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Vol. 16 No. 3 - July 2010

Dual Nature of Reactive Oxygen Species; An Ally or Adversary for Plant

By: 1P. Tripathi*, 2S. Dwivedi*, 3D. Ckakraborty*, 4P.K. Trivedi*, 5R.D.Tripathi*

What are Reactive oxygen species (ROS)?

Palaeo-climatically, the introduction of molecular oxygen (O2) into our atmosphere by O2-evolving photosynthetic organisms 2.7 billion years ago, reactive oxygen species (ROS) have been the undesirable companion of aerobic life. Accordingly, the evolution of all aerobic organisms has been dependent on the development of efficient ROS-scavenging mechanisms. ROS are free radicals produced as by-products of oxidation–reduction (REDOX) reactions. ROS are produced as a consequence of electron transport processes in photosynthesis and aerobic respiration. The total reduction of oxygen produces water, however partial reduction produces ROS including superoxide anion (·-), hydrogen peroxide (H2O2) and the hydroxyl radical (OH·). ROS are generated at very high rates in plants. They are produced by organelles with a high oxidising metabolic activity or intense rate of electron flow, e.g. chloroplasts, mitochondria and microbodies, or by the oxidases, a large class of enzymes. Examples of ROS-producing oxidases include the plasma membrane NADPH oxidases, peroxidases, oxalate oxidases and amine oxidases. The mechanism of activation is by the stepwise monovalent reduction of oxygen to form superoxide (O-), hydrogen peroxide (H2O2), hydroxyl radical (OH·) and finally water according to the scheme. The first step in the reduction of oxygen forming superoxide is endothermic but subsequent reductions are exothermic. Superoxide can act as either an oxidant or a reductant; it can oxidise sulphur, ascorbic acid or NADPH; it can reduce cytochrome C and metal ions. A dismutation reaction leading to the formation of hydrogen peroxide and oxygen can occur spontaneously or is catalysed by the enzyme superoxide dismutase. In its protonated form (pKa = 4.8) superoxide forms the perhydroxyl radical (·OOH) which is a powerful oxidant (Gebicki and Bielski, 1981), but its biological relevance is probably minor because of its low concentration at physiological pH.

Reactive oxygen species as an Ally

According to the most of recent studies, a new task for ROS has been recognized in plants: the control and regulation of a group of essential biological processes, such as, hormone signaling, immune defense, growth and development, seed germination and the alleviation of seed dormancy, programmed cell death; and stress acclimation. At low levels, they are beneficial for plant and act as signaling molecules during local and systemic acquired resistance. In addition to sensing the environment and abiotic stresses, ROS also play an important role in plant defense responses to pathogens. They are involved in the hypersensitive response typical of plant-pathogen incompatible interactions. The hypersensitive response (HR), is an example of programmed cell death (PCD) characterized by cell death at the site of infection. One of the earliest events in HR is the rapid accumulation of ROS, which can be directly toxic to the pathogen, but recent evidence suggests that the ROS, in particular H2O2, are the signal molecules that trigger HR and other defense mechanisms such as systemic acquired resistance (SAR) and activation of defense genes. The production of ROS occurs in two distinct phases: an initial, non-specific phase probably originates from an NADPH-dependent oxidase follows within minutes of pathogen addition, and a secondary phase, dependent on recognition of incompatible pathogens by the host begins 1–3 h after the initial burst. The produced O2.- is dismutated by SOD to H2O2 and will activate benzoic acid 2-hydroxylase leading to salicylic acid (SA) accumulation. This rise in SA, in combination with H2O2, is necessary for inducing defense gene expression. The second, longer-lasting burst is involved in activating defense responses, even if thought to be NAD(P)H dependent. H2O2 and O2.- can induce different genes, in combination or separately, thereby giving more flexibility to the ROS signaling function. ROS are also produced in response to many hormones such as auxin, abscisic acid and salicylic acid. It is no surprise therefore that ROS are important not only in sensing and responding to environmental changes, but also in orchestrating plant movement (stomatal closure and tropism) responses and in development. Liam Dolan and his colleagues at John Innes Centre (UK) have demonstrated a role for ROS in the development of root hairs. The production of the ROS signal for root growth is highly ordered and tightly coordinated to ensure the production of a single root hair. Redox signaling events are also involved in plant responses to temperature stresses. Here the ROS appear to play a more direct role in the induction of heat shock proteins, but this does not rule out other indirect mechanisms. It would appear that stress sensors in the photosynthetic and respiratory electron transport chains activate redox-sensitive transcription factors that in turn up-regulate the expression of genes that encode HSPs and related proteins, ROS-scavenging enzymes and factors involved in the amplification of the ROS signal by activation of NADPH oxidases. According to beyond depiction it will be able to predict that at minimum concentration ROS may act as an ally or defender for plant.

