Capacity Of Alternative Pathway Research Articles

Contribution of the alternative pathway on spring rejuvenation of alfalfa

Alfalfa often suffers from low temperature during spring rejuvenation, so it is important to improve the cold tolerance of alfalfa leaves for its smooth rejuvenation, and the alternative pathway (AP) could effectively improve the plant's tolerance. In this study, the contribution of AP on spring rejuvenation of alfalfa was investigated in Xinmu No.4 and Gannong No.5 with different fall dormancy levels. Though the protein and AP capacity were decreased during the rejuvenation, the ratio of AP/TP were increased in two alfalfa varieties, compared to those in alfalfa before overwintering. This indicated that AP had positive response to alfalfa rejuvenation. The limitation of AP significantly affected the leaf length, leaf width and growth rate of greening alfalfa, showing that AP played an important role in alfalfa rejuvenation. Inhibition of AP resulted in a significant decrease in Pn, Ci, Gs and stomatal structure deformity, suggestion that AP affected photosynthesis by influencing stomatal development during rejuvenation. AP reduces oxidative damage to PSII core protein repair in alfalfa leaves and optimizes photosynthesis by up-regulating NADP-MDH activity, decreasing the accumulation of excess reducing power in the chloroplasts, and by increasing SOD and POD activities and decreasing the accumulation of hydrogen peroxide. The higher proportion of AP keeps it more tolerant to low temperature for rejuvenation in Xinmu No.4 with a lower fall dormancy level.

Nephritis-Associated Plasmin Receptor (NAPlr): An Essential Inducer of C3-Dominant Glomerular Injury and a Potential Key Diagnostic Biomarker of Infection-Related Glomerulonephritis (IRGN).

Nephritis-associated plasmin receptor (NAPlr) was originally isolated from the cytoplasmic fraction of group A Streptococci, and was found to be the same molecule as streptococcal glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and plasmin receptor (Plr) on the basis of nucleotide and amino acid sequence homology. Its main functions include GAPDH activity, plasmin-binding capacity, and direct activation of the complement alternative pathway (A-P). Plasmin trapped by deposited NAPlr triggers the degradation of extracellular matrix proteins, such as glomerular basement membranes and mesangial matrix, and the accumulation of macrophages and neutrophils, leading to the induction of plasmin-related endocapillary glomerular inflammation. Deposited NAPlr at glomerular endocapillary site directly activates the complement A-P, and the endocapillary release of complement-related anaphylatoxins, C3a and C5a, amplify the in situ endocapillary glomerular inflammation. Subsequently, circulating and in situ-formed immune complexes participate in the glomerular injury resulting in NAPlr-mediated glomerulonephritis. The disease framework of infection-related glomerulonephritis (IRGN) has been further expanded. GAPDH of various bacteria other than Streptococci have been found to react with anti-NAPlr antibodies and to possess plasmin-binding activities, allowing glomerular NAPlr and plasmin activity to be utilized as key biomarkers of IRGN.

