RESUMO
5-Enolpyruvylshikimate-3-phosphate synthase (EPSPS), the target enzyme for glyphosate inhibition, catalyzes an essential step in the shikimate pathway for aromatic amino acid biosynthesis. The full-length cDNA of 1,751 nucleotides (CaEPSPS, Genbank accession number: EU698030) from Convolvulus arvensis was cloned and characterized. The CaEPSPS encodes a polypeptide of 520 amino acids with a calculated molecular weight of 55.5 kDa and an isoelectric point of 7.05. The results of homology analysis revealed that CaEPSPS showed highly homologous with EPSPS proteins from other plant species. Tissue expression pattern analysis indicated that CaEPSPS was constitutively expressed in stems, leaves and roots, with lower expression in roots. CaEPSPS expression level could increase significantly with glyphosate treatment, and reached its maximum at 24 h after glyphosate application. We fused CaEPSPS to the CaMV 35S promoter and introduced the chimeric gene into Arabidopsis. The resultant expression of CaEPSPS in transgenic Arabidopsis plants exhibited enhanced tolerance to glyphosate in comparison with control.
Assuntos
3-Fosfoshikimato 1-Carboxiviniltransferase/genética , Clonagem Molecular , Convolvulus/genética , Regulação da Expressão Gênica de Plantas , 3-Fosfoshikimato 1-Carboxiviniltransferase/metabolismo , Sequência de Aminoácidos , Sequência de Bases , Convolvulus/classificação , Convolvulus/metabolismo , DNA Complementar/química , DNA Complementar/genética , Resistência a Medicamentos/genética , Ativação Enzimática , Perfilação da Expressão Gênica , Glicina/análogos & derivados , Glicina/farmacologia , Herbicidas/farmacologia , Dados de Sequência Molecular , Fenótipo , Filogenia , Plantas Geneticamente Modificadas , Regiões Promotoras Genéticas , Alinhamento de Sequência , GlifosatoRESUMO
Anthocyanins contribute greatly to the organoleptic and biochemical properties of grapes and wines. Although there are broadly documented factors involved in grape anthocyanin synthesis, the present work focused on fungal endophytes and their possible role in grape coloration. Our results showed that exposure to endophytic fungi within a dual culture system differentially affected total anthocyanin concentrations and PAL activities in grape cells. Grape cells dual cultured with fungal strains XH-2, R2-21 and B2-17 showed significant differences of their anthocyanin concentrations were subjected to further analysis of their anthocyanidin compositions. Compared to the no-fungus controls, grape cells exposed to fungal strains XH-2 and R2-21 exhibited quantitative promotion of their total anthocyanidin concentrations by 74% and 28%, respectively, whereas treatment with the fungus B2-17 reduced the anthocyanidin content by 19%. A total of 14 species of anthocyanidins were detected from the grape cells in these experiments. Most interestingly, exposure to any of these fungal strains differentially modified the compositional patterns of grape cellular anthocyanidins. The obvious upregulation of the transcription of VvMYB in grape cells treated with fungal strains XH-2 and R2-21 implies that the increased anthocyanin levels in these grape cells may be due to the activated transcriptional factors. In addition, the exposure of grape cells to extracts of these fungi initiated similar responses of anthocyanin contents and PAL activities to exposure to the living fungi and appeared obvious dosage effects. The influence of fungal endophytes on the coloration of grape berries was also examined in this study.