Reactive oxygen species as an Adversary

Nevertheless, the effects of ROS are dose dependent and high levels of ROS production lead to a process that is often referred to as 'oxidative stress' on the cell, and call up reflective changes in gene expression. The adverse effects of ROS have been coupled with aging, carcinogenesis and atherosclerosis in humans. These oxygen radicals are highly reactive, forming hydroperoxides with enes and dienes. Furthermore, specific amino acids, such as histidine, methionine, and tryptophan can be oxidized by O2.-. In the cellular environment, O2.- will also cause lipid peroxidation, thereby weakening cell membranes. The following O2 reduction produces hydrogen peroxide (H2O2), a quite long-lasting molecule (1 µs) that can diffuse some distance from its site of production. The biological toxicity of H2O2 through oxidation of SH groups has long been recognized and it can be enhanced in the presence of metal catalysts through Haber-Weiss or Fenton-type reactions. Likewise, O2.- may react with other molecules such as lipid peroxides. It can also react with nitric oxide, leading to the formation of peroxynitrite, a species considered less reactive than peroxides. These reactions depend on concentration and on the preferential scavenging capacity of the cell. The plant may favor formation of one or the other species by preferentially scavenging H2O2 with antioxidants or, in contrast, accumulate H2O2 by activating superoxide dismutase (SOD). In addition, H2O2 accumulation may itself lead to higher ROS production. The H2O2 disruption of photosynthesis, as well as the probable direct role of H2O2 in the activation of an NAD(P)H-dependent oxidase , suggests that H2O2 itself can stimulate ROS accumulation. The various sites of oxygen activation in the plant cell are highly controlled and firmly coupled to avoid release of intermediate products. Under stress situations, it is probable that this control or coupling breaks down and the process "dysfunctions" leaking activated oxygen. This is probably a common occurrence in plants especially when we consider that a plant has minimal mobility and control of its environment. These uncoupling events are not unfavorable that they are short in duration and that the oxygen scavenging systems are able to detoxify the various forms of activated oxygen. If the production of activated oxygen exceeds the plant's capacity to detoxify it, deleterious degenerative reactions occur, the typical symptoms being loss of osmotic responsiveness, wilting, and necrosis. At the sub cellular level, disintegration of membranes and aggregation of proteins are typical symptoms. Under abiotic stress, such as heavy metal stress condition, a large quantity of ROS generates in the cell and degrades chlorophyll, protein, nucleic acid molecules and causes lipid peroxidation. The degree of lipid peroxidation is measured by MDA( manoldialdehyde), the end product of the reaction. Therefore the undesirable product of ROS action in the cell under abiotic stress illustrates the adverse nature of reactive oxygen species.

