Zoltán Takács

Prolonged dark period modulates the oxidative burst and enzymatic antioxidant systems in the leaves of salicylic acid-treated tomato

Salicylic acid (SA) is an important plant growth regulator playing a role in the hypersensitive reaction (HR) and the induction of systemic acquired resistance. Since the SA-mediated signalling pathways and the formation of reactive oxygen species (ROS) are light-dependent, the time- and concentration-specific induction of oxidative stress was investigated in leaves of tomato plants kept under light and dark conditions after treatments with 0.1mM and 1mM SA. The application of exogenous SA induced early superoxide- and H2O2 production in the leaves, which was different in the absence or presence of light and showed time- and concentration-dependent changes. 1mM SA, which induced HR-like cell death resulted in two peaks in the H2O2 production in the light but the first, priming peak was not detected in the dark. Unlike 0.1mM SA, 1mM SA application induced NADPH oxidase activity leading to increased superoxide production in the first hours of SA treatments in the light. Moreover, SA treatments inhibited catalase (CAT) activity and caused a transient decline in ascorbate peroxidase (APX), the two main enzymes responsible for H2O2 degradation, which led to a fast H2O2 burst in the light. Their activity as well as the expression of some isoenzymes of SOD and APX increased only from the 12th h in the illuminated samples. The activity of NADPH oxidase and expression SlRBOH1 gene encoding a NADPH oxidase subunit was much lower in the dark. In spite of low CAT and APX activity after SA treatments in the dark, the activation of guaiacol-dependent peroxidase (POD) could partially substitute H2O2 scavenging activity of these enzymes in the dark, which reduced the ROS burst and development of lesion formation in the leaves.

Opposite extremes in ethylene/nitric oxide ratio induce cell death in suspension culture and root apices of tomato exposed to salt stress

The plant hormone ethylene or the gaseous signalling molecule nitric oxide (NO) may enhance salt stress tolerance by maintaining ion homeostasis, first of all K+/Na+ ratio of tissues. Ethylene and NO accumulation increased in the root apices and suspension culture cells of tomato at sublethal salt stress caused by 100 mM NaCl, however, the induction phase of programmed cell death (PCD) was different at lethal salt concentration. The production of ethylene by root apices and the accumulation of NO in the cells of suspension culture did not increase during the initiation of PCD after 250 mM NaCl treatment. Moreover, cells in suspension culture accumulated higher amount of reactive oxygen species which, along with NO deficiency contributed to cell death induction. The absence of ethylene in the apical root segments and the absence of NO accumulation in the cell suspension resulted in similar ion disequilibrium, namely K+/Na+ ratio of 1.41 ± 0.1 and 1.68 ± 0.3 in intact plant tissues and suspension culture cells, respectively that was not tolerated by tomato.

In vivo inhibition of polyamine oxidase by a spermine analogue, MDL-72527, in tomato exposed to sublethal and lethal salt stress

The spermine analogue N1,N4-bis-(2,3-butadienyl)-1,4-butanediamine (MDL-72527), an effective inhibitor of polyamine oxidases (PAOs), triggers a systemic response in tomato (Solanum lycopersicum L.) exposed to sublethal (100mM) and lethal (250mM) NaCl concentrations. The accumulation of free polyamines (PAs), the terminal oxidation of PAs by diamine oxidases (DAOs) and PAOs, and the production of H2O2 by PA oxidases depends on the intensity of salt stress. Spermidine and spermine content increased significantly under sublethal salt concentrations, but remained low under lethal salt stress. Along with increased expression of the selected SlDAO1 and SlPAO1 genes in the leaves and roots, respectively, DAO and PAO activities and their product, H2O2, increased and initiated cell death by irreversible loss of electrolytes at 250mM NaCl. MDL-72527 significantly increased spermine, spermidine and/or putrescine contents as a result of reduced activity of PA oxidases; furthermore, it inhibited H2O2 and NO production during salt treatment. These results indicate that PAO contributed to H2O2 and NO production under salt stress, and the terminal activities of DAO and PAO play a role in cell death induction at 250mM NaCl. However, the inhibition of PAO by MDL-72527 does not increase the salt tolerance of plants, since electrolyte leakage increased significantly in the presence of the inhibitor.

H2O2 homeostasis in wild-type and ethylene-insensitive Never ripe tomato in response to salicylic acid treatment in normal photoperiod and in prolonged darkness

Ethylene proved to be an important modulator of salicylic acid (SA) signalling pathway. Since SA may regulate both the production and scavenging of hydrogen peroxide (H2O2), which show light-dependency, the aim of this study was to compare H2O2 metabolism in the leaves of SA-treated wild-type (WT) tomato (Solanum lycopersicum L. cv. Ailsa Craig) and in ethylene receptor Never-ripe (Nr) mutants grown in normal photoperiod or in prolonged darkness. H2O2 accumulation was higher in the WT than in the mutants in normal photoperiod after 1 mM SA treatment, while Nr leaves contained more H2O2 after light deprivation. The expression of certain superoxide dismutase (SOD) genes and activity of the enzyme followed the same tendency as H2O2, which was scavenged by different enzymes in the two genotypes. Catalase (CAT, EC 1.11.1.6) activity was inhibited by SA in WT, while the mutants maintained enhanced enzyme activity in the dark. Thus, in WT, CAT inhibition was the major component of the H2O2 accumulation elicited by 1 mM SA in a normal photoperiod, since the expression and/or activity of ascorbate (APX, EC 1.11.1.11) and guaiacol peroxidases (POD, EC 1.11.1.7) were induced in the leaves. The absence of APX and POD activation in mutant plants suggests that the regulation of these enzymes by SA needs functional ethylene signalling. While the block of ethylene perception in Nr mutants was overwritten in the transcription and activity of certain SOD and CAT isoenzymes during prolonged darkness, the low APX and POD activities led to H2O2 accumulation in these tissues.

Regulation of the key antioxidant enzymes by developmental processes and environmental stresses in the dark

The presence of solar radiation is one of the most important environmental factors, which is required for the optimal growth and development of plants. The absence of it (e.g. in the night period or artificially prolonged darkness) can alter the light-dependent signalling and regulation pathways and may induce new defence responses. Antioxidant enzymes as components of the plant defence system play a crucial role in the detoxification of reactive oxygen species (ROS) induced by several stressors; however, their regulation can be different in the light or in the dark. In this review we summarize the current knowledge about the physiological and molecular aspects of dark-modulated key antioxidant enzymes (superoxide dismutase, catalase, and ascorbate peroxidase) in different plant species and discuss their roles in different developmental processes (seedling growth and development or senescence) and in responses to environmental stresses (cold, chilling, heat, and biotic stress). Moreover, the hormonal regulation of respective gene transcription and the changes in activity of various isoenzymes at subcellular level are also summarized. Based on this knowledge, modification of these antioxidant enzymes may be used to increase the yield and stress tolerance of cultivated plants in the changing environment.