The influence of alpha-amanitin on the NaCl-induced up-regulation of antioxidant enzyme activity in cotton callus tissue.

Liquid suspensions of cotton callus tissue from a NaCl-sensitive cell line and a NaCl-tolerant cell line were subjected to the following treatments: (a) 0 and 150 mM NaCl, respectively (controls); (b) 75 and 250 mM NaCl, respectively; (c) 100 ng ml(-1) alpha-amanitin; or (d) pretreatment for 2 h with 100 ng ml(-1) alpha-amanitin followed by the respective NaCl treatments. The callus tissue was harvested at 0, 0.5, 1, 2, 4, and 8h and analyzed for antioxidant enzyme activity. In the NaCl-tolerant callus, the 250 mM NaCl treatment resulted in transient 2- to 4-fold increases above the control levels in the activities of ascorbate peroxidase, catalase, glutathione reductase, and peroxidase within 1 h after treatment, while superoxide dismutase activity increased 4-fold within 4 h. This rapid increase suggests that the up-regulation of antioxidant capacity is an early response to NaCl stress and perhaps provides protection against oxidative damage until other acclimating mechanisms can be invoked. In the control callus, peroxidase activity remained unchanged, and significant increases in the other enzymes were not observed until 8 h after treatment with 75mM NaCl. Pre-treatment with alpha-amanitin prior to the NaCl treatment completely inhibited the NaCl-induced increase in the activities of all five enzymes in both cell lines. This data supports the conclusion that the NaCl-induced up-regulation of antioxidant enzyme activity in cotton callus tissue is transcriptionally regulated, proceeding via a de novo synthesis of poly(A)+RNA and is not due to the translation of existing transcripts or the mobilization of existing enzyme pools. In addition, the results suggest that it is not only the up-regulation of antioxidant activity that bestows a degree of tolerance to environmental stress, but also the speed with which this response occurs.

The relationship between yield and the antioxidant defense system in tomatoes grown under heat stress.

Four putative heat-tolerant tomato (Lycopersicum esculentum) cultivars (Tamasabro, Heat Wave, LHT-24, and Solar Set) and one putative heat-sensitive tomato cultivar (Floradade) were grown in the field under non-stress (average daily temperature of 26 degrees C) and heat-stress (average daily temperature of 34 degrees C) conditions. At anthesis, approximately five weeks after being transplanted to the field, leaf samples were collected for antioxidant analyses. Yield was determined by harvesting ripe fruit seven weeks after the collection of leaf samples. Heat stress resulted in a 79.1% decrease in yield for the heat-sensitive Floradade, while the fruit yield in the heat-tolerant cultivars Heat Wave, LHT-24, Solar Set, and Tamasabro was reduced 51.5%, 22.1%, 43.8%, and 34.8% respectively. When grown under heat stress, antioxidant activities were also greater in the heat-tolerant cultivars. Superoxide dismutase (SOD) activity increased up to 9-fold in the heat-tolerant cultivars but decreased 83.1% in the heat-sensitive Floradade. Catalase, peroxidase, and ascorbate peroxidase activity increased significantly in all cultivars. Only Heat Wave showed a significant increase in glutathione reductase in response to heat stress but all heat-tolerant cultivars exhibited significantly lower oxidized ascorbate/reduced ascorbate ratios, greater reduced glutathione/oxidized glutathione rations, and greater alpha-tocopherol concentrations compared to the heat-sensitive cultivar Floridade. These data indicate that the more heat-tolerant cultivars had an enhanced capacity for scavenging active oxygen species and a more active ascorbate-glutathione cycle and suggest a strong correlation between the ability to up-regulate the antioxidant defense system and the ability of tomatoes to produce greater yields when grown under heat stress.

Antioxidant Response to NaCl Stress in a Control and an NaCl-Tolerant Cotton Cell Line Grown in the Presence of Paraquat, Buthionine Sulfoximine, and Exogenous Glutathione.