Efficient management of steady state level of ROS

According to dual nature of ROS, it acts as both a defender and as a destroyer to plant. How this dual role is controlled in plants is a little bit mysterious. Although there has been rapid progress in recent years, there are still many uncertainties and gaps in our understanding of how ROS affect the stress response of plants. Nonetheless, it is obvious that the steady-state level of ROS in cells needs to be strongly regulated. Thus, the plant's dilemma is not how to eliminate the activation of oxygen, but how to control and manage the potential reactions of activated oxygen. Complex systems of scavenging activated oxygen therefore exist in plant cells with complimentary and interdependent strategies. Some components such as the carotenoids prevent the formation of activated oxygen by competing for the energy leaked from the photosystems. Other components are lipid soluble and reside in the membrane bilayer to terminate the lipid peroxidation chain reactions. Still others, ascorbate and glutathione, are aqueous scavenger that detoxify activated oxygen directly or serve to recycle other protective components back to their reduced state. The enzymes that catalyse the synthesis, degradation and recycling of these antioxidants are essential to viability. Consequently they are highly conserved among plants, and exist in multiple forms in different subcellular compartments and different tissues to allow precise regulation. For ROS to be effective in these roles Graham Noctor of the University of Paris, notes that “the production and concentration of ROS requires effective regulation by a powerful antioxidant system”. The plant antioxidative system is continuously processing ROS, by acting as electron donors the antioxidants are themselves oxidised in the process of neutralizing the ROS. The omnipresence of O2 in the environment and the various cellular locations where ROS are produced render oxidant scavengers necessary for plant growth and survival. The capacity for ROS to serve as signals adds to the importance of antioxidants to specifically regulate different ROS in various cellular locations. Plants have several antioxidant enzymes and metabolites located in different plant cell compartments, the main ones being SODs, a family of metalloenzymes catalyzing the dismutation of O2.- to H2O2, catalases (CATs), which are heme proteins that catalyze the removal of H2O2, and the enzymes and metabolites of the ascorbate-glutathione cycle that are involved in the removal of H2O2.The majority of enzymes of the ascorbate-glutathione cycle [ascorbate peroxidase (APX), glutathione reductase (GR), and dehydroascorbate reductase (DHAR)] have been found in chloroplasts, cytosol, mitochondria, and peroxisomes. Catalase and the ascorbate-glutathione cycle are important in H2O2 scavenging. Although their properties and requirements are different, they function effectively in parallel. Catalase does not require reducing power and has a high reaction rate but a low affinity for H2O2, thereby only removing the bulk of H2O2. In contrast, APX requires a reductant (ascorbate) and has a higher affinity for H2O2, allowing for the scavenging of small amounts of H2O2 in more specific locations. Non-enzymatic antioxidant include redox buffers such as glutathione, ascorbate, tocopherol, flavonoids, carotenoids and alkaloids vis-ŕ-vis ROS detoxification. Antioxidant capacity is very much dependent on the severity of the stress as well as the species and its developmental stage (Apel & Hirt, 2004).

A highly dynamic and redundant network of more than 150 genes is implicated in Arabidopsis(a model plant for genomics, proteomics and stress related physiology) for managing the concentration of ROS and also encodes ROS-scavenging and ROS-producing proteins. Recent studies have unraveled some of the key players in the network, but many questions related to its mode of regulation, its protective roles and its modulation of signaling networks that control growth, development and stress response remain unanswered. The hunt for ROS receptors in plants is still open. It has been proposed that plant cells sense ROS by at least three different mechanisms: (i) unidentified receptor proteins; (ii) redox-sensitive transcription factors, such as NPR1 or Heat Shock Factors; and (iii) direct inhibition of phosphatases by ROS. Multiple efforts involving genomics, proteomics and metabolomics and other coming up technologies are likely to provide a better picture of the networks involved in different ROS-related plant processes (http://www.scitopics.com/Reactive_Oxygen_Species_in_Plants.html). Since active oxygen species contribute a regulatory function in plants response and adaptation to both biotic and abiotic stress conditions, new insights into the ROS gene network might also allow the identification of genes that can eventually be exploited to modulate ROS-related plant processes that direct the development of better performing yield plants through detoxification of excess ROS at subcellular level. Therefore it is essential to sustain the balance between the production and the scavenging of activated oxygen that is critical to the maintenance of active growth and metabolism of the plant and overall environmental stress tolerance.

*National Botanical Research Institute(CSIR), Lucknow-226001

E-mail: 1[email protected], 2[email protected], 3[email protected], 4[email protected], 5[email protected]

This article has been reproduced from the archives of EnviroNews - Newsletter of ISEB India.

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