The role of proline-associated pentose phosphate pathway in cool-season turfgrasses after UV-B exposure

Physiological adjustments of cool-season turfgrasses were investigated to determine the role of proline-associated pentose phosphate pathway for phenolic biosynthesis and stimulation of antioxidant response system following UV-B exposure. Creeping bentgrass, Kentucky bluegrass, tall fescue, and perennial ryegrass plugs were subjected to artificial UV-B exposure (biologically effective UVB_BE radiation 8 kJ m⁻² d⁻¹.) for one week with 10-h photoperiod. Significant accumulation of phenolics and stimulation of antioxidant enzyme activity was observed in all investigated cool-season turfgrasses after UV-B exposure and this induction corroborated with higher glucose-6-phosphate dehydrogenase activity and high accumulation of proline. Guaiacol peroxidase activity also increased in all investigated cool-season turfgrass species after UV-B exposure. In this study, the shift of carbon flux from glycolysis to pentose phosphate pathway following UV-B exposure and as a result of that, the higher accumulation of phenolics and stimulation of antioxidant response system provides an insight to understand a probable defense mechanism of cool-season turfgrasses against UV stress.     

Relationship of secondary metabolism to growth in oregano (Origanum vulgare L.) shoot cultures under nutritional stress

Micropropagation of Origanum vulgare L. by shoot buds, as a potential model system for studying carbon skeleton diversion from growth to secondary metabolism as adaptive response to nutrient deficiency, has been performed. In addition, the antioxidant phenolic compounds, produced by shoots under nutritional stress or in response to exogenously added proline, have been studied. Caffeic acid, rosmarinic acid, and lithospermic acid B have been isolated in oregano shoot cultures by reversed-phase high-performance liquid chromatography, and their structures have been elucidated by tandem mass spectrometry. Both nutritional stress, which in turn causes a moderate increase of constitutive free proline, and exogenous proline affect growth and antioxidant phenolic content of oregano shoots, compared to control.The role of proline, and the associated redox cycle, as a form of metabolic signaling based on a transfer of redox potential amongst interacting cell pathways, which in turn elicit phenolic metabolism via stimulated carbon flux through oxidative pentose phosphate pathway, is discussed. Furthermore, the potential use of oregano tissue and callus cultures as a new strategy to enable the production of useful secondary metabolites on a commercial scale is also discussed.     

Effect of vitamin C and folic acid on seed vigour response and phenolic-linked antioxidant activity

Folic acid and vitamin C were used in the concentration range of 0-500muM as exogenous growth enhancers to stimulate pea (Pisum sativum) seedling vigour. The results suggest that a concentration of 50muM folic acid and 500muM vitamin C were optimum in maximally enhancing seed vigour and potentially seedling performance according to both agronomic and biochemical seed vigour parameters. Results indicated that germination percentage, shoot weight, shoot height, and root length were enhanced in folic acid and vitamin C treated plants compared to control plants. The levels of enhanced phenolic content in response to folic acid and vitamin C treatments were highest on days 8 and 10. Evaluation of critical biochemical parameters indicated that the average glucose-6-phosphate dehydrogenase (G6PDH) activity and proline content in response to treatments were higher than control and correlated to enhanced phenolic content and DPPH-based antioxidant activity. Key enzymes, guaiacol peroxidase (GPX), superoxide dismutase (SOD), and catalase (CAT) were also higher in response to treatments and correlated to enhanced phenolic content and DPPH-based antioxidant activity. Taken together, these studies support the hypothesis that the proline-linked pentose phosphate pathway stimulates phenolic synthesis and related free-radical scavenging antioxidant activity. Further, this proline-linked pentose phosphate pathway stimulation in response to folic acid and vitamin C was also correlated to antioxidant enzyme response indicated by the stimulation of GPX, SOD, and CAT activities. Therefore, this study indicates the enhancement of seed vigour response by folic acid and vitamin C as reflected in both agronomic and biochemical responses, and this occurred through the stimulation of phenolic-linked antioxidant response that is likely positively modulated through the proline-linked pentose phosphate pathway.     

