeletal phenotype, which define a 32.5 kb interval 607.1-639.6 kb upstream of SOX9, designated XY sex reversal region (XYSR). To localise the suspected testis-specific enhancer, XYSR subfragments were tested in cell transfection and transgenic experiments. While transgenic experiments remained inconclusive, a 1.9 kb SRY-responsive subfragment drove expression specifically in Sertoli-like cells. CONCLUSIONS: Our results indicate that isolated 46,XY and 46,XX DSD can be assigned to two separate regulatory regions, XYSR and XXSR, far upstream of SOX9. The 1.9 kb SRY-responsive subfragment from the XYSR might constitute the core of the Sertoli-cell enhancer of human SOX9, representing the so far missing link in the genetic cascade of male sex determination.
Assuntos
Variações do Número de Cópias de DNA , Transtornos do Desenvolvimento Sexual/genética , Sequências Reguladoras de Ácido Nucleico , Fatores de Transcrição SOX9/genética , Animais , Linhagem Celular , Estudos de Coortes , Feminino , Humanos , Masculino , Camundongos , LinhagemRESUMO
Sox proteins are mechanistically versatile regulators with established relevance to different developmental processes and crucial impact on chromatin structure, DNA conformation, and transcriptional initiation. Here, we show that Sox2 and Sox10, two Sox proteins important for Schwann cell development, also have the capability to activate transcriptional elongation in a Schwann cell line by recruiting the positive transcription elongation factor b. Recruitment is mediated by physical interaction between the carboxyterminal transactivation domains of the two Sox proteins and the Cyclin T1 subunit of positive transcription elongation factor b, with interaction interfaces for the two Sox proteins being mapped to adjacent regions of the central part of Cyclin T1. Supporting the relevance of this interaction to Schwann cell development, transcription of myelin genes appears regulated at the level of elongation. Our results thus add a new facet to the activity of Sox proteins and expand the functional repertoire of this important group of developmental regulators. Sox transcription factors are important regulators of nervous system development. While they are known to regulate transcription by recruiting and stabilizing the RNA polymerase II preinitiation complex directly or with help of the Mediator complex, this study provides evidence that Sox10 and Sox2 additionally influence transcription in glial cells at the elongation stage by recruiting P-TEFb. Cdk9, cyclin-dependent kinase 9; P-TEFb, positive transcription elongation factor b; Pol II, RNA polymerase II; Sox, Sox2 or Sox10 protein.
Assuntos
Fator B de Elongação Transcricional Positiva/metabolismo , Fatores de Transcrição SOXB1/metabolismo , Fatores de Transcrição SOXE/metabolismo , Células de Schwann/metabolismo , Transcrição Gênica/fisiologia , Animais , Linhagem Celular Tumoral , Células HEK293 , Humanos , Camundongos , Ligação Proteica/fisiologia , RatosRESUMO
Mice homozygous for the gray tremor (gt) mutation have a pleiotropic phenotype that includes pigmentation defects, megacolon, whole body tremors, sporadic seizures, hypo- and dys-myelination of the central nervous system (CNS) and peripheral nervous system, vacuolation of the CNS, and early death. Vacuolation similar to that caused by prions was originally reported to be transmissible, but subsequent studies showed the inherited disease was not infectious. The gt mutation mapped to distal mouse chromosome 15, to the same region as Sox10, which encodes a transcription factor with essential roles in neural crest survival and differentiation. As dominant mutations in mouse or human SOX10 cause white spotting and intestinal aganglionosis, we screened the Sox10 coding region for mutations in gt/gt DNA. An adenosine to guanine transversion was identified in exon 2 that changes a highly conserved glutamic acid residue in the SOX10 DNA binding domain to glycine. This mutant allele was not seen in wildtype mice, including the related GT/Le strain, and failed to complement a Sox10 null allele. Gene expression analysis revealed significant down-regulation of genes involved in myelin lipid biosynthesis pathways in gt/gt brains. Knockout mice for some of these genes develop CNS vacuolation and/or myelination defects, suggesting that their down-regulation may contribute to these phenotypes in gt mutants and could underlie the neurological phenotypes associated with peripheral demyelinating neuropathy-central dysmyelinating leukodystrophy-Waardenburg syndrome-Hirschsprung disease, caused by mutations in human SOX10.