A cotton (Gossypium hirsutum L.) control and NaCl-tolerant cell line (cv Coker 312) were grown on media with or without NaCl in the presence or absence of paraquat, buthionine sulfoximine, and oxidized glutathione. On medium with 150 mM NaCl the NaCl-tolerant cell line exhibited no reduction in growth, whereas a 96% reduction was observed in the control line. The NaCl-tolerant cell line that was grown on 150 mM NaCl exhibited significantly greater catalase (341%), peroxidase (319%), glutathione reductase (287%), ascorbate peroxidase (450%), [gamma]-glutamylcysteine synthetase (224%), and glutathione S-transferase (500%) activities than the intolerant control. The NaCl-tolerant cell line had a significantly lower dehydroascorbic acid/ascorbic acid ratio. Paraquat reduced growth by 20 and 53.7%, respectively, in the NaCl-tolerant and control cell line. The NaCl-tolerant cell line also showed a slight tolerance to buthionine sulfoximine. In the buthionine sulfoximine experiments reduced glutathione restored growth in both cell lines, whereas oxidized glutathione restored growth only in the NaCl-tolerant cell line. These data indicate that the NaCl-tolerant cell line exhibited a cross-tolerance to a variety of stress variables and had a more active ascorbate-glutathione cycle.

The effects of NaCl on antioxidant enzyme activities in callus tissue of salt-tolerant and salt-sensitive cotton cultivars (Gossypium hirsutum L.).

To determine NaCl effects on callus growth and antioxidant activity, callus of a salt-tolerant and a salt-sensitive cultivar of cotton was grown on media amended with 0, 75, and 150 mM NaCl. Callus of the salt-tolerant cultivar, Acala 1517-8 8, grown at 150 mM NaCl, showed significant increases in superoxide dismutase, catalase, ascorbate peroxidase, peroxidase and glutathione reductase activities compared to callus tissue grown at 0 mM NaCl. In contrast, callus tissue of the salt-sensitive cultivar, Deltapine 50, grown at 0, 75, and 150 mM NaCl, showed no difference in the activities of these enzymes. At the 150 mM NaCl treatment, peroxidase was the only antioxidant enzyme from Deltapine 50 with an activity as high as that observed in Acala 1517-88. The NaCl-induced increase in the activity of these enzymes in Acala 1517-88 indicates that callus tissue from the more salt-tolerant cultivar has a higher capacity for scavenging and dismutating superoxide, an increased ability to decompose H2O2, and a more active ascorbate-glutathione cycle when grown on media amended with NaCl.

Carbohydrate partitioning and the capacity of apparent nitrogen fixation of soybean plants grown outdoors.

Patterns of leaf carbohydrate partitioning and nodule activity in soybean plants grown under natural conditions and the irradiance level required to produce sufficient carbohydrate to obtain maximum rates of apparent N(2)-fixation (acetylene reduction) were measured. Soybean plants, grown outdoors, maintained constant levels of leaf soluble sugars while leaf starch pools varied diurnally. When root temperature was kept at 25 degrees C and shoot temperature was allowed to vary with ambient temperature, the plants maintained constant rates of apparent N(2)-fixation and root+nodule respiration. Results from a second experiment, in which the entire plant was kept at 25 degrees C, were similar to those of the first experiment. Shoot carbon exchange rate of plants from the second experiment was light saturated at photosynthetic photon flux densities between 400 and 600 micromoles per square meter per second. When plants were subjected to an extended 40-hour dark period to deplete carbohydrate reserves, apparent N(2)-fixation was unaffected during the first 10 hours of darkness, decreased rapidly between 10 and 16 hours, and plateaued at one-third the initial level thereafter. After the extended dark period, plants were exposed to photosynthetic photon flux density from 200 to 1000 micromoles per square meter per second for 10 hours. Photosynthetic photon flux densities of 200 micromoles per square meter per second and greater resulted in maximum leaf soluble sugar content and nodule activity. Leaf starch content increased with irradiance levels up to 600 micromoles per square meter per second with no further increase at higher irradiance levels. Results presented here indicate that maximum nodule activity occurs at irradiance levels that do not saturate the plant's photosynthetic apparatus. This response would allow for maximum N(2)-fixation to occur in a nodulated legume during periods of inclement weather.