Complement Activation Profiles in Patients with Juvenile Idiopathic Arthritis

Objectives: Besides T- and B-cells also the innate immunity is playing an effective role in the pathogenesis of Juvenile idiopathic arthritis (JIA) reported in literature. The complement system (CS) is activated by the three different ways classical pathway (CP), lectin pathway (LP) and alternative pathway (AP). This is a controlled prospective observational single center study focused on the three pathways of CS and its effector, the terminal complement complex (TCC), associated with disease activity in all clinical JIA subgroups. Methods: Peripheral blood (PB) samples (n=159) of 60 JIA patients (partially also longitudinal visits) were analyzed for specific complement pathway activation (COMPL300 ELISA), complement factor H (CFH)-autoantibodies (CFHAb ELISA), and the soluble TCC (sC5b-9 ELISA) in serum (S) and EDTA plasma (P). TCC was also analyzed in synovial fluid (SF) of JIA patients (n=9) controlled with samples of healthy adults (n=12) obtained by arthroscopy (primarily gonarthrosis). As healthy controls (HC) adults (n=100) and children (n=18) without inflammatory diseases were tested. The JADAS10 defined the acute phase of the disease. Results: We compared the elevated serum TCC levels of JIA patients in the acute phase of the disease (13.32 IQR 8.09-19.48; p<0,001) with the controls' levels (7.78 IQR 4.93-10.31). Subgroup analyses showed statistically significant (s.s.) elevation in PArf+, OAper, and OAext. COMPL300 analyses revealed that JIA patients with acute disease had lower CP (82% IQR 37-97% vs 105% IQR 98-115%; p<0.001), LP (31% IQR 2-86% vs 80% IQR 20-101%; p<0.001), and AP (25% IQR 2-88% vs 85% IQR 70-98%; p<0.001) capacity compared to the HC average due to chronic activation in all three pathways. Both TCC and COMPL300 analyses during acute phase and remission showed a clear tendency, especially in PArf+ but only s.s. in the individual course. Remarkable sJIA patients neither in TCC nor in COMPL300 analyses showed complement activation. No evidence of CFH-autoantibodies was found in our study group. The TCC levels in SF showed a median of 2.48 IQR 1.71-2.76 AU/ml in JIA patients and surprisingly high levels in HC with 6.7 IQR 3.9-9.24 AU/ml. Conclusion: Specific subgroups of JIA patients (extOA, PARf+, ERA) showed increased CS activation in TCC in PB in the acute course of the disease. Furthermore, the decreased CS capacity, especially in the CP and AP, confirms the consumption of complement components as an additional contributor to the disease pathogenesis and acute course. Due to the CS activation throughout all disease activity states, we could not determine s.s. between the acute and remission phases in our cohort. However, COMPL300 in combination with TCC seems to be a helpful acute disease biomarker in the individual course. The pharmacological modulation of CS components might be a valid alternative approach in treatmentresistant patients, although further investigations are needed.

Alternative Pathway Is Involved in Hydrogen Peroxide-Enhanced Cadmium Tolerance in Hulless Barley Roots.

Hulless barley, grown in the Qinghai Tibet Plateau, has a wide range of environmental stress tolerance. Alternative pathway (AP) and hydrogen peroxide (H2O2) are involved in enhancing plant tolerance to environmental stresses. However, the relationship between H2O2 and AP in hulless barley tolerance to cadmium (Cd) stress remains unclear. In the study, the role and relationship of AP and H2O2 under Cd stress were investigated in hulless barley (Kunlun14) and common barley (Ganpi6). Results showed that the expression level of alternative oxidase (AOX) genes (mainly AOX1a), AP capacity (Valt), and AOX protein were clearly induced more in Kunlun14 than in Ganpi 6 under Cd stress; moreover, these parameters were further enhanced by applying H2O2. Malondialdehyde (MDA) content, electrolyte leakage (EL) and NAD(P)H to NAD(P) ratio also increased in Cd-treated roots, especially in Kunlun 14, which can be markedly alleviated by exogenous H2O2. However, this mitigating effect was aggravated by salicylhydroxamic acid (SHAM, an AOX inhibitor), suggesting AP contributes to the H2O2-enhanced Cd tolerance. Further study demonstrated that the effect of SHAM on the antioxidant enzymes and antioxidants was minimal. Taken together, hulless barley has higher tolerance to Cd than common barley; and in the process, AP exerts an indispensable function in the H2O2-enhanced Cd tolerance. AP is mainly responsible for the decrease of ROS levels by dissipating excess reducing equivalents.

