RESUMO
The TOR (target of rapamycin) pathway is a phylogenetically conserved transduction system in eukaryotes linking the energy status of the cell to the protein synthesis apparatus and to cell growth. The TOR protein is specifically inhibited by a rapamycin-FKBP12 complex (where FKBP stands for FK506-binding protein) in yeast and animal cells. Whereas plants appear insensitive to rapamycin, Arabidopsis thaliana harbours a single TOR gene, which is essential for embryonic development. It was found that the product of this gene was capable of binding to rapamycin and yeast FKBP12. In-frame fusion with a GUS reporter gene shows that the TOR protein is produced essentially in proliferating zones, whereas the TOR mRNA can be detected in all organs suggesting a translational regulation of TOR. Phenotypic analysis of Arabidopsis TOR mutants indicates that the plant TOR pathway fulfils the same role in controlling cell growth as its other eukaryotic counterparts.
Assuntos
Desenvolvimento Vegetal , Biossíntese de Proteínas , Regulação da Expressão Gênica de Plantas , Plantas/genética , Processamento Pós-Transcricional do RNARESUMO
In mammalian, insect, and yeast cells, TOR proteins are essential regulators of cell growth in response to environmental signals including nutrients, mitogens, and stresses. Although many aspects of the TOR-dependent signalling pathway are conserved between animals and fungi, important differences have also been found and are likely to be related to the ecophysiological adaptations of these organisms. The TOR protein also exists in plants. This review will first discuss specific aspects of plants concerning the contribution of cell growth to overall growth, as well as their responses to nutrient starvation, with emphasis on recent results obtained through genetic analysis in the model plant Arabidopsis thaliana. This is followed by the current status of the genetic analysis of the TOR gene in this plant and the search for potential members of a TOR pathway in the Arabidopsis genome.
Assuntos
Arabidopsis/crescimento & desenvolvimento , Fosfatidilinositol 3-Quinases/fisiologia , Fosfotransferases (Aceptor do Grupo Álcool)/fisiologia , Proteínas de Plantas/fisiologia , Arabidopsis/enzimologia , Ciclo Celular/fisiologia , Fosfatidilinositol 3-Quinases/metabolismo , Fosfotransferases (Aceptor do Grupo Álcool)/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Transdução de Sinais/fisiologia , Sirolimo/metabolismo , Sirolimo/farmacologiaRESUMO
We characterized two Arabidopsis thaliana cDNAs coding for class I valyl-tRNA synthetase and class II threonyl-tRNA synthetase. The proteins display characteristics of cytosolic enzymes, yet possess an N-terminal extension relative to their prokaryotic homologs. The proximal part of the N-terminal extension is a mitochondrial-targeting signal. Through transient expression of GFP fusions in tobacco cells, we demonstrated that both genes encode the cytosolic and mitochondrial forms of the enzymes by alternative use of two in-frame initiation codons. A long, mitochondrial form of the enzyme is translated from a first initiation codon at reduced levels because of a poor sequence context and a shorter, cytosolic form is translated from a second in-phase AUG, which is in a better context for translation initiation. Primer extension experiments revealed several transcript ends mapping upstream of the first AUG and between the two AUGs. Distal to the mitochondrial transit peptide both valyl-tRNA synthetase and threonyl tRNA synthetase possess an NH2-appended domain compared with their prokaryotic counterparts. This domain's amphiphilic helix is conserved between yeast and A. thaliana valyl-tRNA synthetase, suggesting an important role in translation. Based on the high structural similarities between yeast and A. thaliana valyl-tRNA synthetase, we propose that the acquisition of bifunctionality of valyl-tRNA synthetase predates the divergence of these two organisms.
Assuntos
Arabidopsis/enzimologia , Arabidopsis/genética , Genes de Plantas , Treonina-tRNA Ligase/genética , Valina-tRNA Ligase/genética , Sequência de Aminoácidos , Sequência de Bases , Clonagem Molecular , Códon de Iniciação/genética , Sequência Conservada , Citosol/enzimologia , Primers do DNA/genética , DNA Complementar/genética , DNA de Plantas/genética , Mitocôndrias/enzimologia , Dados de Sequência Molecular , Plantas Geneticamente Modificadas , Plantas Tóxicas , Proteínas Recombinantes de Fusão/genética , Saccharomyces cerevisiae/enzimologia , Saccharomyces cerevisiae/genética , Homologia de Sequência de Aminoácidos , Nicotiana , TransfecçãoRESUMO
One-fifth of the tRNAs used in plant mitochondrial translation is coded for by chloroplast-derived tRNA genes. To understand how aminoacyl-tRNA synthetases have adapted to the presence of these tRNAs in mitochondria, we have cloned an Arabidopsis thaliana cDNA coding for a methionyl-tRNA synthetase. This enzyme was chosen because chloroplast-like elongator tRNAMet genes have been described in several plant species, including A. thaliana. We demonstrate here that the isolated cDNA codes for both the chloroplastic and the mitochondrial methionyl-tRNA synthetase (MetRS). The protein is transported into isolated chloroplasts and mitochondria and is processed to its mature form in both organelles. Transient expression assays using the green fluorescent protein demonstrated that the N-terminal region of the MetRS is sufficient to address the protein to both chloroplasts and mitochondria. Moreover, characterization of MetRS activities from mitochondria and chloroplasts of pea showed that only one MetRS activity exists in each organelle and that both are indistinguishable by their behavior on ion exchange and hydrophobic chromatographies. The high degree of sequence similarity between A. thaliana and Synechocystis MetRS strongly suggests that the A. thaliana MetRS gene described here is of chloroplast origin.