Stimulation of total phenolics, L-DOPA and antioxidant activity through proline-linked pentose phosphate pathway in response to proline and its analogue in germinating fava beans (Vicia faba)

The rationale behind the study was to enhance azetedine-2-carboxylate (A2C)-linked stress in the germinating seeds to which it responds by increased proline synthesis to overcome inhibition of proline dehydrogenase (PDH). A2C is a competitive inhibitor of proline that inhibits its transport from cytosol to the mitochondria for further metabolic recycling by binding on to the active sites of PDH. The enhanced synthesis of proline would increase the ratio of the cofactors NADP/NADPH₂. The increase in the cofactors would result in the over-expression of the phenypropanoid pathway required for the phenolic acids and L-DOPA synthesis via pentose phosphate pathway through the activity of glucose-6-phosphate dehydrogenase (G6PDH). Fava bean were chosen since it has high phenolic and L-DOPA levels and could be an important part of the diet especially for patients suffering from Parkinson's Disease. The objectives were investigated by assaying for total phenolic content, the corresponding antioxidant activity by β-carotene oxidation method, proline levels and enzymes such as G6PDH and guaiacol peroxidase (GPX) using spectrophotometric methods. L-DOPA was quantified using HPLC. The fava bean seeds were primed with water, 200 μM A2C, 0.25 mM proline and a combination of A2C and proline treatments. After the priming stage, seeds were dark germinated and grown for a period of 8 days, for biochemical analysis. L-DOPA levels did not change in comparison to the control treatments while total phenolic content, proline and G6PDH were all enhanced by the treatments. During the early stages of germination the phenolic acids were antioxidant in nature, which later was reduced as they become polymerized to lignins and lignans via the GPX activity. Total phenolic synthesis was both coupled and uncoupled to PLPPP depending on the treatments. The three treatments over-expressed PLPPP since large difference between control and the treatments were observed for all parameters, except L-DOPA content.     

Transfer of 1-pyrroline-5-carboxylate as oxidizing potential from hepatocytes to erythrocytes.

The interconversions of proline and 1-pyrroline-5-carboxylate form an intercellular cycle that is the basis of a metabolic interaction between hepatocytes and erythrocytes. The cycle transfers oxidizing potential from hepatocytes to erythrocytes, which stimulates pentose phosphate pathway in erythrocytes. This interaction depends on the differential metabolism of proline and 1-pyrroline-5-carboxylate in erythrocytes and hepatocytes and consists of the following: in hepatocytes proline oxidase converts proline into 1-pyrroline-5-carboxylate, which is released into the medium and taken up by erythrocytes; erythrocyte 1-pyrroline-5-carboxylate reductase converts 1-pyrroline-5-carboxylate into proline and concomitantly generates NADP+; the generated oxidizing potential drives glucose metabolism through the pentose phosphate pathway in erythrocytes; finally, erythrocytes release proline into the medium, enabling it to re-enter hepatocytes and repeat the cycle. The increased activity of the pentose phosphate pathway in erythrocytes may enhance the production of 5-phosphoribosyl pyrophosphate, a necessary moiety for the processing of purines.     

Poly-β-Hydroxybutyrate Metabolism Is Affected by Changes in Respiratory Enzymatic Activities Due to Cold Stress in Two Psychrotrophic Strains of Rhizobium

Cold stress resulted in a decrease in the poly-β-hydroxybutyrate (PHB) content of non-cold-acclimated Rhizobium DDSS69 cultures. Analysis of the specific activity of β-ketothiolase and β-hydroxybutyrate dehydrogenase revealed that decrease in PHB levels was a result of the inhibition of synthesis of PHB rather than an increase in its breakdown. Rhizobium ATR1, a cold-acclimated strain, revealed the presence of a stable PHB metabolism that did not show any significant differences either in PHB levels or in the activity of enzymes of the PHB metabolism under cold stress, suggesting that PHB is not involved in cold tolerance. Analysis of specific activities of glucose-6-phosphate dehydrogenase and 6-phosphogluconate dehydrogenase of the pentose phosphate pathway showed the upward regulation of alternate pathways of carbohydrate metabolism under cold stress to rapidly generate energy to overcome the stress. There is diversity in the switching mechanisms of carbon metabolism among cold-acclimated and non-cold-acclimated Rhizobium isolates. Upward regulation of malate dehydrogenase in both isolates suggests that it is a critical input for cold tolerance. Received: 26 June 2000 / Accepted: 31 July 2000     

