RESUMO
Endoplasmic reticulum (ER) stress, defective autophagy and genomic instability in the central nervous system are often associated with severe developmental defects and neurodegeneration. Here, we reveal the role played by Rint1 in these different biological pathways to ensure normal development of the central nervous system and to prevent neurodegeneration. We found that inactivation of Rint1 in neuroprogenitors led to death at birth. Depletion of Rint1 caused genomic instability due to chromosome fusion in dividing cells. Furthermore, Rint1 deletion in developing brain promotes the disruption of ER and Cis/Trans Golgi homeostasis in neurons, followed by ER-stress increase. Interestingly, Rint1 deficiency was also associated with the inhibition of the autophagosome clearance. Altogether, our findings highlight the crucial roles of Rint1 in vivo in genomic stability maintenance, as well as in prevention of ER stress and autophagy.
Assuntos
Autofagia , Encéfalo/metabolismo , Estresse do Retículo Endoplasmático , Instabilidade Genômica , Proteínas Supressoras de Tumor/genética , Proteínas de Transporte Vesicular/genética , Animais , Apoptose , Encéfalo/citologia , Células Cultivadas , Camundongos da Linhagem 129 , Camundongos Transgênicos , Mitose , Células-Tronco Neurais/fisiologia , Cultura Primária de Células , Células de Purkinje/fisiologia , Proteínas Supressoras de Tumor/metabolismo , Proteínas de Transporte Vesicular/metabolismoRESUMO
The identification of key tumorigenic events in Sonic Hedgehog (SHH) subgroup medulloblastomas (MBSHH) will be essential for the development of individualized therapies and improved outcomes. However, beyond confirmation of characteristic SHH pathway mutations, recent genome-wide sequencing studies have not revealed commonly mutated genes with widespread relevance as potential therapeutic targets. We therefore examined any role for epigenetic DNA methylation events in MBSHH using a cross-species approach to candidate identification, prioritization and validation. MBSHH-associated DNA methylation events were first identified in 216 subgrouped human medulloblastomas (50 MBSHH, 28 Wnt/Wingless, 44 Group 3 and 94 Group 4) and their conservation then assessed in tumors arising from four independent murine models of Shh medulloblastoma, alongside any role in tumorigenesis using functional assessments in mouse and human models. This strategy identified widespread regional CpG hypo-methylation of VAV1, leading to its elevated expression, as a conserved aberrant epigenetic event, which characterizes the majority of MBSHH tumors in both species, and is associated with a poor outcome in MBSHH patients. Moreover, direct modulation of VAV1 in mouse and human models revealed a critical role in tumor maintenance, and its abrogation markedly reduced medulloblastoma growth. Further, Vav1 activity regulated granule neuron precursor germinal zone exit and migration initiation in an ex vivo model of early postnatal cerebellar development. These findings establish VAV1 as a critical epigenetically regulated oncogene with a key role in MBSHH maintenance, and highlight its potential as a validated therapeutic target and prognostic biomarker for the improved therapy of medulloblastoma.
Assuntos
Metilação de DNA/genética , Epigênese Genética , Meduloblastoma/genética , Proteínas Proto-Oncogênicas c-vav/genética , Animais , Proliferação de Células , Transformação Celular Neoplásica/genética , Ilhas de CpG/genética , Humanos , Meduloblastoma/patologia , Camundongos , Neurônios/metabolismo , Neurônios/patologia , Proteínas Proto-Oncogênicas c-vav/biossíntese , Transdução de Sinais , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Normal development of an organism requires the ability to respond to DNA damage. A particularly deleterious lesion is a DNA double-strand break (DSB). The cellular response to DNA DSBs occurs via an integrated sensing and signaling network that maintains genomic stability. The outcomes of defective DNA DSB repair are related to the developmental stage of an organism, and often show striking tissue specificity. Many human diseases are associated with deficiencies in DNA DSB repair and can be characterized by neuropathology, immune deficiency, growth retardation or predisposition to cancer. This review will focus on the requirements of the DNA DSB response that function to maintain homeostasis during mammalian development.
