Mitochondrial Reactive Oxygen Species. Contribution to Oxidative Stress and Interorganellar Signaling

Abstract

The inner membrane of a plant mitochondrion contains the mitochondrial electron transport chain (mtETC), consisting of protein complexes that use an energy source-derived reductant to form a proton gradient across the membrane. This proton gradient drives ATP synthesis, a primary mitochondrial function. Ultimately, electrons in the mtETC are transferred through one of two pathways from ubiquinone to a terminal oxidase, where oxygen is reduced to water. Specifically, electrons pass through either the “standard” cytochrome pathway, which includes two sites of proton pumping downstream of the ubiquinone pool, to cytochrome c oxidase (COX), or the alternative pathway, which consists of the cyanide-insensitive alternative oxidase (AOX) and has no proton-pumping sites after the ubiquinone pool (Finnegan et al., 2004). Because the mtETC harbors electrons with sufficient free energy to directly reduce molecular oxygen, it is considered the unavoidable primary source of mitochondrial reactive oxygen species (mtROS) production, a necessary accompaniment to aerobic respiration. Formation of mtROS takes place under normal respiratory conditions but can be enhanced in response to a range of abnormal conditions, including exposure to biotic and abiotic stresses. The marked reactivity of ROS toward biological molecules, including lipids, proteins, and nucleic acids, requires multiple mechanisms for keeping mtROS levels under control, including pathways that attenuate mtROS formation in response to imposed stresses and protective, antioxidant enzyme systems. However, when mtROS formation exceeds normal levels despite the operation of these protective mechanisms, there are downstream consequences for the cell, including altered gene expression and even programmed cell death (PCD). Not simply agents of damage, mtROS also play roles in the signaling required to bring about these changes. The cell ultimately must strike a balance between the level of ROS required to elicit an appropriate response to a changing condition while at the same time keeping ROS levels sufficiently low to prevent large-scale cellular damage. On the other hand, cells must be able to determine a mtROS-initiated, extreme measure at the cellular level is appropriate for the good of the plant as a whole (e.g. the hypersensitive response to pathogen attack). This article provides an overview of our current understanding of plant mtROS. While much remains to be established, mtROS clearly play important roles in the responses of plants to the variety of environmental conditions they experience on a regular basis, as well as more extreme environmental stresses. The known sites of mtROS production in the mtETC are complexes I and III, where superoxide anion (O2 −) is formed and in turn is reduced by dismutation to H2O2 (Raha and Robinson, 2000; Møller, 2001; Sweetlove and Foyer, 2004). H2O2, a compound of relatively low toxicity, can react with reduced Fe2+ and Cu+ to produce highly toxic hydroxyl radicals and, being uncharged, can also penetrate membranes and leave the mitochondrion (Grene, 2002; Sweetlove and Foyer, 2004). The ubisemiquinone intermediate formed at complexes I and III is the principal electron donor to oxygen, although other complex I sites are also potential donors (Raha and Robinson, 2000; Sweetlove and Foyer, 2004). Thus, the overall reduction level of the mitochondrial ubiquinone pool will be the primary determinate of mtROS output (Sweetlove and Foyer, 2004). The amount of ROS produced by mitochondria and the fraction of total cellular ROS that come from mitochondria are difficult to determine, in part because ROS levels in general are difficult to measure accurately (Veljovic-Jovanovic et al., 2002; Halliwell and Whiteman, 2004). For isolated mitochondria and submitochondrial particles, ROS evolution varies with conditions but falls within the wide range of 0.2 to 30.0 nmol min−1 mg protein−1 for either H2O2 or superoxide (studies compiled in Møller, 2001; Popov et al., 2003). While we are unaware of any quantitative measurements of ROS production by mitochondria in vivo, estimates (Foyer and Noctor, 2003) indicate that the in situ level of mtROS evolution normally will be considerably less than that of chloroplasts or peroxisomes in the light due to the operation of photosynthesis and photorespiration. However, in the dark or in non-green tissues, mitochondria will be a major source of ROS (e.g. Puntarulo et al., 1988). Production of mtROS will increase if the rate of electrons leaving the mtETC through the terminal oxidases is slowed and/or the rate of electron input increases in excess of the ability of the two respiratory pathways to process the electrons, leading to an overreduced ubiquinone pool. This principle has been demonstrated in isolated mitochondria, where rates of H2O2 and superoxide generation show a substrate-dependent increase upon the addition of specific inhibitors of either the cytochrome pathway or AOX (Popov et al.