Regulation and function of proline oxidase under nutrient stress

Under conditions of nutrient stress, cells switch to a survival mode catabolizing cellular and tissue constituents for energy. Proline metabolism is especially important in nutrient stress because proline is readily available from the breakdown of extracellular matrix (ECM), and the degradation of proline through the proline cycle initiated by proline oxidase (POX), a mitochondrial inner membrane enzyme, can generate ATP. This degradative pathway generates glutamate and α‐ketoglutarate, products that can play an anaplerotic role for the TCA cycle. In addition the proline cycle is in a metabolic interlock with the pentose phosphate pathway providing another bioenergetic mechanism. Herein we have investigated the role of proline metabolism in conditions of nutrient stress in the RKO colorectal cancer cell line. The induction of stress either by glucose withdrawal or by treatment with rapamycin, stimulated degradation of proline and increased POX catalytic activity. Under these conditions POX was responsible, at least in part, for maintenance of ATP levels. Activation of AMP‐activated protein kinase (AMPK), the cellular energy sensor, by 5‐aminoimidazole‐4‐carboxamide ribonucleoside (AICAR), also markedly upregulated POX and increased POX‐dependent ATP levels, further supporting its role during stress. Glucose deprivation increased intracellular proline levels, and expression of POX activated the pentose phosphate pathway. Together, these results suggest that the induction of proline cycle under conditions of nutrient stress may be a mechanism by which cells switch to a catabolic mode for maintaining cellular energy levels. J. Cell. Biochem. 107: 759–768, 2009. © 2009 Wiley‐Liss, Inc.     

Metabolite Responses to Exogenous Application of Nitrogen,Cytokinin, and Ethylene Inhibitors in Relation to Heat-Induced Senescence in Creeping Bentgrass

The exogenous application of ethylene inhibitors, cyotkinins, or nitrogen has previously been shown to suppress heat-induced senescence and improve heat tolerance in cool -season grasses. The objectives of this study were to examine metabolic profiles altered by exogenous treatment of creeping bentgrass with an ethylene inhibitor, cytokinin or nitrogen under heat stress and to determine metabolic pathways regulated by those compounds in association with their effectiveness for improving heat tolerance. Creeping bentgrass (Agostis stolonifera) plants (cv. Penncross) were foliar sprayed with 18 mM carbonyldiamide (N source), 25μM aminoethoxyvinylglycine (AVG, ethylene inhibitor), 25μM zeatin riboside (ZR, cytokinin), or a water control, and then exposed to 20/15°C (day/night) or 35/30°C (heat stress) in growth chambers. All three exogenous treatments suppressed leaf senescence, as manifested by increased turf quality and chlorophyll content, and reduced electrolyte leakage under heat stress. Polar metabolite profiling identified increases in the content of certain organic acids (i.e. citric and malic acid), sugar alcohols, disaccharides (sucrose), and decreased accumulations of monosaccharides (i.e. glucose and fructose) with exogenous treatment of N, AVG, or ZR at the previously mentioned concentrations when compared to the untreated control under heat stress. Nitrogen stimulated amino acid accumulation whereas AVG and ZR reduced amino acid accumulation compared to the untreated control under heat stress. These results revealed that the alleviation of heat-induced leaf senescence by N, AVG, and ZR could be due to changes in the accumulation of metabolites involved in osmoregulation, antioxidant metabolism, carbon and nitrogen metabolism, as well as stress signaling molecules.     

Effects of SAG12-ipt expression on cytokinin production,growth and senescence of creeping bentgrass (<Emphasis Type="Italic">Agrostis stolonifera</Emphasis> L.) under heat stress

The study was conducted to determine the effects of expression of a transgene encoding adenine isopentenyl transferase (ipt), which controls cytokinin synthesis, on growth and leaf senescence of creeping bentgrass (Agrostis stolonifera L.), subjected to heat stress. Creeping bentgrass (cv. Penncross) was transformed with ipt ligated to a senescence-activated promoter (SAG12). Eight SAG12-ipt transgenic lines exhibiting desirable turf quality and a transgenic control line (transformed with the empty vector) were evaluated for morphological and physiological changes under normal growth temperature (20°C) and after 14 days of heat stress (35°C) in growth chambers. Six of the SAG12-ipt lines developed more tillers than the control line during establishment under normal growth temperature of 20°C. Following 14 days of heat stress, four of the SAG12-ipt lines had increased 65–83% of roots and for all six SAG12-ipt lines root elongation continued, whereas root production ceased and total root length decreased for the control line. Root isopentenyl adenine (iPA) content increased 2.5–3.5 times in five of the SAG12-ipt lines, whereas in the control line iPA decreased 20% after 14 days at 35°C. Total zeatin riboside (ZR) content was maintained at the original level or increased in five of the SAG12-ipt lines, whereas in the control line ZR decreased under heat stress. Our results suggest expression of SAG12-ipt in creeping bentgrass stimulated tiller formation and root production, and delayed leaf senescence under heat stress, suggesting a role for cytokinins in regulating cool-season grass tolerance to heat stress.     