Assuntos
Quebras de DNA de Cadeia Dupla , Reparo do DNA , Mamíferos/embriologia , Recombinação Genética , Animais , Humanos , Mamíferos/fisiologiaRESUMO
The Sonic Hedgehog (SHH) signaling pathway is indispensable for development, and functions to activate a transcriptional program modulated by the GLI transcription factors. Here, we report that loss of a regulator of the SHH pathway, Suppressor of Fused (Sufu), resulted in early embryonic lethality in the mouse similar to inactivation of another SHH regulator, Patched1 (Ptch1). In contrast to Ptch1+/- mice, Sufu+/- mice were not tumor prone. However, in conjunction with p53 loss, Sufu+/- animals developed tumors including medulloblastoma and rhabdomyosarcoma. Tumors present in Sufu+/-p53-/- animals resulted from Sufu loss of heterozygosity. Sufu+/-p53-/- medulloblastomas also expressed a signature gene expression profile typical of aberrant SHH signaling, including upregulation of N-myc, Sfrp1, Ptch2 and cyclin D1. Finally, the Smoothened inhibitor, hedgehog antagonist, did not block growth of tumors arising from Sufu inactivation. These data demonstrate that Sufu is essential for development and functions as a tumor suppressor.
Assuntos
Transformação Celular Neoplásica/genética , Genes Letais , Predisposição Genética para Doença , Meduloblastoma/genética , Proteínas Repressoras/fisiologia , Rabdomiossarcoma/genética , Animais , Apoptose , Transformação Celular Neoplásica/metabolismo , Perfilação da Expressão Gênica , Genes Supressores de Tumor/fisiologia , Perda de Heterozigosidade , Meduloblastoma/patologia , Camundongos , Camundongos Knockout , Receptores Patched , Receptor Patched-1 , Receptor Patched-2 , Receptores de Superfície Celular/genética , Receptores de Superfície Celular/fisiologia , Proteínas Repressoras/genética , Rabdomiossarcoma/patologia , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/fisiologiaRESUMO
Within the nervous system appropriate responses to DNA damage are required to maintain homeostasis and prevent disease. In this tissue, DNA double-strand breaks (DSBs) initiate a molecular response to repair DNA, or in many cases, activate apoptosis. The repair of DNA DSBs occurs via nonhomologous end-joining (NHEJ) or homologous recombination (HR). These mechanistically distinct pathways are critical for maintenance of genomic integrity. During nervous system development there are discrete requirements for each DNA DSB repair pathway at different stages of development. For example, in the nervous system HR is particularly important for proliferating cells, while NHEJ is critical for differentiating cells. Inactivation of either of these pathways can lead to embryonic lethality, neurodegeneration or brain tumors. Human syndromes that result from defective responses to DNA damage often feature overt neuropathology. A prime example is the neurodegenerative syndrome ataxia telangiectasia (A-T), which results from inactivation of the ATM kinase, a crucial nexus for the cellular response to DNA DSBs. This type of DNA damage activates ATM via the Mre11-Rad50-NBS1 (MRN) complex, which leads to selective phosphorylation of ATM substrates resulting in apoptosis or cell cycle arrest and DNA repair. Furthermore, DNA DSBs resulting from chronic genotoxic stress can also result in tumorigenesis, as inactivation of either HR or NHEJ can lead to certain types of brain tumors. Thus, there are distinct requirements for each DNA DSB repair pathway during neural development, which have important implications for understanding diseases of the nervous system.
