Amount Of Alternative Oxidase Protein Research Articles

Responses of woody plant Dalbergia odorifera treated with glycine betaine to drought and cold stresses: involvement of the alternative oxidase

Woody plants faced multiple abiotic stresses in forest plantation that can influence their growth and development. Dalbergia odorifera T. Chen is a vulnerably endangered tree species, and references about its responses to abiotic stresses are very rare in literature. Furthermore, the mechanisms underlying the abiotic stress tolerance in plants induced by exogenous glycine betaine (GB) remains unclear. Indeed, the alternative oxidase (AOX) is one of the major components of antioxidant enzymatic machinery, and there are no studies that focused on the effects of GB on the amount of AOX protein in plants under drought or cold stresses. Thus, the aim of this study was to investigate the effect of exogenous GB on the phenotype, osmoprotectants, photosynthetic pigments, and antioxidant systems in D. odorifera under cold and drought stresses. The layout of the trial was a factorial experiment in a completely randomized design using two factors including abiotic stress (drought and cold) and GB. Moreover, the principal component analysis based on the measured parameters manifested how the selected ROS scavengers were dispersed throughout the treatments. The results showed huge beneficial impacts of GB on the phenotypic traits of D. odorifera; GB also influences positively the antioxidant machinery, photosynthetic pigments, redox-homeostasis, and water status in D. odorifera seedlings under both stresses. Moreover, exogenous GB affects more the AOX pathway in D. odorifera under cold stress than under drought stress.

Alternative Pathway is Involved in Nitric Oxide-Enhanced Tolerance to Cadmium Stress in Barley Roots.

Alternative pathway (AP) has been widely accepted to be involved in enhancing tolerance to various environmental stresses. In this study, the role of AP in response to cadmium (Cd) stress in two barley varieties, highland barley (Kunlun14) and barley (Ganpi6), was investigated. Results showed that the malondialdehyde (MDA) content and electrolyte leakage (EL) level under Cd stress increased in two barley varieties. The expressions of alternative oxidase (AOX) genes (mainly AOX1a), AP capacity (Valt), and AOX protein amount were clearly induced more in Kunlun14 under Cd stress, and these parameters were further enhanced by applying sodium nitroprussid (SNP, a NO donor). Moreover, H2O2 and O2− contents were raised in the Cd-treated roots of two barley varieties, but they were markedly relieved by exogenous SNP. However, this mitigating effect was aggravated by salicylhydroxamic acid (SHAM, an AOX inhibitor), suggesting that AP contributes to NO-enhanced Cd stress tolerance. Further study demonstrated that the effect of SHAM application on reactive oxygen species (ROS)-related scavenging enzymes and antioxidants was minimal. These observations showed that AP exerts an indispensable function in NO-enhanced Cd stress tolerance in two barley varieties. AP was mainly responsible for regulating the ROS accumulation to maintain the homeostasis of redox state.

Coordinated regulation of photosynthetic and respiratory components is necessary to maintain chloroplast energy balance in varied growth conditions.

Mitochondria have a non-energy-conserving alternative oxidase (AOX) proposed to support photosynthesis, perhaps by promoting energy balance under varying growth conditions. To investigate this, wild-type (WT) Nicotiana tabacum were compared with AOX knockdown and overexpression lines. In addition, the amount of AOX protein in WT plants was compared with that of chloroplast light-harvesting complex II (LHCB2), whose amount is known to respond to chloroplast energy status. With increased growth irradiance, WT leaves maintained higher rates of respiration in the light (RL), but no differences in RL or photosynthesis were seen between the WT and transgenic lines, suggesting that, under non-stress conditions, AOX was not critical for leaf metabolism, regardless of growth irradiance. However, under drought, the AOX amount became an important determinant of RL, which in turn was an important determinant of chloroplast energy balance (measured as photosystem II excitation pressure, EP), and photosynthetic performance. In the WT, the AOX amount increased and the LHCB2 amount decreased with increased growth irradiance or drought severity. These changes in protein amounts correlated strongly, in opposing ways, with growth EP. This suggests that a signal deriving from the photosynthetic electron transport chain status coordinately controls the amounts of AOX and LHCB2, which then both contribute to maintaining chloroplast energy balance, particularly under stress conditions.

Alternative oxidase: a respiratory electron transport chain pathway essential for maintaining photosynthetic performance during drought stress.