Nitric oxide-mediated alternative pathway alleviates aluminum-induced programmed cell death in soybean root tips

Alternative pathway (AP) plays essential roles in plant adaptation to environmental stress. However, the exact role of AP in response to aluminum (Al) toxicity remains elusive. We here provide solid evidences that the activated AP capacity in root tips of soybean alleviated Al toxicity. Furthermore, inhibition of AP by pharmacological or transgenic approach aggravated Al-induced programmed cell death (PCD) occurrence mediated through reactive oxygen species (ROS)-dependent mitochondrial pathway. Our results also demonstrated that nitric oxide (NO) plays a negative role in PCD occurrence caused by Al in soybean root tips. Interestingly, the alleviating effect of NO on Al-induced PCD could be blocked by AP inhibition. Further investigation showed that NO mediates the induction of AP resulting from the upregulation of AOX expression and pyruvate content in Al-treated root tips of soybean. Taken together, our results clearly suggest that AP participates in the alleviation of Al toxicity and also plays a critical role in the alleviating effect of NO on Al-induced PCD occurrence, which will open up new avenues for the improvement of plant growth in acidic soils.

A specific promoter-type in ribonuclease L gene is associated with phagocytic activity in pigs.

We have previously generated Large White pigs with high immune competence using a selection strategy based on phagocytic activity (PA), capacity of alternative complement pathway, and antibody response after vaccination against swine erysipelas. In this study, to identify the genetic changes caused by the immune selection pressure, we compared gene expression and polymorphisms in the promoter region between pigs subjected to the immune selection (immune-selected pigs) and those that were not (non-selected pigs). After lipid A stimulation, using a microarray analysis, 37 genes related to immune function and transcription factor activity showed a greater than three-fold difference in expression between macrophages derived from immune-selected and non-selected pigs. We further performed a polymorphic analysis of the promoter region of the differentially expressed genes, and elucidated the predominant promoter-types in the immune-selected and non-selected pigs, respectively, in the genes encoding ribonuclease L (RNASEL), sterile α motif and histidine-aspartate domain containing deoxynucleoside triphosphate triphosphohydrolase 1, signal transducer and activator of transcription 3, and tripartite motif containing 21. Analysis of the association between these promoter genotypes and the immune phenotypes revealed that the immune-selected promoter-type in RNASEL was associated with increased PA and was inherited recessively. Considering that RNASEL has been reported to be involved in antimicrobial immune response of mice, it may be possible to enhance the PA of macrophages and improve disease resistance in pig populations using RNASEL promoter-type as a DNA marker for selection.

Involvement of active MKK9-MAPK3/MAPK6 in increasing respiration in salt-treated Arabidopsis callus.

Mitogen-activated protein kinase kinase 9 (MKK9) is an upstream activator of mitogen-activated protein kinase 3 (MAPK3) and MAPK6 in planta. To investigate MKK9 roles in mitochondrial respiration in Arabidopsis, MKK9DD, the active allele with mutations of Thr-201 and Ser-205 to Asp, and MKK9KR, the allele lacking MKK9 activity with a mutation of Lys-76 to Arg, were used. Results showed that the total respiratory rate (Vt), alternative pathway capacity (Valt) and cytochrome pathway capacity (Vcyt) increased under 0-100mM NaCl treatments but decreased under 150-300mM NaCl treatments in Col-0 callus. However, the activation of MKK9 by dexamethasone (DEX) increased Vt, Valt and Vcyt under 200mM NaCl treatment; moreover, Valt showed more increase than Vcyt. The activation of MKK9 in MKK9DD callus sharply increased AOX protein expression under normal and NaCl conditions, but the increase was not observed in MKK9KR callus. Further results indicated that MAPK3 and MAPK6 were involved in the MKK9-induced increase of AOX protein levels. qRT-PCR results showed that MKK9-MAPK3/MAPK6 was involved in the NaCl-induced AOX1b and AOX1d expression, but only MKK9-MAPK3 was necessary for AOX2 expression; in addition, MAPK3 regulated the AOX1a transcription in an MKK9-independent manner. MKK9 positively regulated SOD and CAT activities by affecting MAPK3 and MAPK6 and negatively regulated APX and POD activities by affecting MAPK3. Moreover, MKK9 functions as a positive factor in H2O2 accumulation under salt stress. The regulation of ethylene on alternative respiration was also associated with MKK9 under salt stress. Taken together, the MKK9-MAPK3/MAPK6 pathway plays a pivotal role in increasing alternative respiration in the salt-treated Arabidopsis callus.

