RESUMEN
Reactive oxygen species and nitric oxide (NOâ¢) concomitantly play essential roles in guard cell signaling. Studies using catalase mutants have revealed that the inducible and constitutive elevations of intracellular hydrogen peroxide (H2O2) have different roles: only the inducible H2O2 production transduces the abscisic acid (ABA) signal leading stomatal closure. However, the involvement of inducible or constitutive NO⢠productions, if exists, in this process remains unknown. We studied H2O2 and NO⢠mobilization in guard cells of catalase mutants. Constitutive H2O2 level was higher in the mutants than that in wild type, but constitutive NO⢠level was not different among lines. Induced NO⢠and H2O2 levels elicited by ABA showed a high correlation with each other in all lines. Furthermore, NO⢠levels increased by exogenous H2O2 also showed a high correlation with stomatal aperture size. Our results demonstrate that ABA-induced intracellular H2O2 accumulation triggers NO⢠production leading stomatal closure. ABBREVIATIONS: ABA: abscisic acid; CAT: catalase; cGMP: cyclic guanosine monophosphate; DAF-2DA: 4,5-diaminofluorescein-2 diacetate; H2DCF-DA: 2',7'-dichlorodihydrofluorescein diacetate; MeJA: methyljasmonate; NOS: nitric oxide synthetase; NR: nitrate reductase; POX: peroxidase; ROS: reactive oxygen species; SNAP: S-nitroso-N-acetyl-DL-penicillamine; SNP: sodium nitroprusside; NOX: NADP(H) oxidase.
Asunto(s)
Ácido Abscísico/farmacología , Peróxido de Hidrógeno/farmacología , Espacio Intracelular/metabolismo , Óxido Nítrico/metabolismo , Estomas de Plantas/efectos de los fármacos , Estomas de Plantas/metabolismo , Transducción de Señal/genética , Ácido Abscísico/metabolismo , Arabidopsis , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Catalasa/genética , Catalasa/metabolismo , GMP Cíclico/metabolismo , Peróxido de Hidrógeno/metabolismo , Nitroprusiato/farmacología , Plantas Modificadas GenéticamenteRESUMEN
Arsenic causes physiological and structural disorders in plants. Proline is accumulated as a compatible solute in plants under various stress conditions and mitigates stresses. Here, we investigated the effects of exogenous proline on tobacco Bright Yellow-2 (BY-2) cultured cells under [Formula: see text] stress. Arsenate did not inhibit BY-2 cell growth at 40 and 50 µM but did it at 60 µM. Proline at 0.5 to 10 mM did not affect the cell growth but delayed it at 20 mM. At 40 µM [Formula: see text], neither 0.5 mM nor 1 mM proline affected the cell growth but 10 mM proline inhibited it. In the presence of [Formula: see text], 10 mM proline increased the number of Evans Blue-stained (dead) cells and decreased the number of total cells. Together, our results suggest that exogenous proline does not alleviate arsenate toxicity but enhances the sensitivity of BY-2 cells to arsenate.
Asunto(s)
Arseniatos/toxicidad , Contaminantes Ambientales/toxicidad , Nicotiana/citología , Prolina/farmacología , Recuento de Células , Muerte Celular/efectos de los fármacos , Línea Celular , Proliferación Celular/efectos de los fármacos , Sinergismo FarmacológicoRESUMEN
Methyl jasmonate (MeJA)-induced stomatal closure is accompanied by the accumulation of hydrogen peroxide (H2O2) in guard cells. In this study, we investigated the roles of catalases (CATs) in MeJA-induced stomatal closure using cat mutants cat2, cat3-1 and cat1 cat3, and the CAT inhibitor, 3-aminotriazole (AT). When assessed with 2',7'-dichlorodihydrofluorescein, the reduction of catalase activity by means of mutations and the inhibitor accumulated higher basal levels of H2O2 in guard cells whereas they did not affect stomatal aperture in the absence of MeJA. In contrast, the cat mutations and the treatment with AT potentiated MeJA-induced stomatal closure and MeJA-induced H2O2 production. These results indicate that CATs negatively regulate H2O2 accumulation in guard cells and suggest that inducible H2O2 production rather than constitutive elevation modulates stomatal apertures in Arabidopsis.