Assuntos
Regulação da Expressão Gênica/genética , Doenças Neurodegenerativas/genética , Doenças Neurodegenerativas/fisiopatologia , Fatores de Transcrição SOXE/metabolismo , Animais , Vias Biossintéticas/genética , Análise Mutacional de DNA , Primers do DNA/genética , Galactosídeos , Perfilação da Expressão Gênica , Humanos , Indóis , Camundongos , Camundongos Knockout , Camundongos Mutantes , Repetições de Microssatélites/genética , Mutação de Sentido Incorreto/genética , Bainha de Mielina/metabolismo , Fatores de Transcrição SOXE/genéticaRESUMO
Lineage progression and diversification is regulated by the coordinated action of unique sets of transcription factors. Oligodendrocytes (OL) and astrocytes (AS) comprise the glial sub-lineages in the CNS, and the manner in which their associated regulatory factors orchestrate lineage diversification during development and disease remains an open question. Sox10 and NFIA are key transcriptional regulators of gliogenesis associated with OL and AS. We found that NFIA inhibited Sox10 induction of OL differentiation through direct association and antagonism of its function. Conversely, we found that Sox10 antagonized NFIA function and suppressed AS differentiation in mouse and chick systems. Using this developmental paradigm as a model for glioma, we found that this relationship similarly regulated the generation of glioma subtypes. Our results describe the antagonistic relationship between Sox10 and NFIA that regulates the balance of OL and AS fate during development and demonstrate for the first time, to the best of our knowledge, that the transcriptional processes governing glial sub-lineage diversification oversee the generation of glioma subtypes.
Assuntos
Glioma/classificação , Glioma/metabolismo , Neuroglia/metabolismo , Fatores de Transcrição SOXE/metabolismo , Animais , Animais Recém-Nascidos , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Diferenciação Celular/efeitos dos fármacos , Células Cultivadas , Córtex Cerebral/citologia , Embrião de Galinha , Imunoprecipitação da Cromatina , Eletroporação , Embrião de Mamíferos , Glioma/genética , Proteínas de Fluorescência Verde/genética , Proteínas de Homeodomínio/metabolismo , Camundongos , Proteína Básica da Mielina/genética , Proteína Básica da Mielina/metabolismo , Fatores de Transcrição NFI/genética , Fatores de Transcrição NFI/metabolismo , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Neuroglia/efeitos dos fármacos , Fator de Transcrição 2 de Oligodendrócitos , Fatores de Transcrição SOXE/genética , TransfecçãoRESUMO
Sensory nerves of the brainstem are mostly composed of placode-derived neurons, neural crest-derived neurons and neural crest-derived Schwann cells. This mixed origin of cells has made it difficult to dissect interdependence for fiber guidance. Inner ear-derived neurons are known to connect to the brain after delayed loss of Schwann cells in ErbB2 mutants. However, the ErbB2 mutant related alterations in the ear and the brain compound interpretation of the data. We present here a new model to evaluate exclusively the effect of Schwann cell loss on inner ear innervation. Conditional deletion of the neural crest specific transcription factor, Sox10, using the rhombic lip/neural crest specific Wnt1-cre driver spares Sox10 expression in the ear. We confirm that neural crest-derived cells provide a stop signal for migrating spiral ganglion neurons. In the absence of Schwann cells, spiral ganglion neurons migrate into the center of the cochlea and even out of the ear toward the brain. Spiral ganglion neuron afferent processes reach the organ of Corti, but many afferent fibers bypass the organ of Corti to enter the lateral wall of the cochlea. In contrast to this peripheral disorganization, the central projection to cochlear nuclei is normal. Compared to ErbB2 mutants, conditional Sox10 mutants have limited cell death in spiral ganglion neurons, indicating that the absence of Schwann cells alone contributes little to the embryonic survival of neurons. These data suggest that neural crest-derived cells are dispensable for all central and some peripheral targeting of inner ear neurons. However, Schwann cells provide a stop signal for migratory spiral ganglion neurons and facilitate proper targeting of the organ of Corti by spiral ganglion afferents.