Photosynthesis and respiration are the hubs of energy metabolism in plants. Drought strongly perturbs photosynthesis as a result of both diffusive limitations resulting from stomatal closure, and in some cases biochemical limitations that are associated with a reduced abundance of key photosynthetic components. The effects of drought on respiration, particularly respiration in the light (RL ), are less understood. The plant mitochondrial electron transport chain includes a non-energy conserving terminal oxidase called alternative oxidase (AOX). Several studies have shown that drought increases AOX transcript, protein and maximum capacity. Here we review recent studies comparing wild-type (WT) tobacco to transgenic lines with altered AOX protein amount. Specifically during drought, RL was compromised in AOX knockdown plants and enhanced in AOX overexpression plants, compared with WT. Significantly, these differences in RL were accompanied by dramatic differences in photosynthetic performance. Knockdown of AOX increased the susceptibility of photosynthesis to drought-induced biochemical limitations, while overexpression of AOX delayed the development of such biochemical limitations, compared with WT. Overall, the results indicate that AOX is essential to maintaining RL during drought, and that this non-energy conserving respiration maintains photosynthesis during drought by promoting energy balance in the chloroplast. This review also outlines several areas for future research, including the possibility that enhancement of non-energy conserving respiratory electron sinks may be a useful biotechnological approach to increase plant performance during stress.

Plant respiration in a high CO2 world: How will alternative oxidase respond to future atmospheric and climatic conditions?

Wang, J., Cheung, M., Rasooli, L., Amirsadeghi, S. and Vanlerberghe, G. C. 2014. Plant respiration in a high CO2 world: How will alternative oxidase respond to future atmospheric and climatic conditions? Can. J. Plant Sci. 94: 1091–1101. Plant mitochondria contain an alternative oxidase (AOX) that reduces the energy yield of respiration. While respiration and photosynthesis are known to interact, the role of AOX in the light remains poorly understood. This gap in our understanding of leaf metabolism extends to future conditions of high CO2 and climate change. While studies indicate that AOX respiration is quite responsive to growth conditions, few studies have examined AOX respiration at high CO2 and little is known regarding the combined impact of changes in both CO2 and other climatic factors such as temperature and water availability. Given its non-energy conserving nature, a fundamental response by AOX to these future conditions could impact the net carbon gain that results from the combined processes of photosynthesis and respiration. Here, we show that leaf AOX protein amount in Nicotiana tabacum is dependent upon growth irradiance and CO2 level, that AOX is subject to biochemical control by intermediates of photorespiration, and that photosynthesis is impacted in transgenic plants lacking AOX. We also review findings that tobacco AOX respiration is responsive to climatic variables (temperature, water availability), thus providing an excellent experimental system to investigate the interplay between AOX, photosynthesis at high CO2, and climate change.

Response of mitochondrial thioredoxin PsTrxo1, antioxidant enzymes, and respiration to salinity in pea (Pisum sativum L.) leaves.

Mitochondria play an essential role in reactive oxygen species (ROS) signal transduction in plants. Redox regulation is an essential feature of mitochondrial function, with thioredoxin (Trx), involved in disulphide/dithiol interchange, playing a prominent role. To explore the participation of mitochondrial PsTrxo1, Mn-superoxide dismutase (Mn-SOD), peroxiredoxin (PsPrxII F), and alternative oxidase (AOX) under salt stress, their transcriptional and protein levels were analysed in pea plants growing under 150 mM NaCl for a short and a long period. The activities of mitochondrial Mn-SOD and Trx together with the in vivo activities of the alternative pathway (AP) and the cytochrome pathway (CP) were also determined, combined with the characterization of the plant physiological status as well as the mitochondrial oxidative indicators. The analysis of protein and mRNA levels and activities revealed the importance of the post-transcriptional and post-translational regulation of these proteins in the response to salt stress. Increases in AOX protein amount correlated with increases in AP capacity, whereas in vivo AP activity was maintained under salt stress. Similarly, Mn-SOD activity was also maintained. Under all the stress treatments, photosynthesis, stomatal conductance, and CP activity were decreased although the oxidative stress in leaves was only moderate. However, an increase in lipid peroxidation and protein oxidation was found in mitochondria isolated from leaves under the short-term salinity conditions. In addition, an increase in mitochondrial Trx activity was produced in response to the long-term NaCl treatment. The results support a role for PsTrxo1 as a component of the defence system induced by NaCl in pea mitochondria, providing the cell with a mechanism by which it can respond to changing environment protecting mitochondria from oxidative stress together with Mn-SOD, AOX, and PrxII F.