Asunto(s)
Acetatos/metabolismo , Proteínas de Arabidopsis/metabolismo , Catalasa/metabolismo , Ciclopentanos/metabolismo , Oxilipinas/metabolismo , Estomas de Plantas/citología , Estomas de Plantas/enzimología , Transducción de Señal , Acetatos/farmacología , Amitrol (Herbicida)/farmacología , Proteínas de Arabidopsis/genética , Señalización del Calcio/efectos de los fármacos , Catalasa/antagonistas & inhibidores , Catalasa/genética , Ciclopentanos/farmacología , Inhibidores Enzimáticos/farmacología , Mutación/genética , Oxilipinas/farmacología , Estomas de Plantas/efectos de los fármacos , Estomas de Plantas/fisiología , Especies Reactivas de Oxígeno/metabolismo , Transducción de Señal/efectos de los fármacosRESUMEN
We investigated the roles of catalase (CAT) in abscisic acid (ABA)-induced stomatal closure using a cat2 mutant and an inhibitor of CAT, 3-aminotriazole (AT). Constitutive reactive oxygen species (ROS) accumulation due to the CAT2 mutation and AT treatment did not affect stomatal aperture in the absence of ABA, whereas ABA-induced stomatal closure, ROS production, and [Ca(2+)](cyt) oscillation were enhanced.
Asunto(s)
Ácido Abscísico/metabolismo , Arabidopsis/citología , Arabidopsis/enzimología , Catalasa/metabolismo , Transducción de Señal , Amitrol (Herbicida)/farmacología , Arabidopsis/efectos de los fármacos , Catalasa/antagonistas & inhibidores , Catalasa/genética , Inhibidores Enzimáticos/farmacología , Mutación , Estomas de Plantas/citología , Estomas de Plantas/efectos de los fármacos , Estomas de Plantas/enzimología , Transducción de Señal/efectos de los fármacosRESUMEN
Reactive oxygen species (ROS), including hydrogen peroxide (H(2)O(2)), are among the important second messengers in abscisic acid (ABA) signaling in guard cells. In this study, to investigate specific roles of H(2)O(2) in ABA signaling in guard cells, we examined the effects of mutations in the guard cell-expressed catalase (CAT) genes, CAT1 and CAT3, and of the CAT inhibitor 3-aminotriazole (AT) on stomatal movement. The cat3 and cat1 cat3 mutations significantly reduced CAT activities, leading to higher basal level of H(2)O(2) in guard cells, when assessed by 2',7'-dichlorodihydrofluorescein, whereas they did not affect stomatal aperture size under non-stressed condition. In addition, AT-treatment at concentrations that abolish CAT activities, showed trivial affect on stomatal aperture size, while basal H(2)O(2) level increased extensively. In contrast, cat mutations and AT-treatment potentiated ABA-induced stomatal closure. Inducible ROS production triggered by ABA was observed in these mutants and wild type as well as in AT-treated guard cells. These results suggest that ABA-inducible cytosolic H(2)O(2) elevation functions in ABA-induced stomatal closure, while constitutive increase of H(2)O(2) do not cause stomatal closure.
Asunto(s)
Ácido Abscísico/farmacología , Amitrol (Herbicida)/farmacología , Arabidopsis/fisiología , Catalasa/metabolismo , Inhibidores Enzimáticos/farmacología , Peróxido de Hidrógeno/metabolismo , Arabidopsis/efectos de los fármacos , Arabidopsis/enzimología , Proteínas de Arabidopsis/efectos de los fármacos , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Catalasa/efectos de los fármacos , Catalasa/genética , Citosol , Expresión Génica , Regulación Enzimológica de la Expresión Génica , Regulación de la Expresión Génica de las Plantas , Mutagénesis Insercional , Hojas de la Planta/efectos de los fármacos , Hojas de la Planta/enzimología , Hojas de la Planta/fisiología , Estomas de Plantas/efectos de los fármacos , Protoplastos , ARN de Planta/genética , Especies Reactivas de Oxígeno/metabolismo , Transducción de Señal/efectos de los fármacosRESUMEN
Efficient detoxification of the reactive oxygen species, nitric oxide (NO) and methylglyoxal (MG), provides protection against NaCl-induced damage in plants. To elucidate the protective mechanisms of proline and glycinebetaine (betaine) against NaCl stress, intracellular levels of hydrogen peroxide (H(2)O(2)), superoxide (O(2)(-)), NO, and MG were investigated in tobacco Bright Yellow-2 cells. The Levels of H(2)O(2), O(2)(-), NO and MG were higher in the short-term and long-term NaCl-stressed cells than in the non-stressed cells, whereas the O(2)(-) level was higher in the long-term stressed cells. Exogenous proline and betaine decreased the H(2)O(2) level in both the short-term and the long-term NaCl-stressed cells and the MG level in the long-term NaCl-stressed cells, but did not change the O(2)(-) or NO levels. Under salt stress, both proline and betaine increased the transcription levels of glutathione peroxidase, which can contribute to the reduction of H(2)O(2). In conclusion, proline and betaine mitigated salt stress via reduction of H(2)O(2) accumulation during short-term incubation and via reduction of the accumulation of H(2)O(2) and MG during long-term incubation.