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.

Altered levels of AOX1a expression result in changes in metabolic pathways in Arabidopsis thaliana plants acclimated to low dose rates of ultraviolet B radiation

UV-B is a damaging component of solar radiation that inevitably reaches the Earth’s surface. Plants have developed response mechanisms to adapt to UVB exposure. The alternative oxidase (AOX) catalyzes the ATP-uncoupling cyanide-resistant alternative pathway (AP) in plant mitochondria and is thought to be an important part of the cellular defense network under stress conditions. This study aimed to unravel the poorly understood functional significance of AOX1a induction in Arabidopsis thaliana leaves exposed to ecologically relevant doses of UVB radiation, by comparing wild-type (WT) plants with plants with modified expression of the AOX1a gene, either downregulated by antisense (AS-12) or overexpressed (XX-2). UVB exposure resulted in a phenotypic difference between lines. AOX1a overexpression resulted in the highest induction of AOX1A synthesis and MnSOD activity, and the lowest ROS level without pronounced changes in the phenotype relative to other genotypes. In AS-12 plants, expression of the majority of the genes encoding AOX was detected, other non-phosphorylating pathway components and antioxidant enzymes increased along with anthocyanin accumulation in leaves, and the ROS content was lower than in the WT. In addition to the expected AOX1 protein size (34 kDa), an AOX1 30 kDa band appeared under UVB exposure in all genotypes. However, in AS-12, the alterations in the transcript level and in the abundance of AOX1 protein isoforms induced by UVB could not fully functionally compensate for the lack of AOX1A. This was confirmed by the observed low AP capacity and increased levels of the oxidized form of ascorbate. These results highlight the importance of AOX in plant response to UVB for the control of a balanced metabolism, and indicate that AOX1a plays a key role in the regulation of the stress response.

Mitochondrial respiration after combined action of dehydration and low temperature in pea seedlings

The impact of the individual and combined action of dehydration and low temperature on the respiratory activity of mitochondria isolated from epicotyls of pea (Pisum sativum L.) seedlings was investigated. These organelles were isolated immediately after the treatments and after two and three days of recovery. After the treatments, the capacity of the energy-producing cytochrome pathway (CP) of mitochondrial oxidation decreased, while the capacity of the non-phosphorylating alternative respiratory pathway (AP) catalyzed by a CN-resistant oxidase was elevated. After returning the seedlings to normal growing conditions, a gradual recovery of the respiratory activity of mitochondria was observed due to the inactivation of the AP and an elevation of the activity of the CP. The dehydration was a strong stress factor and led to a more significant disturbance of mitochondrial functioning than low temperature, and the complete recovery of the respiratory activity was observed only after the combined action of these stress factors. The post-stress reactivation of malate rather than succinate oxidation required a longer maintenance of seedlings under the control conditions.

Mitochondrial alternative cyanide-resistant oxidase is involved in an increase of heat stress tolerance in spring wheat

The aim of this study was to determine the influence of different heat treatments on the alternative cyanide-resistant oxidase (AOX) capacity and establish a relation between the heat stress tolerance of spring wheat (Triticum aestivum L.), content of water-soluble carbohydrates in leaves and the alternative respiratory pathway (AP) capacity. We identified a positive relation between these studied parameters. Heat exposure at 39 °C for 24 h increased the heat stress tolerance of seedlings, content of water-soluble carbohydrates and AOX capacity, and the AOX capacity was also high after the subsequent influence of heat shock (50 °C for 3 h). The increased AOX capacity correlated with an increased level of water-soluble carbohydrates in leaves. The content of the AOX protein increased after heat exposure at 39 °C (for 3 h and 24 h) and after the subsequent influence of heat shock (50 °C for 1 and 3 h) at 39 °C for 24 h. We also detected that the content of AOX protein isoforms depends on the duration and intensity of heat treatment. It was concluded that AOX plays an important role in the acclimation of plants to high temperatures.