Changes of alternative oxidase activity, capacity and protein content in leaves of Cucumis sativus wild‐type and MSC16 mutant grown under different light intensities

In vitro studies demonstrated that alternative oxidase (AOX) is biochemically regulated by a sulfhydryl-disulfide system, interaction with alpha-ketoacids, ubiquinone pool redox state and protein content among others. However, there is still scarce information about the in vivo regulation of the AOX. Cucumis sativus wild-type (WT) and MSC16 mutant plants were grown under two different light intensities and were used to analyze the relationship between the amount of leaf AOX protein and its in vivo capacity and activity at night and day periods. WT and MSC16 plants presented lower total respiration (V(t)), cytochrome oxidase pathway (COP) activity (v(cyt)) and alternative oxidase pathway (AOP) activity (v(alt)) when grown at low light (LL), although growth light intensity did not change the amount of cytochrome oxidase (COX) nor AOX protein. Changes of v(cyt) related to growing light conditions suggested a substrate availability and energy demand control. On the other hand, the total amount of AOX protein present in the tissue does not play a role in the regulation neither of the capacity nor of the activity of the AOP in vivo. Soluble carbohydrates were directly related to the activity of the AOP. However, although differences in soluble sugar contents mostly regulate the capacity of the AOP at different growth light intensities, additional regulatory mechanisms are necessary to fully explain the observed results.

Influence of Chloroplastic Photo-Oxidative Stress on Mitochondrial Alternative Oxidase Capacity and Respiratory Properties: A Case Study with Arabidopsis yellow variegated 2

Mitochondrial alternative oxidase (AOX), the unique respiratory terminal oxidase in plants, catalyzes the energy-wasteful cyanide (CN)-resistant respiration. Although it has been demonstrated that leaf AOX is up-regulated under high-light (HL) conditions, the in vivo mechanism of AOX up-regulation by light is still unknown. In the present study, we examined whether the photo-oxidative stress in the chloroplast modulates mitochondrial respiratory properties, especially the AOX capacity, using Arabidopsis leaf-variegated mutant yellow variegated 2 (var2) and exposing plants to HL. var2 mutants lack FtsH2 metalloprotease required for the repair of damaged PSII. Indeed, var2-1 suffered from photo-oxidative stress even before the HL treatments. While the activities of tricarboxylic acid cycle enzymes and cytochrome c oxidase in var2-1 were almost identical to those in the wild type, the amount of AOX protein and the CN-resistant respiration rate were higher in var2-1. Real-time PCR analysis revealed that HL treatment induced the expression of some energy-dissipating respiratory genes, including AOX1a, NDB2 and UCP5, more strongly in var2-1. Western blotting using var2-1 leaf extracts specific to green or white sectors, containing functional or non-functional photosynthetic apparatus, respectively, revealed that more AOX protein was induced in the green sectors by the HL treatment. These results indicate that photo-oxidative stress by excess light is involved in the regulation of respiratory gene expression and the modulation of respiratory properties, especially the AOX up-regulation.

Induction of alternative oxidase chain under salt stress conditions

This paper describes the effect of NaCl on the respiration of Citrus cell suspensions namely on the induction of the alternative oxidase. The exposure of two Citrus (cvs. Carvalhal tangor and Valencia late) cell suspensions to 200 or 400 mM NaCl lead to a reduction on cell respiration rates. Under these conditions, the respiration rate decreased less in the presence of KCN indicating a stimulation of the capacity of the alternative oxidase (AOX). In addition, immunoblots showed an increase on the amount of AOX protein. Antibodies raised against the Sauromatum guttatum enzyme recognized the reduced form of the enzyme near the 35 kDa band. The protein accumulation was correlated with the significantly higher AOX capacity observed for cv. Carvalhal tangor.

Regulation of alternative oxidase activity during phosphate deficiency in bean roots (Phaseolus vulgaris).