Asunto(s)
Betaína/farmacología , Peróxido de Hidrógeno/metabolismo , Nicotiana/citología , Prolina/farmacología , Piruvaldehído/metabolismo , Cloruro de Sodio/efectos adversos , Estrés Fisiológico/efectos de los fármacos , Células Cultivadas , Depuradores de Radicales Libres/farmacología , Regulación de la Expresión Génica de las Plantas/efectos de los fármacos , Espacio Intracelular/efectos de los fármacos , Espacio Intracelular/metabolismo , Óxido Nítrico/metabolismo , Superóxidos/metabolismo , Factores de Tiempo , Nicotiana/efectos de los fármacos , Nicotiana/metabolismo , Nicotiana/fisiologíaRESUMEN
Cadmium (Cd) stress significantly decreased membrane integrity and impaired the ascorbate (ASC)-glutathione (GSH) cycle in tobacco Bright Yellow-2 cells. Exogenous application of proline and glycinebetaine (betaine) significantly restored the membrane integrity and increased the activities of ASC-GSH cycle enzymes under Cd stress without maintenance of the rich ASC or GSH pools. Moreover, proline offered more efficient protection against Cd stress than betaine.
Asunto(s)
Ácido Ascórbico/metabolismo , Betaína/farmacología , Cadmio/toxicidad , Glutatión/metabolismo , Nicotiana/efectos de los fármacos , Nicotiana/enzimología , Prolina/farmacología , Línea Celular , Membrana Celular/efectos de los fármacos , Membrana Celular/metabolismo , Glutatión Transferasa/metabolismo , Peróxido de Hidrógeno/metabolismo , Espacio Intracelular/efectos de los fármacos , Espacio Intracelular/metabolismo , Estrés Fisiológico/efectos de los fármacos , Nicotiana/citología , Nicotiana/metabolismoRESUMEN
Environmental stress, including heavy metal stress, can cause oxidative damage to plants. Up-regulation of the antioxidant defense system induced by proline and glycinebetaine (betaine) alleviates the damaging effects of oxidative stress in plants. Here, we investigated the protective effects of exogenously applied proline and betaine on growth, accumulation of proline and betaine, lipid peroxidation and activity of antioxidant enzymes in cultured tobacco Bright Yellow-2 (BY-2) cells exposed to cadmium (Cd) stress. Cadmium stress (at 100 microM Cd) caused a significant inhibition of the growth of BY-2 cells, and both proline and betaine significantly mitigated this inhibition. In addition, the mitigating effect of proline was more pronounced than that of betaine. Cadmium stress leads to an accumulation of Cd and endogenous proline in cultured cells, increased lipid peroxidation and peroxidase (POX) activity, and decreased activity of superoxide dismutase (SOD) and catalase (CAT). Exogenous application of proline resulted in a decrease in lipid peroxidation and an increase in SOD and CAT activities without reducing Cd contents under Cd stress, while application of betaine resulted in a decrease in lipid peroxidation and an increase in CAT activity with reducing Cd accumulation. Furthermore, exogenous proline and betaine intensified the accumulation of proline and betaine in Cd-stressed BY-2 cells, respectively. The present study suggests that proline and betaine confer tolerance to Cd stress in tobacco BY-2 cells by different mechanisms.