Assuntos
Quebras de DNA de Cadeia Dupla , Dano ao DNA/genética , Doenças do Sistema Nervoso/genética , Sistema Nervoso/metabolismo , Animais , Ataxia Telangiectasia/genética , Ataxia Telangiectasia/metabolismo , Ataxia Telangiectasia/fisiopatologia , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/metabolismo , Reparo do DNA/genética , Enzimas Reparadoras do DNA/genética , Enzimas Reparadoras do DNA/metabolismo , Humanos , Sistema Nervoso/fisiopatologia , Doenças do Sistema Nervoso/metabolismo , Doenças do Sistema Nervoso/fisiopatologiaRESUMO
Ataxia-telangiectasia (A-T) is a neurodegenerative syndrome resulting from dysfunction of ATM (ataxia telangiectasia mutated). The molecular details of ATM function in the nervous system are unclear, although the neurological lesions in A-T are probably developmental because they appear during childhood. The nervous systems of Atm-null mice show a pronounced defect in apoptosis that is induced by DNA damage, suggesting that ATM may function to eliminate DNA-damaged neurons. Here we show that Atm-dependent apoptosis occurs at discrete stages of neurogenesis. Analysis of gamma-irradiated mouse embryos showed that Atm-dependent apoptosis occurred only in the postmitotic populations that were present in the neuroepithelial subventricular zone of the developing nervous system. Notably, Atm deficiency did not prevent radiation-induced apoptosis in multipotent precursor cells residing in the proliferating ventricular zone. Atm-dependent apoptosis required p53 and coincided with the specific phosphorylation of p53 and caspase-3 activation. Thus, these data show that Atm functions early in neurogenesis and underscore the selective requirement for Atm in eliminating damaged postmitotic neural cells. Furthermore, these data demonstrate that the differentiation status of neural cells is a critical determinant in the activation of certain apoptotic pathways.
Assuntos
Apoptose/efeitos da radiação , Raios gama , Sistema Nervoso/citologia , Sistema Nervoso/efeitos da radiação , Proteínas Serina-Treonina Quinases/metabolismo , Animais , Apoptose/fisiologia , Ataxia Telangiectasia/genética , Proteínas Mutadas de Ataxia Telangiectasia , Caspase 3 , Caspases/metabolismo , Proteínas de Ciclo Celular , Diferenciação Celular/fisiologia , Diferenciação Celular/efeitos da radiação , Divisão Celular/genética , Divisão Celular/efeitos da radiação , Dano ao DNA , Reparo do DNA , Proteínas de Ligação a DNA , Embrião de Mamíferos/citologia , Embrião de Mamíferos/efeitos da radiação , Feminino , Camundongos , Camundongos Knockout , Mitose/efeitos da radiação , Sistema Nervoso/embriologia , Sistema Nervoso/metabolismo , Neurônios/citologia , Neurônios/efeitos da radiação , Fosforilação/efeitos da radiação , Gravidez , Efeitos Tardios da Exposição Pré-Natal , Proteínas Serina-Treonina Quinases/deficiência , Proteínas Serina-Treonina Quinases/genética , Retina/citologia , Retina/crescimento & desenvolvimento , Retina/efeitos da radiação , Células-Tronco/citologia , Células-Tronco/efeitos da radiação , Proteína Supressora de Tumor p53/deficiência , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo , Proteínas Supressoras de TumorRESUMO
Despite the rarity of the human autosomal recessive disease ataxia telangiectasia (A-T) (affecting approximately 1/40000-1/100000), interest in the function of the mutated gene product (ATM) in this syndrome is intense. Mutation of this single gene can lead to a diverse array of features, including cancer, immune defects, infertility and radiosensitivity. However, it is the pronounced and debilitating neurodegeneration that is the hallmark of this disease. Thus, from a clinical perspective, it is ATM function in the nervous system that, arguably, is the most important to understand. Although the case for DNA damage as a causative factor for neurodegeneration in A-T is compelling, new data point to a possible link to defects in neurogenesis. Thus, whereas ATM is important for nervous system development, it could also be important for adult neurogenesis.