Cyanide-resistant respiration was studied in mitochondria isolated from the roots of bean plants (Phaseolus vulgaris L. cv. Złota Saxa) grown hydroponically up to 16 days on a phosphate-sufficient (+P, control) or phosphate-deficient (-P) medium. Western blotting indicated that the alternative oxidase (AOX) was present only in its reduced (active) form, both in phosphate-sufficient and phosphate-deficient roots, but in the latter, the amount of AOX protein was greater. Addition of pyruvate to the isolation, washing and reaction media made mitochondria from +P roots cyanide-insensitive, similar to mitochondria from -P roots. The doubled activity of NAD-malic enzyme (NAD-ME) in -P compared with +P root mitochondria may suggest increased pyruvate production in -P mitochondria. Lower cytochrome c oxidase (COX) activity and no uncoupler effect on respiration indicated limited cytochrome chain activity in -P mitochondria. In -P mitochondria, the oxygen uptake decreased and the level of Q reduction increased from 60 to 80%. With no pyruvate present (AOX not fully activated), inhibition of the cytochrome pathway resulted in an increased level of the ratio of reduced ubiquinone (Qr) to total ubiquinone (Qt) (Qr/Qt) in +P mitochondria, but did not change Qr/Qt in -P mitochondria. When pyruvate was present, the kinetics for AOX were similar in mitochondria from -P and +P roots. It is suggested that AOX participation in -P respiration may provide an acclimation to phosphate deficiency. Stabilization of the ubiquinone reduction level by AOX might prevent the harmful effect of an increased formation of reactive oxygen species.

The effect of growth and measurement temperature on the activity of the alternative respiratory pathway

A postulated role of the CN-resistant alternative respiratory pathway in plants is the maintenance of mitochondrial electron transport at low temperatures that would otherwise inhibit the main phosphorylating pathway and prevent the formation of toxic reactive oxygen species. This role is supported by the observation that alternative oxidase protein levels often increase when plants are subjected to growth at low temperatures. We used oxygen isotope fractionation to measure the distribution of electrons between the main and alternative pathways in mung bean (Vigna radiata) and soybean (Glycine max) following growth at low temperature. The amount of alternative oxidase protein in mung bean grown at 19 degrees C increased over 2-fold in both hypocotyls and leaves compared with plants grown at 28 degrees C but was unchanged in soybean cotyledons grown at 14 degrees C compared with plants grown at 28 degrees C. When the short-term response of tissue respiration was measured over the temperature range of 35 degrees C to 9 degrees C, decreases in the activities of both main and alternative pathway respiration were observed regardless of the growth temperature, and the relative partitioning of electrons to the alternative pathway generally decreased as the temperature was lowered. However, cold-grown mung bean plants that up-regulated the level of alternative oxidase protein maintained a greater electron partitioning to the alternative oxidase when measured at temperatures below 19 degrees C supporting a role for the alternative pathway in response to low temperatures in mung bean. This response was not observed in soybean cotyledons, in which high levels of alternative pathway activity were seen at both high and low temperatures.

Differential expression of alternative oxidase genes in soybean cotyledons during postgerminative development

The expression of the alternative oxidase (AOX) was investigated during cotyledon development in soybean (Glycine max [L.] Merr.) seedlings. The total amount of AOX protein increased throughout development, not just in earlier stages as previously thought, and was correlated with the increase in capacity of the alternative pathway. Each AOX isoform (AOX1, AOX2, and AOX3) showed a different developmental trend in mRNA abundance, such that the increase in AOX protein and capacity appears to involve a shift in gene expression from AOX2 to AOX3. As the cotyledons aged, the size of the mitochondrial ubiquinone pool decreased. We discuss how this and other factors may affect the alternative pathway activity that results from the developmental regulation of AOX expression.

The Regulation of Electron Partitioning between the Cytochrome and Alternative Pathways in Soybean Cotyledon and Root Mitochondria.

The regulation of electron partitioning between the cytochrome (Cyt) and alternative pathways in soybean (Glycine max L. cv Ransom) mitochondria in the absence of added inhibitors has been studied using the oxygen isotope fractionation technique. This regulation can depend on several factors, including the amount of alternative oxidase protein, the redox status of the alternative oxidase regulatory sulfhydryl-disulfide system, the degree of activation by [alpha]-keto acids, and the concentration and redox state of the ubiquinone pool. We studied electron partitioning onto the alternative pathway in mitochondria isolated from etiolated and light-grown cotyledons and roots to ascertain how these factors interact in different tissues. In light-grown cotyledon mitochondria there is some partitioning to the alternative pathway in state 4, which is increased dramatically by either pyruvate or dithiothreitol. In etiolated cotyledon mitochondria, the alternative pathway shows little ability to compete for electrons with the Cyt pathway under any circumstances. In root mitochondria, control of alternative pathway activity is exercised by both the ubiquinone pool and the regulatory sulfhydryl-disulfide system. In addition, oxygen isotope fractionation by the Cyt and alternative pathways in mitochondria were identical to the fractionation for the respective pathways seen in intact tissue, suggesting that residual respiration is not present in the absence of inhibitors.