Effect of the exogenous anthocyanin extract on key metabolic pathways and antioxidant status of Brazilian elodea (Egeria densa (Planch.) Casp.) exposed to cadmium and manganese

Present study deals with the effect of 24 h pre-incubation with exogenous anthocyanins (ANTH), extracted from red cabbage leaves, on key metabolic processes (photosynthesis and respiration) and pro-/antioxidant balance in the aquatic macrophyte Egeria densa (Planch.) Casp., Hydrocharitaceae family, treated with Cd and Mn (in sulfate form) at a concentration of 100 μmol. After five days of metal treatments, Cd was accumulated and the damage caused to metabolic processes was stronger than Mn. In Cd-treated leaves, the protein level, chlorophyll concentration and maximal photochemical efficiency of PS II decreased twofold, and net-photosynthesis was significantly inhibited, whereas lipid peroxidation and H2O2 production increased. In turn, protective responses developed, including an increase in the total soluble thiols, alternative respiratory pathway capacity and the activity of superoxide dismutase and peroxidases. Pre-incubation in the ANTH-enriched extract caused an increase in foliar ANTH content, enhanced Cd and reduced Mn uptake into the tissue. A decrease in the level of oxidative reactions, an increase in the protein and chlorophyll concentration compared to the control values and a partial improvement of the photosynthetic parameters confirmed the ability of ANTH to reduce Cd-induced damage effects and to mitigate ROS-driven stress reactions. Stimulation of catalase and ascorbate peroxidase activity, an alternative respiration capacity and non-enzymatic antioxidant (carotenoids, ascorbate and proline) synthesis by ANTH were also revealed. These data suggest that ANTH-enriched extract from red cabbage leaves has a protective action against metal toxicity in Egeria plants.

H<sub>2</sub>O<sub>2</sub>-induced acclimation of photosystem II to excess light is mediated by alternative respiratory pathway and salicylic acid

Acclimation to excess light is required for optimizing plant performance under natural environment. The present work showed that the treatment of Arabidopsis leaves with exogenous H2O2 can increase the acclimation of PSII to excess light. Treatments with H2O2 also enhanced the capacity of the mitochondrial alternative respiratory pathway and salicylic acid (SA) content. Our work also showed that the lack in alternative oxidase (AOX1a) in AtAOX1a antisense line and the SA deficiency in NahG (salicylate hydroxylase gene) transgenic mutant attenuated the H2O2-induced acclimation of PSII to excess light. It indicates that the H2O2-induced acclimation of PSII to excess light could be mediated by the alternative respiratory pathway and SA.

The Chlamydomonas alternative oxidase 1 is regulated by cadmium stress: New insights into control of expression

Although mitochondrial alternative oxidase (AOX) has been proposed to play essential roles in stress tolerance of plants, the effects of cadmium (Cd), an important environmental pollutant, on AOX are largely unknown. The green alga Chlamydomonas reinhardtii reacts to Cd stress conditions with the induced expression of many genes. In this work, we demonstrated a sharp increase in AOX1 transcript and protein abundance under Cd stress. Furthermore, C. reinhardtii cells displayed a large increase in alternative respiration in response to Cd treatment. Cd-induced AOX1 production and alternative pathway capacity were lower in illuminated cells than in dark-adapted cells. Moreover, in dark-adapted unstressed cells, AOX1 was significantly expressed. The results clearly indicate that the dark-mediated signals function in an additive manner with Cd-mediated signals, and thus likely act in parallel pathways to regulate the expression of AOX1.

Alternative pathway is involved in the tolerance of highland barley to the low-nitrogen stress by maintaining the cellular redox homeostasis.