Assuntos
Ataxia Telangiectasia/genética , Ataxia Telangiectasia/patologia , Animais , Proteínas Mutadas de Ataxia Telangiectasia , Encéfalo/metabolismo , Proteínas de Ciclo Celular , Sobrevivência Celular , Dano ao DNA , Proteínas de Ligação a DNA , Humanos , Camundongos , Camundongos Knockout , Mutação , Neurônios/metabolismo , Proteínas Serina-Treonina Quinases/genética , Células-Tronco/metabolismo , Proteínas Supressoras de TumorRESUMO
Ataxia telangiectasia results from mutations of ATM and is characterized by severe neurodegeneration and defective responses to DNA damage. Inactivation of certain DNA repair genes such as DNA ligase IV results in massive neuronal apoptosis and embryonic lethality in the mouse, indicating the occurrence of endogenously formed DNA double-strand breaks during nervous system development. Here we report that Atm is required for apoptosis in all areas of the DNA ligase IV-deficient developing nervous system. However, Atm deficiency failed to rescue deficits in immune differentiation in DNA ligase IV-null mice. These data indicate that ATM responds to endogenous DNA lesions and functions during development to eliminate neural cells that have incurred genomic damage. Therefore, ATM could be important for preventing accumulation of DNA-damaged cells in the nervous system that might eventually lead to the neurodegeneration observed in ataxia telangiectasia.
Assuntos
DNA Ligases/deficiência , Neurônios/patologia , Proteínas Serina-Treonina Quinases/metabolismo , Animais , Animais Recém-Nascidos , Antígenos Nucleares , Apoptose/fisiologia , Ataxia Telangiectasia , Proteínas Mutadas de Ataxia Telangiectasia , Encéfalo/anatomia & histologia , Encéfalo/crescimento & desenvolvimento , Caspase 3 , Caspases/metabolismo , Proteínas de Ciclo Celular , Dano ao DNA , DNA Ligase Dependente de ATP , DNA Ligases/genética , Proteínas de Ligação a DNA , Embrião de Mamíferos/patologia , Camundongos , Camundongos Knockout , Proteínas Nucleares/metabolismo , Proteínas Serina-Treonina Quinases/genética , Transdução de Sinais , Linfócitos T/imunologia , Linfócitos T/patologia , Proteína Supressora de Tumor p53/metabolismo , Proteínas Supressoras de TumorRESUMO
Many human pathological conditions with genetic defects in DNA damage responses are also characterized by neurological deficits. These neurological deficits can manifest themselves during many stages of development, suggesting an important role for DNA repair or processing during the development and maintenance of the nervous system. Although the molecular neuropathology associated with such deficits is largely unknown, many of the responsible gene defects have been identified. The current rapid progress in elucidation of molecular details following gene identification should provide further insight into the importance of DNA processing in nervous system function.
Assuntos
Dano ao DNA/fisiologia , Transtornos Heredodegenerativos do Sistema Nervoso/genética , Transtornos Heredodegenerativos do Sistema Nervoso/fisiopatologia , Degeneração Neural/genética , Degeneração Neural/fisiopatologia , Ataxia Telangiectasia/fisiopatologia , Síndrome de Bloom , Reparo do DNA , Anemia de Fanconi/fisiopatologia , Humanos , Síndrome de Rothmund-Thomson/fisiopatologia , Síndrome de Werner/fisiopatologia , Xeroderma Pigmentoso/fisiopatologiaRESUMO
Ataxia-telangiectasia is a hereditary multisystemic disease resulting from mutations of ataxia telangiectasia, mutated (ATM) and is characterized by neurodegeneration, cancer, immune defects, and hypersensitivity to ionizing radiation. The molecular details of ATM function in the nervous system are unclear, although the neurological lesion in ataxia-telangiectasia becomes apparent early in life, suggesting a developmental origin. The central nervous system (CNS) of Atm-null mice shows a pronounced defect in apoptosis induced by genotoxic stress, suggesting ATM functions to eliminate neurons with excessive genomic damage. Here, we report that the death effector Bax is required for a large proportion of Atm-dependent apoptosis in the developing CNS after ionizing radiation (IR). Although many of the same regions of the CNS in both Bax-/- and Atm-/- mice were radioresistant, mice nullizygous for both Bax and Atm showed additional reduction in IR-induced apoptosis in the CNS. Therefore, although the major IR-induced apoptotic pathway in the CNS requires Atm and Bax, a p53-dependent collateral pathway exists that has both Atm- and Bax-independent branches. Further, Atm- and Bax-dependent apoptosis in the CNS also required caspase-3 activation. These data implicate Bax and caspase-3 as death effectors in neurodegenerative pathways.