Assuntos
Movimento Celular , Orelha Interna/citologia , Deleção de Genes , Marcação de Genes , Neurônios/citologia , Fatores de Transcrição SOXE/metabolismo , Proteína Wnt1/metabolismo , Animais , Apoptose , Feminino , Integrases/metabolismo , Metaloproteinases da Matriz Associadas à Membrana/metabolismo , Camundongos Knockout , Modelos Biológicos , Mutação/genética , Fatores de Crescimento Neural/metabolismo , Órgão Espiral/citologia , Recombinação Genética/genética , Reprodutibilidade dos Testes , Células de Schwann/citologia , Células de Schwann/metabolismo , Gânglio Espiral da Cóclea/citologia , Vestíbulo do Labirinto/citologia , Vestíbulo do Labirinto/metabolismoRESUMO
Loss of the Merlin tumour suppressor causes abnormal de-differentiation and proliferation of Schwann cells and formation of schwannoma tumours in patients with neurofibromatosis type 2. Within the mature peripheral nerve the normal development, differentiation and maintenance of myelinating and non-myelinating Schwann cells is regulated by a network of transcription factors that include SOX10, OCT6 (now known as POU3F1), NFATC4 and KROX20 (also known as Egr2). We have examined for the first time how their regulation of Schwann cell development is disrupted in primary human schwannoma cells. We find that induction of both KROX20 and OCT6 is impaired, whereas enforced expression of KROX20 drives both myelin gene expression and cell cycle arrest in Merlin-null cells. Importantly, we show that human schwannoma cells have reduced expression of SOX10 protein and messenger RNA. Analysis of mouse SOX10-null Schwann cells shows they display many of the characteristics of human schwannoma cells, including increased expression of platelet derived growth factor receptor beta (PDGFRB) messenger RNA and protein, enhanced proliferation, increased focal adhesions and schwannoma-like morphology. Correspondingly, reintroduction of SOX10 into human Merlin-null cells restores the ability of these cells to induce KROX20 and myelin protein zero (MPZ), localizes NFATC4 to the nucleus, reduces cell proliferation and suppresses PDGFRB expression. Thus, we propose that loss of the SOX10 protein, which is vital for normal Schwann cell development, is also key to the pathology of Merlin-null schwannoma tumours.
Assuntos
Técnicas de Silenciamento de Genes , Neurilemoma/genética , Neurofibromatose 2/genética , Neurofibromina 2/deficiência , Fenótipo , Fatores de Transcrição SOXE/deficiência , Animais , Células Cultivadas , Humanos , Camundongos , Camundongos Transgênicos , Neurilemoma/metabolismo , Neurilemoma/patologia , Neurofibromatose 2/metabolismo , Neurofibromatose 2/patologia , Neurofibromina 2/genética , Fatores de Transcrição SOXE/fisiologiaRESUMO
BACKGROUND: Because of their beneficial cardiovascular effects, several studies have recently advocated starting statins at a young age for primary prevention. However, some reports suggest that statin therapy may be associated with an increased incidence of musculoskeletal and neoplastic diseases. This study was conducted to investigate the incidence of various musculoskeletal and neoplastic diseases in statin users and nonusers. METHODS: A retrospective cohort study of patients in the San Antonio Military Multi-Service Market during the period from October 1, 2003, to March 5, 2010, was conducted. The International Classification of Diseases, 9th edition, diagnosis codes between 2 cohort groups-statin users and nonusers-were compared. Statin users were those patients with at least one 3-month prescription for a statin in the fiscal year 2004. Nonusers were those patients who received a prescription-but not a statin-during the period of the study. Both groups were assessed for the development of musculoskeletal and neoplastic diseases in the following 4-year period (October 1, 2004, to September 30, 2009). RESULTS: A total of 92,360 patients were identified: 12,980 statin users and 45,997 nonusers. After adjusting for age, sex and Charlson comorbidity index, statin users had significantly higher rates of osteoarthritis and arthropathy (odds ratio: 1.26; 95% confidence interval: 1.19-1.33), and dorsopathies, rheumatism and chondropathies (odds ratio: 1.20; 95% confidence interval: 1.12-1.27). CONCLUSIONS: In this retrospective analysis, statin use was associated with an increased incidence of musculoskeletal diseases, including arthropathy. Further studies are needed to provide physicians and their patients with adequate information regarding statin therapy, particularly if recommended for primary prevention in younger populations.
