RESUMO
Nonsense-mediated mRNA decay (NMD) is a conserved mechanism that targets aberrant mRNAs for destruction. NMD has also been found to regulate the expression of large numbers of genes in diverse organisms, although the biological role for this is unclear and few evolutionarily conserved targets have been identified. Expression analyses of three Arabidopsis thaliana lines deficient in NMD reveal that the vast majority of NMD-targeted transcripts are associated with response to pathogens. Congruently, NMD mutants, in which these transcripts are elevated, confer partial resistance to Pseudomonas syringae. These findings suggest a biological rationale for the regulation of gene expression by NMD in plants and suggest that manipulation of NMD could offer a new approach for crop protection. Amongst the few non-pathogen responsive NMD-targeted genes, one potential NMD targeted signal, the evolutionarily conserved upstream open reading frame (CuORF), was found to be hugely over-represented, raising the possibility that this feature could be used to target specific physiological mRNAs for control by NMD.
Assuntos
Arabidopsis/genética , Arabidopsis/metabolismo , Regulação da Expressão Gênica de Plantas , Mutação , Degradação do RNAm Mediada por Códon sem Sentido , RNA Mensageiro/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Transporte/genética , Cicloeximida/farmacologia , Evolução Molecular , Genes de Plantas , Análise de Sequência com Séries de Oligonucleotídeos , Fases de Leitura Aberta , Plantas Geneticamente Modificadas , Proteômica/métodos , Pseudomonas syringae/metabolismo , Estabilidade de RNARESUMO
The gaseous hormone ethylene is perceived by a family of ethylene receptors which interact with the Raf-like kinase CTR1. SlTPR1 encodes a novel TPR (tetratricopeptide repeat) protein from tomato that interacts with the ethylene receptors NR and LeETR1 in yeast two-hybrid and in vitro protein interaction assays. SlTPR1 protein with a GFP fluorescent tag was localized in the plasmalemma and nuclear membrane in Arabidopsis, and SlTPR1-CFP and NR-YFP fusion proteins were co-localized in the plasmalemma and nuclear membrane following co-bombardment of onion cells. Overexpression of SlTPR1 in tomato resulted in ethylene-related pleiotropic effects including reduced stature, delayed and reduced production of inflorescences, abnormal and infertile flowers with degenerate styles and pollen, epinasty, reduced apical dominance, inhibition of abscission, altered leaf morphology, and parthenocarpic fruit. Similar phenotypes were seen in Arabidopsis overexpressing SlTPR1. SlTPR1 overexpression did not increase ethylene production but caused enhanced accumulation of mRNA from the ethylene responsive gene ChitB and the auxin-responsive gene SlSAUR1-like, and reduced expression of the auxin early responsive gene LeIAA9, which is known to be inhibited by ethylene and to be associated with parthenocarpy. Cuttings from the SlTPR1-overexpressors produced fewer adventitious roots and were less responsive to indole butyric acid. It is suggested that SlTPR1 overexpression enhances a subset of ethylene and auxin responses by interacting with specific ethylene receptors. SlTPR1 shares features with human TTC1, which interacts with heterotrimeric G-proteins and Ras, and competes with Raf-1 for Ras binding. Models for SlTPR1 action are proposed involving modulation of ethylene signalling or receptor levels.
Assuntos
Etilenos/metabolismo , Ácidos Indolacéticos/metabolismo , Proteínas de Plantas/metabolismo , Receptores de Superfície Celular/metabolismo , Solanum lycopersicum/crescimento & desenvolvimento , Solanum lycopersicum/metabolismo , Motivos de Aminoácidos , Sequência de Aminoácidos , Sequência de Bases , Regulação da Expressão Gênica no Desenvolvimento , Solanum lycopersicum/química , Solanum lycopersicum/genética , Dados de Sequência Molecular , Proteínas de Plantas/química , Proteínas de Plantas/genética , Ligação Proteica , Receptores de Superfície Celular/química , Receptores de Superfície Celular/genética , Alinhamento de Sequência , Técnicas do Sistema de Duplo-HíbridoRESUMO
Aberrant mRNAs containing premature termination codons (PTCs) have the potential to be translated into truncated proteins, which could act to the detriment of the organism by interfering with normal cellular processes. Eukaryotes have mechanisms of mRNA quality control that identify PTC-containing transcripts and target them for destruction, a process known as nonsense-mediated mRNA decay (NMD). Surprising differences have been reported in the mechanisms of NMD between different organisms. UPF1 and UPF3 are structurally unrelated proteins, which function in the NMD pathway in yeast, mammals, Drosophila and Caenorhabditis elegans. Here we show that NMD in plants requires UPF1, as mRNAs containing PTCs become stabilized in upf1-5 mutants. However, in contrast to NMD in mammals, UPF1-dependent NMD is capable of targeting both spliced and unspliced PTC-containing mRNAs. An allelic series of upf1 mutants exhibits a range of unexpected vegetative and floral abnormalities, including jagged leaves, late flowering, fused flowers and seedling lethality. We also show that mutants in UPF3 share these abnormalities. As both UPF1 and UPF3 are required for NMD, the similar phenotypes of the upf1 and upf3 mutants suggest that NMD regulates a common set of genes required for plant development and survival. Finally, gene silencing by an inverted repeat transgene is impaired in upf1-5 mutants, indicating a connection between UPF1 and RNA interference in plants.