Assuntos
Apoptose/efeitos da radiação , Sistema Nervoso Central/efeitos da radiação , Proteínas Serina-Treonina Quinases/fisiologia , Proteínas Proto-Oncogênicas c-bcl-2 , Proteínas Proto-Oncogênicas/fisiologia , Animais , Apoptose/genética , Proteínas Mutadas de Ataxia Telangiectasia , Western Blotting , Caspase 3 , Caspases/metabolismo , Proteínas de Ciclo Celular , Sistema Nervoso Central/citologia , Sistema Nervoso Central/metabolismo , Cerebelo/metabolismo , Cerebelo/efeitos da radiação , Proteínas de Ligação a DNA , Giro Denteado/metabolismo , Giro Denteado/efeitos da radiação , Ativação Enzimática/efeitos da radiação , Genótipo , Camundongos , Camundongos Knockout , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas/metabolismo , Radiação Ionizante , Retina/metabolismo , Retina/efeitos da radiação , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo , Proteína Supressora de Tumor p53/fisiologia , Proteínas Supressoras de Tumor , Proteína X Associada a bcl-2RESUMO
Ataxia-telangiectasia is a human syndrome resulting from mutations of the ATM protein kinase that is characterized by radiation sensitivity and neurodegeneration. Although neuroprotective, the molecular details of ATM function in the nervous system are uncertain. However, in the mouse, Atm is essential for ionizing radiation-induced apoptosis in select postmitotic populations of the developing nervous system. Atm-dependent apoptosis in the nervous system also requires p53, consistent with the well-established link of p53 as a major substrate of ATM. Furthermore, the proapoptotic effector Bax is also required for most, but not all, Atm-dependent apoptosis. Therefore, after DNA damage in the developing nervous system, Atm initiates a p53-dependent apoptotic cascade in differentiating neural cells. Together, these data suggest ATM-dependent apoptosis may be important for elimination of neural cells that have accumulated genomic damage during development, thus preventing dysfunction of these cells later in life.
Assuntos
Apoptose/genética , Ataxia Telangiectasia/genética , Sistema Nervoso Central/anormalidades , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Animais , Ataxia Telangiectasia/metabolismo , Ataxia Telangiectasia/fisiopatologia , Proteínas Mutadas de Ataxia Telangiectasia , Proteínas de Ciclo Celular , Sistema Nervoso Central/metabolismo , Sistema Nervoso Central/fisiopatologia , Proteínas de Ligação a DNA , Humanos , Degeneração Neural/genética , Degeneração Neural/metabolismo , Degeneração Neural/fisiopatologia , Transdução de Sinais/genética , Proteína Supressora de Tumor p53/metabolismo , Proteínas Supressoras de TumorRESUMO
Sc1 is an extracellular matrix-associated protein whose function is unknown. During early embryonic development, Sc1 is widely expressed, and from embryonic day 12 (E12), Sc1 is expressed primarily in the developing nervous system. This switch in Sc1 expression at E12 suggests an importance for nervous-system development. To gain insight into Sc1 function, we used gene targeting to inactivate mouse Sc1. The Sc1-null mice showed no obvious deficits in any organs. These mice were born at the expected ratios, were fertile, and had no obvious histological abnormalities, and their long-term survival did not differ from littermate controls. Therefore, the function of Sc1 during development is not critical or, in its absence, is subserved by another protein.
