Assuntos
Translocador Nuclear Receptor Aril Hidrocarboneto/metabolismo , Leucemia/metabolismo , Receptores de Hidrocarboneto Arílico/metabolismo , Doença Aguda , Translocador Nuclear Receptor Aril Hidrocarboneto/genética , Humanos , Proteínas de Fusão Oncogênica/genética , Proteínas de Fusão Oncogênica/metabolismo , Proteínas Proto-Oncogênicas c-ets/genética , Proteínas Proto-Oncogênicas c-ets/metabolismo , Receptores de Hidrocarboneto Arílico/genética , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Variante 6 da Proteína do Fator de Translocação ETSRESUMO
Macrophages play an essential role in the resolution of tissue damage through removal of necrotic cells, thus paving the way for tissue regeneration. Macrophages also directly support the formation of new tissue to replace the injury, through their acquisition of an anti-inflammatory, or M2, phenotype, characterized by a gene expression program that includes IL-10, the IL-13 receptor, and arginase 1. We report that deletion of two CREB-binding sites from the Cebpb promoter abrogates Cebpb induction upon macrophage activation. This blocks the downstream induction of M2-specific Msr1, Il10, II13ra, and Arg-1 genes, whereas the inflammatory (M1) genes Il1, Il6, Tnfa, and Il12 are not affected. Mice carrying the mutated Cebpb promoter (betaDeltaCre) remove necrotic tissue from injured muscle, but exhibit severe defects in muscle fiber regeneration. Conditional deletion of the Cebpb gene in muscle cells does not affect regeneration, showing that the C/EBPbeta cascade leading to muscle repair is muscle-extrinsic. While betaDeltaCre macrophages efficiently infiltrate injured muscle they fail to upregulate Cebpb, leading to decreased Arg-1 expression. CREB-mediated induction of Cebpb expression is therefore required in infiltrating macrophages for upregulation of M2-specific genes and muscle regeneration, providing a direct genetic link between these two processes.
Assuntos
Proteína beta Intensificadora de Ligação a CCAAT/fisiologia , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/fisiologia , Regulação da Expressão Gênica , Macrófagos/fisiologia , Músculo Esquelético/fisiologia , Animais , Linfócitos B/fisiologia , Sítios de Ligação , Células da Medula Óssea/fisiologia , Proteína beta Intensificadora de Ligação a CCAAT/genética , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/genética , Camundongos , Regiões Promotoras Genéticas , Regeneração , Fatores de Transcrição/metabolismoRESUMO
The transcriptional regulators that couple interfollicular basal keratinocyte proliferation arrest to commitment and differentiation are yet to be identified. Here we report that the basic region leucine zipper transcription factors C/EBPalpha and C/EBPbeta are co-expressed in basal keratinocytes, and are coordinately upregulated as keratinocytes exit the basal layer and undergo terminal differentiation. Mice lacking both C/EBPalpha and beta in the epidermis showed increased proliferation of basal keratinocytes and impaired commitment to differentiation. This led to ectopic expression of keratin 14 (K14) and DeltaNp63 in suprabasal cells, decreased expression of spinous and granular layer proteins, parakeratosis and defective epidermal water barrier function. Knock-in mutagenesis revealed that C/EBP-E2F interaction was required for control of interfollicular epidermis (IFE) keratinocyte proliferation, but not for induction of spinous and granular layer markers, whereas C/EBP DNA binding was required for DeltaNp63 downregulation and K1/K10 induction. Finally, loss of C/EBPalpha/beta induced stem cell gene expression signatures in the epidermis. C/EBPs, therefore, couple basal keratinocyte cell cycle exit to commitment to differentiation through E2F repression and DNA binding, respectively, and may act to restrict the epidermal stem cell compartment.