Alternative pathway (AP) is involved in the tolerance of highland barley seedlings to the low-nitrogen stress by dissipating excessive reducing equivalents generated by photosynthesis and maintaining the cellular redox homeostasis. Low nitrogen (N) is a major limiting factor for plant growth and crop productivity. In this study, we investigated the roles of the alternative pathway (AP) in the tolerance of two barley seedlings, highland barley (Kunlun12) and barley (Ganpi6), to low-N stress. The results showed that the chlorophyll content and the fresh weight decreased more in Ganpi6 than those in Kunlun12 under low-N stress, suggesting that Kunlun12 has higher tolerance to low-N stress than Ganpi6. AP capacity was markedly induced by low-N stress; and it was higher in Kunlun12 than in Ganpi6. Comparatively, the cytochrome pathway capacity was not affected under all conditions. Western-blot analysis showed that the protein level of the alternative oxidase (AOX) increased under low-N stress in Kunlun12 but not in Ganpi6. Under low-N stress, the NAD(P)H content and the NAD(P)H to NAD(P)(+)+NAD(P)H ratio in Ganpi6 increased more than those in Kunlun12. Furthermore, photosynthetic parameters (Fv/Fm, qP, ETR and Yield) decreased markedly and qN increased, indicating photoinhibition occurred in both barley seedlings, especially in Ganpi6. When AP was inhibited by salicylhydroxamic acid (SHAM), the NAD(P)H content and the NAD(P)H to NAD(P)(+)+NAD(P)H ratio dramatically increased under all conditions, resulting in the marked accumulation of H(2)O(2) and malondialdehyde in leaves of both barley seedlings. Meanwhile, the malate-oxaloacetate shuttle activity and the photosynthetic efficiency were further inhibited. Taken together, AP is involved in the tolerance of highland barley seedlings to low-N stress by dissipating excess reducing equivalents and maintaining the cellular redox homeostasis.

Effect of 24-epibrassinolide on Arabidopsis thaliana alternative respiratory pathway under salt stress

Using wild-type (WT) Arabidopsis thaliana plants and two transgenic lines—overexpressing alternative oxidase AOX1a (AOX1a-OE) and antisense AOX1a (AOX1a-AS)—it was observed that brassinosteroids (BRs) are involved in regulation of respiration in A. thaliana leaf, particularly alternative respiratory pathway, under salt stress conditions. This study demonstrates that BRs stimulate total respiration rate, alternative respiratory pathway, and shift capacity of alternative respiratory pathway comparing to cyanide-sensitive pathway. Lowering the level of endogenous BRs with the specific inhibitor of BRs biosynthesis—brassinazole—caused the decrease of alternative respiratory pathway activity in WT plants and transgenic AOX1a-OE and AOX1a-AS lines.

Light regulation of mitochondrial alternative oxidase pathway during greening of etiolated wheat seedlings

This study deals with effects of de-etiolation (48h) of spring wheat (Triticum aestivum L., var. Irgina) seedlings on differential expression of AOX1 genes, levels of AOX protein and the alternative respiratory pathway (AP) capacity. As a result of exposure to continuous irradiation of dark-grown wheat seedlings, the respiratory activity and AP capacity in leaves significantly increased during the first 6h of studies. Expression of AOX1a was up-regulated by light and proved consistent with changes in the AP capacity. Effects on expression of AOX1c were less pronounced. Immunoblot analysis showed three distinct bands of AOX with molecular weights of 34, 36 and 38kDa, with no significant changes in the relative levels during de-etiolation. The lack of a clear correlation between AOX protein amount, AOX1a expression, and AP capacity suggests post-translational control of the enzyme activation. The AOX1a suppression and a decrease in the AP capacity correlated with the sugar pool depletion after 24h of the de-etiolation, which may mean a possible substrate dependence of the AOX activity in the green cells. More efficient malate oxidation by mitochondria as well as the higher AOX capacity during the first 6h of de-etiolation was detected, whereas respiration and AOX capacity with exogenous NADH and glycine increased after 6 and 24h, respectively. We conclude that AOX plays an important role during development of an actively photosynthesizing cell, and can rapidly adapt to changes in metabolism and photosynthesis.

Effects of modifying alternative respiration on nitric oxide-induced virus resistance and PR1 protein accumulation.