Assuntos
Molécula de Adesão de Leucócito Ativado/metabolismo , Envelhecimento/metabolismo , Desenvolvimento Embrionário e Fetal , Matriz Extracelular/metabolismo , Molécula de Adesão de Leucócito Ativado/genética , Animais , Epitélio/metabolismo , Éxons/genética , Fertilidade/genética , Deleção de Genes , Perfilação da Expressão Gênica , Genes Essenciais/genética , Hibridização In Situ , Longevidade/genética , Mesoderma/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Sistema Nervoso/embriologia , Sistema Nervoso/crescimento & desenvolvimento , Sistema Nervoso/metabolismo , RNA Mensageiro/análise , RNA Mensageiro/genéticaRESUMO
The cancer-prone neurodegenerative disorder, ataxia telangiectasia (A-T), results from mutations of ATM (ataxia telangiectasia mutated). Individuals with A-T are also hypersensitive to ionizing radiation (IR). Cultured cells from A-T individuals or Atm-/- mice have cell cycle and growth defects and are generally considered radiosensitive. However, it has been shown recently that cell populations in the Atm-/- central nervous system are radioresistant. To define specific IR sensitivities of neural populations, we analyzed Atm-/- astrocytes. Here we show that Atm-/- astrocytes exhibit premature senescence, express constitutively high levels of p21, and have impaired p53 stabilization. However, in contrast to radiosensitive Atm-/- fibroblasts and radioresistant Atm-/- neurons, survival of Atm-/- astrocytes after IR was similar to wild-type astrocytes. Additionally, p53-null astrocytes, but not fibroblasts, were moderately more radioresistant than their wild-type counterparts, suggesting that the deficit in p53 stabilization observed in Atm-null cells is not a measure of radiation susceptibility. Thus, in astrocytes, the function of Atm in cellular growth and radiosensitivity is distinct. These data may have implications for ATM disruption strategies as a radiosensitizing treatment for brain tumors.
Assuntos
Astrócitos/efeitos da radiação , Ataxia Telangiectasia/genética , Proteínas Serina-Treonina Quinases , Proteínas/fisiologia , Tolerância a Radiação , Animais , Astrócitos/fisiologia , Proteínas Mutadas de Ataxia Telangiectasia , Proteínas de Ciclo Celular , Divisão Celular , Células Cultivadas , Proteínas de Ligação a DNA , Humanos , Camundongos , Camundongos Knockout , Mutação , Antígeno Nuclear de Célula em Proliferação/análise , Proteínas/genética , Proteína Supressora de Tumor p53/fisiologia , Proteínas Supressoras de TumorRESUMO
The alternative reading frame product (p19ARF) of the mouse INK4a/ARF locus is induced by oncoproteins such as Myc and E1A as part of a checkpoint response that limits cell cycle progression in response to hyperproliferative signals. ARF binds directly to Mdm2 to prevent down-regulation of p53 and thereby promotes p53-dependent transcription and cell cycle arrest. However, ARF is not required for p53 induction in response to ionizing radiation or other forms of DNA damage. Animals lacking a functional ataxia telangiectasia (Atm) gene are exquisitely sensitive to ionizing radiation; Atm-null mouse embryo fibroblasts (MEFs) undergo premature replicative arrest, which is relieved by the loss of p53. Here we show that the loss of ARF expands the life expectancy of Atm-null MEFs, but alters neither the sensitivity of Atm-null mice to ionizing radiation nor their propensity to develop lymphomas early in life. Therefore, whereas ARF and Atm signal to p53 through distinct pathways, the loss of ARF can modify p53-dependent features of the Atm-null phenotype.
Assuntos
Proteínas Nucleares , Proteínas Serina-Treonina Quinases , Proteínas/genética , Proteínas/fisiologia , Tolerância a Radiação/genética , Animais , Proteínas Mutadas de Ataxia Telangiectasia , Ciclo Celular/genética , Proteínas de Ciclo Celular , Células Cultivadas , Dano ao DNA , Reparo do DNA/genética , Proteínas de Ligação a DNA , Embrião de Mamíferos/citologia , Fibroblastos/citologia , Fibroblastos/efeitos da radiação , Linfoma/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fenótipo , Fosforilação , Processamento de Proteína Pós-Traducional , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Proto-Oncogênicas c-mdm2 , Proteína Supressora de Tumor p14ARF , Proteína Supressora de Tumor p53/metabolismo , Proteínas Supressoras de TumorRESUMO
Ataxia-telangiectasia is a human autosomal recessive disease characterized by neurodegeneration, cancer predisposition and sensitivity to ionizing radiation. One of the earliest features of this disease is ataxia, which is thought to be attributable to a progressive cerebellar degeneration associated with a disruption of Purkinje cell cytoarchitecture and positioning. To investigate the neuropathology of ataxia-telangiectasia, we used in situ hybridization to map Atm (the gene mutated in ataxia-telangiectasia) expression during mouse development. Atm expression was highest in the embryonic mouse nervous system, where it was predominantly associated with regions undergoing mitosis. During the period of Purkinje cell neurogenesis, Atm was highly expressed in the area containing Purkinje cell precursors (the ventricular zone of the fourth ventricle). However, in the postnatal cerebellum, Atm expression in Purkinje cells was very low, while expression in proliferating granule neurons was high. The only region of the adult nervous system that exhibited elevated Atm expression were the postmitotic sensory neurons of the dorsal root ganglia. The data suggest an early developmental requirement for ATM in the cerebellum, and other regions of the central nervous system, and a potential contribution of the dorsal root ganglia/sensory input pathway to the ataxic phenotype of ataxia-telangiectasia.
