RESUMO
Unlike adult mammals, adult frogs regrow their optic nerve following a crush injury, making Xenopus laevis a compelling model for studying the molecular mechanisms that underlie neuronal regeneration. Using Translational Ribosome Affinity Purification (TRAP), a method to isolate ribosome-associated mRNAs from a target cell population, we have generated a transcriptional profile by RNA-Seq for retinal ganglion cells (RGC) during the period of recovery following an optic nerve injury. Based on bioinformatic analysis using the Xenopus laevis 9.1 genome assembly, our results reveal a profound shift in the composition of ribosome-associated mRNAs during the early stages of RGC regeneration. As factors involved in cell signaling are rapidly down-regulated, those involved in protein biosynthesis are up-regulated alongside key initiators of axon development. Using the new genome assembly, we were also able to analyze gene expression profiles of homeologous gene pairs arising from a whole-genome duplication in the Xenopus lineage. Here we see evidence of divergence in regulatory control among a significant proportion of pairs. Our data should provide a valuable resource for identifying genes involved in the regeneration process to target for future functional studies, in both naturally regenerative and non-regenerative vertebrates.
Assuntos
Proteínas do Olho/biossíntese , Regulação da Expressão Gênica , Regeneração Nervosa/fisiologia , Proteínas do Tecido Nervoso/biossíntese , Traumatismos do Nervo Óptico/fisiopatologia , Proteínas de Xenopus/biossíntese , Xenopus laevis/genética , Sequência de Aminoácidos , Animais , Animais Geneticamente Modificados , Proteínas do Olho/genética , Ontologia Genética , Anotação de Sequência Molecular , Compressão Nervosa , Proteínas do Tecido Nervoso/genética , RNA Mensageiro/genética , RNA Mensageiro/isolamento & purificação , Células Ganglionares da Retina/metabolismo , Ribossomos/metabolismo , Alinhamento de Sequência , Análise de Sequência de RNA , Homologia de Sequência de Aminoácidos , Transdução de Sinais , Proteínas de Xenopus/genética , Xenopus laevis/fisiologiaRESUMO
BACKGROUND: Translating Ribosome Affinity Purification (TRAP), a method recently developed to generate cell type-specific translational profiles, relies on creating transgenic lines of animals in which a tagged ribosomal protein is placed under regulatory control of a cell type-specific promoter. An antibody is then used to affinity purify the tagged ribosomes so that cell type-specific mRNAs can be isolated from whole tissue lysates. RESULTS: Here, cell type-specific transgenic lines were generated to enable TRAP studies for retinal ganglion cells and rod photoreceptors in the Xenopus laevis retina. Using real time quantitative PCR for assessing expression levels of cell type-specific mRNAs, the TRAP method was shown to selectively isolate mRNAs expressed in the targeted cell and was efficient at purifying mRNAs expressed at both high and low levels. Statistical measures used to distinguish cell type-specific RNAs from low level background and non-specific RNAs showed TRAP to be highly effective in Xenopus. CONCLUSIONS: TRAP can be used to purify mRNAs expressed in rod photoreceptors and retinal ganglion cells in X. laevis. The generated transgenic lines will enable numerous studies into the development, disease, and injury of the X. laevis retina.
Assuntos
Proteínas do Olho/biossíntese , Regulação da Expressão Gênica/fisiologia , Células Fotorreceptoras de Vertebrados/metabolismo , Biossíntese de Proteínas/fisiologia , Células Ganglionares da Retina/metabolismo , Proteínas de Xenopus/biossíntese , Animais , Perfilação da Expressão Gênica/métodos , Células Fotorreceptoras de Vertebrados/citologia , RNA Mensageiro/biossíntese , Reação em Cadeia da Polimerase em Tempo Real/métodos , Células Ganglionares da Retina/citologia , Xenopus laevisRESUMO
The present study examined Cromer's (1983) claim that children with language impairments have a hierarchical planning deficit that affects language as well as performance on complex construction tasks. Subjects were 30 boys (ages 5-7 years), 15 with specific language impairments (SLI) and 15 with normally developing language. Children were asked to build four hierarchical structures: a block construction, a puzzle construction, a simple straw construction, and a complex straw construction. Children who failed to complete the complex straw construction were taught how to construct the model using a sequential strategy. The two groups tended to perform comparably on the block and complex straw construction, the easiest and hardest of the four constructions. The two groups performed least comparably on the puzzle, simple straw construction, and the training task. On the basis of these findings and recent work by Greenfield (1991), we concluded that it is time to reject the notion that a central hierarchical planning mechanism underlies language and non-language structures that contain hierarchical components. The possible exception is early in development before language and manual actions become more autonomous and modular in nature.
Assuntos
Transtornos da Linguagem/diagnóstico , Criança , Linguagem Infantil , Pré-Escolar , Cognição , Humanos , Desenvolvimento da Linguagem , Testes de Linguagem , MasculinoRESUMO
5(18.5%) of a group 27 coppersmiths exposed to cadmium fume had stone disease. When compared with a control group of assembly workers in the same factory they had evidence of renal impairment as shown by blood biochemistry and proteinuria. A greater tendency to liver damage was found in the coppersmiths. There was evidence that restrictive airways disease was more common in the coppersmiths than in the control group. Blood-cadmium concentrations were significantly higher in the coppersmiths and in the assembly workers than in a reference population.