Nitric oxide (NO) is an important defensive signal in plants but its effects on virus infection are not well understood. Administration of NO-releasing compounds immediately before inoculation of tobacco leaves with potato virus X and tobacco mosaic virus decreased the accumulation of virus, indicating that NO can induce resistance rapidly. Resistance induction was inhibited by co-administration with an NO-scavenging compound or when experiments were done in transgenic tobacco plants expressing increased alternative respiratory pathway capacity due to constitutive expression of the plant mitochondrial enzyme, alternative oxidase (AOX). These results indicate that NO, which inhibits electron transport chain activity, is triggering defensive signalling by inducing changes in mitochondrial reactive oxygen species levels that are in turn regulated by AOX. Experiments using nahG-transgenic plants, which cannot accumulate the defensive plant hormone salicylic acid (SA) showed that NO rapidly induces resistance to virus infection independently of SA. However, this initial state of resistance may be transient. Subsequently, by 5 days post-treatment, NO had caused an increase in pathogenesis-related protein 1 (PR1) expression (a proxy for increased SA biosynthesis), which correlated with a longer-term state of resistance to virus infection. The induction by NO of PR1 accumulation was modified in AOX-transgenic plants. This indicates that the influence of NO on defensive gene expression is in part mediated through its effects on mitochondria.

Involvement of alternative oxidase in the regulation of sensitivity of Sclerotinia sclerotiorum to the fungicides azoxystrobin and procymidone

Sclerotinia sclerotiorum is a filamentous fungal pathogen that can infect many economically important crops and vegetables. Alternative oxidase is the terminal oxidase of the alternative respiratory pathway in fungal mitochondria. The function of alternative oxidase was investigated in the regulation of sensitivity of S. sclerotiorum to two commercial fungicides, azoxystrobin and procymidone which have different fungitoxic mechanisms. Two isolates of S. sclerotiorum were sensitive to both fungicides. Application of salicylhydroxamic acid, a specific inhibitor of alternative oxidase, significantly increased the values of effective concentration causing 50% mycelial growth inhibition (EC50) of azoxystrobin to both S. sclerotiorum isolates, whereas notably decreased the EC50 values of procymidone. In mycelial respiration assay azoxystrobin displayed immediate inhibitory effect on cytochrome pathway capacity, but had no immediate effect on alternative pathway capacity. In contrast, procymidone showed no immediate impact on capacities of both cytochrome and alternative pathways in the mycelia. However, alternative oxidase encoding gene (aox) transcript and protein levels, alternative respiration pathway capacity of the mycelia were obviously increased by pre-treatment for 24 h with both azoxystrobin and procymidone. These results indicate that alternative oxidase was involved in the regulation of sensitivity of S. sclerotiorum to the fungicides azoxystrobin and procymidone, and that both fungicides could affect aox gene expression and the alternative respiration pathway capacity development in mycelia of this fungal pathogen.

Metabolic interconnectivity among alternative respiration, residual respiration, carbohydrates and phenolics in leaves of Salvinia minima exposed to Cr(VI)

Floating and submerged leaves of the aquatic fern Salvinia minima were used to analyze a metabolic interconnectivity among mitochondrial alternative respiration, residual respiration (Rresp), carbohydrate metabolism and soluble phenolics (SP) accumulation occurring under Cr(VI) stress. Treatment with Cr enhanced alternative pathway capacity (APcap) and (Rresp) in both leaf types. APcap/Tresp ratio revealed an increasing relative contribution of the alternative respiration to total respiration rate under Cr(VI) treatment. Sucrose content increased in Cr-treated leaves, but glucose and starch decreased. Enzyme profile showed that sucrose synthase (SS) rather than soluble acid invertase (AI) seems to be involved in sucrose metabolism of Cr-treated plants. Accumulation of SP showed a positive correlation with both APcap and Rresp in floating leaves. Decreases of SP in submerged leaves can be explained by an increased synthesis of polymerized phenolics. Results provide important new insights about influence of alternative and residual respirations on the synthesis of phenylpropanoid-derivative compounds. This work could also represent the first communication about involvement of the Rresp in defence mechanism of S. minima against Cr(VI) toxicity.