Assuntos
Ataxia Telangiectasia/genética , Regulação da Expressão Gênica no Desenvolvimento/genética , Regulação da Expressão Gênica no Desenvolvimento/fisiologia , Genes/genética , Mutação/fisiologia , Sistema Nervoso Periférico/fisiologia , Animais , Northern Blotting , Química Encefálica/genética , Cerebelo/crescimento & desenvolvimento , Cerebelo/metabolismo , Gânglios Espinais/crescimento & desenvolvimento , Gânglios Espinais/metabolismo , Humanos , Hibridização In Situ , Camundongos , Camundongos Endogâmicos , FenótipoRESUMO
Ataxia telangiectasia (AT) is characterized by progressive neurodegeneration that results from mutation of the ATM gene. However, neither the normal function of ATM in the nervous system nor the biological basis of the degeneration in AT is known. Resistance to apoptosis in the developing central nervous system (CNS) of Atm-/- mice was observed after ionizing radiation. This lack of death occurred in diverse regions of the CNS, including the cerebellum, which is markedly affected in AT. In wild-type, but not Atm-/- mice, up-regulation of p53 coincided with cell death, suggesting that Atm-dependent apoptosis in the CNS is mediated by p53. Further, p53 null mice showed a similar lack of radiation-induced cell death in the developing nervous system. Atm may function at a developmental survival checkpoint that serves to eliminate neurons with excessive DNA damage.
Assuntos
Apoptose , Encéfalo/citologia , Encéfalo/efeitos da radiação , Neurônios/citologia , Proteínas Serina-Treonina Quinases , Proteínas/fisiologia , Animais , Proteínas Mutadas de Ataxia Telangiectasia , Proteínas de Ciclo Celular , Cerebelo/citologia , Cerebelo/efeitos da radiação , Proteínas de Ligação a DNA , Genes p53 , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Neurônios/efeitos da radiação , Fenótipo , Proteínas/genética , Radiação Ionizante , Retina/citologia , Timo/citologia , Timo/efeitos da radiação , Proteína Supressora de Tumor p53/fisiologia , Proteínas Supressoras de Tumor , Regulação para CimaRESUMO
Sc1 and Sparc are two extracellular proteins sharing similarity in their carboxyl terminus, with 63% identity over a 232-amino-acid region. We have cloned and mapped the genomic locus of mouse Sc1. The mouse Sc1 gene contains 11 exons spanning approximately 35 kb of DNA. The genomic structure (exon/intron boundaries) of Sc1 exons 6 to exon 11 is identical to those of the similar portion of the Sparc gene. This suggests that Sc1 and Sparc originated from a common ancestral gene. Using fluorescence in situ hybridization analysis, Sc1 was localized to band 5E4 of mouse chromosome 5.
