RESUMO
Jasmonates are well documented for their ability to modulate the expression of plant genes and to influence specific aspects of disease/pest resistance traits. We and others have been studying the synthesis of sesquiterpene phytoalexins in elicitor/pathogen-challenged plants and have sought to determine if methyl jasmonate (MeJA) could substitute for fungal elicitors in the induction of capsidiol accumulation by tobacco cell cultures. The current results demonstrate that MeJA does in fact induce phytoalexin accumulation, but with a much more delayed induction time course than elicitor. While elicitor treatment induced strong but transient changes in key enzymes of sesquiterpene biosynthesis, sesquiterpene cyclase, and aristolochene/deoxy-capsidiol hydroxylase, MeJA did not. Instead, MeJA caused a protracted induction of cyclase activity and only a low level of hydroxylase activity. MeJA induced the expression of at least two sesquiterpene cyclase genes, including one that had not been observed previously in elicitor-induced mRNA populations. Only a small portion of the total sesquiterpene cyclase mRNA induced by MeJA was associated with polysomal RNA, suggesting that the MeJA treatment imposed both transcriptional and posttranscriptional regulation in tobacco cells. These results are not consistent with MeJA playing a role in orchestrating defense responses in elicitor-treated tobacco cells, but do provide evidence that MeJA induces a subset of genes coding for the biosynthesis of sesquiterpene phytoalexins.
Assuntos
Acetatos/farmacologia , Ciclopentanos/farmacologia , Nicotiana/metabolismo , Plantas Tóxicas , Sesquiterpenos/metabolismo , Sequência de Aminoácidos , Sequência de Bases , Carbono-Carbono Liases/genética , Células Cultivadas , Celulase/farmacologia , Primers do DNA/genética , DNA de Plantas/genética , Proteínas Fúngicas/farmacologia , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Genes de Plantas , Dados de Sequência Molecular , Oxilipinas , Reguladores de Crescimento de Plantas/farmacologia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , RNA de Plantas/genética , RNA de Plantas/metabolismo , Homologia de Sequência de Aminoácidos , Homologia de Sequência do Ácido Nucleico , Nicotiana/efeitos dos fármacos , Nicotiana/genéticaRESUMO
Legumes obtain a substantial portion of their nitrogen (N) from symbiotic N2 fixation in root nodules. The glutamine synthetase (GS, EC 6.3.1.2)/glutamate synthase (GOGAT) cycle is responsible for the initial N assimilation. This report describes the analysis of a transgenic alfalfa (Medicago sativa L.) line containing an antisense NADH-GOGAT (EC 1.4.1.14) under the control of the nodule-enhanced aspartate amino-transferase (AAT-2) promoter. In one transgenic line, NADH-GOGAT enzyme activity was reduced to approximately 50%, with a corresponding reduction in protein and mRNA. The transcript abundance for cytosolic GS, ferredoxin-dependent GOGAT (EC 1.4.7.1), AAT-2 (EC 2.6.1.1), asparagine synthase (EC 6.3.5.4), and phosphoenolpyruvate carboxylase (PEPC, EC 4.1.1.31) were unaffected, as were enzyme activities for AAT, PEPC and GS. Antisense NADH-GOGAT plants grown under symbiotic conditions were moderately chlorotic and reduced in growth and N content, even though symbiotic N2 fixation was not significantly reduced. The addition of nitrate relieved the chlorosis and restored growth and N content. Surprisingly, the antisense NADH-GOGAT plants were male sterile resulting from inviable pollen. A reduction in NADH-GOGAT enzyme activity and transcript abundance in the antisense plants was measured during the early stages of flower development. Inheritance of the transgene was stable and resulted in progeny with a range of NADH-GOGAT activity. These data indicate that NADH-GOGAT plays a critical role in the assimilation of symbiotically fixed N and during pollen development.
Assuntos
Elementos Antissenso (Genética) , Glutamato Sintase/metabolismo , Medicago sativa/enzimologia , NAD/metabolismo , Transformação Genética , Transgenes , Glutamato Sintase/genética , Medicago sativa/genética , Raízes de Plantas/enzimologiaRESUMO
Development of root nodules, specifically induction of cortical cell division for nodule initiation, requires expression of specific genes in the host and microsymbiont. A full-length cDNA clone and the corresponding genomic clone encoding a MAP (mitogen-activated protein) kinase homolog were isolated from alfalfa (Medicago sativa). The genomic clone, TDY1, encodes a 68.9-kDa protein with 47.7% identity to MMK4, a previously characterized MAP kinase homolog from alfalfa. TDY1 is unique among the known plant MAP kinases, primarily due to a 230 amino acid C-terminal domain. The putative activation motif, Thr-Asp-Tyr (TDY), also differs from the previously reported Thr-Glu-Tyr (TEY) motif in plant MAP kinases. TDY1 messages were found predominantly in root nodules, roots, and root tips. Transgenic alfalfa and Medicago truncatula containing a chimeric gene consisting of 1.8 kbp of 5' flanking sequence of the TDY1 gene fused to the beta-glucuronidase (GUS) coding sequence exhibited GUS expression primarily in the nodule parenchyma, meristem, and vascular bundles, root tips, and root vascular bundles. Stem internodes stained intensely in cortical parenchyma, cambial cells, and primary xylem. GUS activity was observed in leaf mesophyll surrounding areas of mechanical wounding and pathogen invasion. The promoter was also active in root tips and apical meristems of transgenic tobacco. Expression patterns suggest a possible role for TDY1 in initiation and development of nodules and roots, and in localized responses to wounding.