Assuntos
Inibidores de Hidroximetilglutaril-CoA Redutases/efeitos adversos , Doenças Musculoesqueléticas/epidemiologia , Neoplasias/epidemiologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Estudos de Coortes , Feminino , Humanos , Incidência , Masculino , Pessoa de Meia-Idade , Doenças Musculoesqueléticas/induzido quimicamente , Doenças Musculoesqueléticas/diagnóstico , Neoplasias/induzido quimicamente , Neoplasias/diagnóstico , Estudos Retrospectivos , VeteranosRESUMO
Giant congenital naevi are pigmented childhood lesions that frequently lead to melanoma, the most aggressive skin cancer. The mechanisms underlying this malignancy are largely unknown, and there are no effective therapies. Here we describe a mouse model for giant congenital naevi and show that naevi and melanoma prominently express Sox10, a transcription factor crucial for the formation of melanocytes from the neural crest. Strikingly, Sox10 haploinsufficiency counteracts Nras(Q61K)-driven congenital naevus and melanoma formation without affecting the physiological functions of neural crest derivatives in the skin. Moreover, Sox10 is also crucial for the maintenance of neoplastic cells in vivo. In human patients, virtually all congenital naevi and melanomas are SOX10 positive. Furthermore, SOX10 silencing in human melanoma cells suppresses neural crest stem cell properties, counteracts proliferation and cell survival, and completely abolishes in vivo tumour formation. Thus, SOX10 represents a promising target for the treatment of congenital naevi and melanoma in human patients.
Assuntos
Melanoma/fisiopatologia , Nevo/patologia , Fatores de Transcrição SOXE/metabolismo , Animais , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Western Blotting , Linhagem Celular , Linhagem Celular Tumoral , Criança , Modelos Animais de Doenças , Feminino , Regulação Neoplásica da Expressão Gênica , Haploinsuficiência , Humanos , Imuno-Histoquímica , Lactente , Masculino , Melanoma/genética , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C3H , Camundongos Endogâmicos C57BL , Análise em Microsséries , Nevo/fisiopatologia , Reação em Cadeia da Polimerase em Tempo Real , Fatores de Transcrição SOXE/genéticaRESUMO
Schwann cells produce myelin sheaths and thereby permit rapid saltatory conductance in the vertebrate peripheral nervous system. Their stepwise differentiation from neural crest cells is driven by a defined set of transcription factors. How this is linked to chromatin changes is not well understood. Here we show that the glial transcription factor Sox10 functions in Schwann cells by recruiting Brg1-containing chromatin-remodeling complexes via Baf60a to regulatory regions of Oct6 and Krox20 genes. It thereby stimulates expression of these transcriptional regulators that then cooperate with Sox10 to convert immature into myelinating Schwann cells. The functional interaction between Sox10 and Brg1 is evident from gain- and loss-of-function studies, similar neuropathies in the corresponding mouse mutants, and an aggravated neuropathy in compound mutants. Our results demonstrate that the transcription factor-mediated recruitment of the chromatin-remodeling machinery to specific genomic loci is an essential driving force for Schwann cell differentiation and myelination.