Assuntos
Proteínas da Matriz Extracelular/genética , Proteínas do Tecido Nervoso/genética , Osteonectina/genética , Molécula de Adesão de Leucócito Ativado , Sequência de Aminoácidos , Animais , Sequência de Bases , Mapeamento Cromossômico , Cromossomos , Clonagem Molecular , Evolução Molecular , Éxons/genética , Hibridização in Situ Fluorescente , Camundongos , Dados de Sequência Molecular , Análise de Sequência , Homologia de SequênciaRESUMO
Astrocytes are the most abundant cell type in the mammalian central nervous system (CNS), and are involved in many processes critical for normal CNS maintenance and function. We have used double-label immunocytochemistry and in situ analysis to show that the SPARC (secreted protein acidic and rich in cysteine)-related protein SC1, co-localizes with the astrocyte marker glial fibrillary acidic protein (GFAP) in the adult rodent brain. Thus, SC1 is an astrocyte marker that may be used to investigate astrocyte heterogeneity and analyze glial cell lineages during neural development. Consistent with the presence of SC1 and GFAP in astrocytes, both proteins were markedly upregulated following reactive astrocytosis induced by focal mechanical trauma. Therefore, SC1 may play an important role in reactive astrocytosis subsequent to a wide variety of neural trauma, including neurodegenerative diseases and acute neural damage.
Assuntos
Astrócitos/metabolismo , Encéfalo/metabolismo , Proteínas da Matriz Extracelular/metabolismo , Gliose/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Molécula de Adesão de Leucócito Ativado , Animais , Sequência de Bases , Encéfalo/patologia , Lesões Encefálicas/metabolismo , Lesões Encefálicas/patologia , Proteínas da Matriz Extracelular/genética , Proteína Glial Fibrilar Ácida/metabolismo , Gliose/patologia , Camundongos , Camundongos Endogâmicos , Sondas Moleculares/genética , Dados de Sequência Molecular , Proteínas do Tecido Nervoso/genética , Reação em Cadeia da Polimerase , RNA Mensageiro/metabolismo , Distribuição Tecidual , Ferimentos Perfurantes/metabolismo , Ferimentos Perfurantes/patologiaRESUMO
The rod and cone transducins are specific G proteins originally thought to be present only in photoreceptor cells of the vertebrate retina. Transducins convert light stimulation of photoreceptor opsins into activation of cyclic GMP phosphodiesterase (reviewed in refs. 5-7). A transducin-like G protein, gustducin, has been identified and cloned from rat taste cells. We report here that rod transducin is also present in vertebrate taste cells, where it specifically activates a phosphodiesterase isolated from taste tissue. Furthermore, the bitter compound denatonium in the presence of taste-cell membranes activates transducin but not Gi. A peptide that competitively inhibits rhodopsin activation of transducin also blocks taste-cell membrane activation of transducin, arguing for the involvement of a seven-transmembrane-helix G-protein-coupled receptor. These results suggest that rod transducin transduces bitter taste by coupling taste receptor(s) to taste-cell phosphodiesterase. Phosphodieterase-mediated degradation of cyclic nucleotides may lead to taste-cell depolarization through the recently identified cyclic-nucleotide-suppressible conductance.
Assuntos
Diester Fosfórico Hidrolases/metabolismo , Papilas Gustativas/metabolismo , Transducina/metabolismo , Animais , Bovinos , Ativação Enzimática , Proteínas de Ligação ao GTP/metabolismo , Células Fotorreceptoras/metabolismo , Reação em Cadeia da Polimerase , Testes de Precipitina , Compostos de Amônio Quaternário/farmacologia , RNA Mensageiro/metabolismo , Ratos , Rodopsina/metabolismo , Papilas Gustativas/efeitos dos fármacos , Papilas Gustativas/enzimologiaRESUMO
Pineal opsin (P-opsin), an opsin from chick that is highly expressed in pineal but is not detectable in retina, was cloned by the polymerase chain reaction. It is likely that the P-opsin lineage diverged from the retinal opsins early in opsin evolution. The amino acid sequence of P-opsin is 42 to 46 percent identical to that of the retinal opsins. P-opsin is a seven-membrane spanning, G protein-linked receptor with a Schiff-base lysine in the seventh membrane span and a Schiff-base counterion in the third membrane span. The primary sequence of P-opsin suggests that it will be maximally sensitive to approximately 500-nanometer light and produce a slow and prolonged phototransduction response consistent with the nonvisual function of pineal photoreception.