Assuntos
Proteína alfa Estimuladora de Ligação a CCAAT/metabolismo , Proteína beta Intensificadora de Ligação a CCAAT/metabolismo , Diferenciação Celular/fisiologia , Proliferação de Células , Queratinócitos/metabolismo , Animais , Proteína alfa Estimuladora de Ligação a CCAAT/genética , Proteína beta Intensificadora de Ligação a CCAAT/genética , Diferenciação Celular/genética , Embrião de Mamíferos/metabolismo , Células Epidérmicas , Epiderme/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Queratina-14/genética , Queratina-14/metabolismo , Queratinócitos/citologia , Camundongos , Camundongos Knockout , Proteínas Nucleares/metabolismoRESUMO
BACKGROUND: Extracellular signaling through receptors for neurotrophins mediates diverse neuronal functions, including survival, migration and differentiation in the central nervous system, but the transcriptional targets and regulators that mediate these diverse neurotrophin functions are not well understood. RESULTS: We have identified the immediate-early (IE) genes Fos, Egr1 and Egr2 as transcriptional targets of brain derived neurotrophic factor (BDNF)/TrkB signaling in primary cortical neurons, and show that the Fos serum response element area responds to BDNF/TrkB in a manner dependent on a combined C/EBP-Ebox element. The Egr1 and Egr2 promoters contain homologous regulatory elements. We found that C/EBPalpha/beta and NeuroD formed complexes in vitro and in vivo, and were recruited to all three homologous promoter regions. C/EBPalpha and NeuroD co-operatively activated the Fos promoter in transfection assays. Genetic depletion of Trk receptors led to impaired recruitment of C/EBPs and NeuroD in vivo, and elimination of Cebpa and Cebpb alleles reduced BDNF induction of Fos, Egr1 and Egr2 in primary neurons. Finally, defective differentiation of cortical dendrites, as measured by MAP2 staining, was observed in both compound Cebp and Ntrk knockout mice. CONCLUSION: We here identify three IE genes as targets for BDNF/TrkB signaling, show that C/EBPalpha and -beta are recruited along with NeuroD to target promoters, and that C/EBPs are essential mediators of Trk signaling in cortical neurons. We show also that C/EBPs and Trks are required for cortical dendrite differentiation, consistent with Trks regulating dendritic differentiation via a C/EBP-dependent mechanism. Finally, this study indicates that BDNF induction of IE genes important for neuronal function depends on transcription factors (C/EBP, NeuroD) up-regulated during neuronal development, thereby coupling the functional competence of the neuronal cells to their differentiation.
Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Encéfalo/metabolismo , Proteínas Estimuladoras de Ligação a CCAAT/metabolismo , Genes Precoces/genética , Proteínas do Tecido Nervoso/metabolismo , Neurônios/metabolismo , Receptor trkB/metabolismo , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Encéfalo/anormalidades , Encéfalo/citologia , Fator Neurotrófico Derivado do Encéfalo/genética , Fator Neurotrófico Derivado do Encéfalo/metabolismo , Proteína alfa Estimuladora de Ligação a CCAAT/genética , Proteína alfa Estimuladora de Ligação a CCAAT/metabolismo , Proteína beta Intensificadora de Ligação a CCAAT/genética , Proteína beta Intensificadora de Ligação a CCAAT/metabolismo , Proteínas Estimuladoras de Ligação a CCAAT/genética , Diferenciação Celular/genética , Células Cultivadas , Dendritos/metabolismo , Dendritos/patologia , Proteína 1 de Resposta de Crescimento Precoce/genética , Proteína 1 de Resposta de Crescimento Precoce/metabolismo , Proteína 2 de Resposta de Crescimento Precoce/genética , Proteína 2 de Resposta de Crescimento Precoce/metabolismo , Regulação da Expressão Gênica no Desenvolvimento/genética , Camundongos , Camundongos Knockout , Camundongos Transgênicos , Células NIH 3T3 , Proteínas do Tecido Nervoso/genética , Neurônios/citologia , Regiões Promotoras Genéticas/genética , Proteínas Proto-Oncogênicas c-fos/genética , Proteínas Proto-Oncogênicas c-fos/metabolismo , Receptor trkB/genética , Elemento de Resposta Sérica/genética , Ativação Transcricional/genéticaRESUMO
TEL is a frequent target of chromosomal translocations in human cancer and an alleged tumor suppressor gene. TEL encodes two isoforms: a major TEL-M1 isoform as well as TEL-M43, which lacks the first 42 amino acid residues of TEL-M1. Both isoforms are potent transcriptional repressors that can inhibit RAS-induced transformation. Here we show that the v-SRC protein-tyrosine kinase relieves the repressive activity of TEL-M1, an activity that is associated with the v-SRC-induced delocalization of TEL-M1 from the nucleus to the cytoplasm. TEL-M1 delocalization requires the kinase activity of v-SRC and is not induced by oncogenic RAS or AKT. Cytoplasmic delocalization of TEL-M1 in response to v-SRC critically depends upon its unique amino-terminal domain (SRCD domain) because (i). v-SRC did not inhibit the repressive properties of TEL-M43, nor affected TEL-M43 nuclear localization; (ii). fusion of the first 52 amino acid residues of TEL-M1 to FLI-1, an ETS protein insensitive to v-SRC-induced delocalization, is sufficient to confer v-SRC-induced delocalization to this TEL/FLI-1 chimeric protein. The v-SRC-induced nucleo-cytoplasmic delocalization of TEL-M1 does not involve phosphorylation of the SRCD and does not require TEL self-association and repressive domains. Finally, enforced expression of the v-SRC-insensitive TEL-M43, but not of TEL-M1, inhibits v-SRC-induced transformation of NIH3T3 fibroblasts. These results identify a regulatory domain in TEL that specifically impinges on the subcellular localization of its major TEL-M1 isoform. They, furthermore, indicate that inhibition of TEL-M1 nuclear function is required for v-SRC to induce cellular transformation.