Stimulation of the proline cycle and anthraquinone accumulation in <Emphasis Type="Italic">Rubia tinctorum</Emphasis> cell suspension cultures in the presence of glutamate and two proline analogs

Anthraquinone biosynthesis in Rubia tinctorum L. involves different metabolic routes. Chorismic acid, the end-product of the shikimate pathway, becomes the branch point between primary and secondary metabolism. It has been proposed that the proline cycle could be coupled with the pentose phosphate pathway (PPP), since the NADP⁺ generated by proline reduction from glutamate could act as a cofactor of the first enzymes of the PPP. This pathway generates erythrose-4-phosphate, the substrate of the shikimate pathway. The aim of the present work was to study the effect of the addition of glutamate and two proline analogs, azetidine-2-carboxylic acid and thiazolidine-4-carboxylic acid (T4C), on the PPP, the proline cycle, and anthraquinone production in R. tinctorum cell suspension cultures. The addition of 5 mM of glutamate enhanced both anthraquinone (up to 30%) and total phenolic content (12%), which correlated well with proline accumulation. Only the addition of 200 μM of T4C resulted in an increase in anthraquinone production, which was accompanied by a rise in the proline content. Neither the addition of glutamate nor proline analogs resulted in the induction of PPP, so this route was not a limiting factor as a carbon donor to the shikimate pathway.     

Protein accumulation in leaves and roots associated with improved drought tolerance in creeping bentgrass expressing an ipt gene for cytokinin synthesis

Cytokinins (CKs) may be involved in the regulation of plant adaptation to drought stress. The objectives of the study were to identify proteomic changes in leaves and roots in relation to improved drought tolerance in transgenic creeping bentgrass (Agrostis stolonifera) containing a senescence-activated promoter (SAG12) and the isopentyl transferase (ipt) transgene that increases endogenous CK content. Leaves of SAG12-ipt bentgrass exhibited less severe senescence under water stress, as demonstrated by maintaining lower electrolyte leakage and lipid peroxidation, and higher photochemical efficiency (F(v)/F(m)), compared with the null transformant (NT) plants. SAG12-ipt plants had higher root/shoot ratios and lower lipid peroxidation in leaves under water stress than the NT plants. The suppression of drought-induced leaf senescence and root dieback in the transgenic plants was associated with the maintenance of greater antioxidant enzyme activities (superoxide dismutase, peroxidase, and catalase). The SAG12-ipt and NT plants exhibited differential protein expression patterns under well-watered and drought conditions in both leaves and roots. Under equivalent leaf water deficit (47% relative water content), SAG12-ipt plants maintained higher abundance of proteins involved in (i) energy production within both photosynthesis and respiration [ribulose 1,5-bisphosphate carboxylase (RuBisCO) and glyceraldehyde phosphate dehydrogenase (GAPDH)]; (ii) amino acid synthesis (methionine and glutamine); (iii) protein synthesis and destination [chloroplastic elongation factor (EF-Tu) and protein disulphide isomerases (PDIs)]; and (iv) antioxidant defence system (catalase and peroxidase) than the NT plants. These results suggest that increased endogenous CKs under drought stress may directly or indirectly regulate protein abundance and enzymatic activities involved in the above-mentioned metabolic processes, thereby enhancing plant drought tolerance.     

Transgenic tobacco plants expressing grass <Emphasis Type="Italic">AstEXPA1</Emphasis> gene show improved performance to several stresses

Expansins are cell wall-loosening proteins and now widely accepted to associate with the plant resistance against various abiotic stresses. In this study, we cloned an expansin gene of AstEXPA1 from Agrostis stolonifera, a heat-resistant creeping bentgrass cultivar, and transformed it into tobacco plants. Physiological index test showed that the transgenic lines were resistant to various abiotic stresses of drought, heat, cold, and salt in comparison to non-transgenic plants. Comprehensive analysis of four physiological response indices showed that the transgenic plants performed much better resistance to drought, following to heat, cold and salt stress, respectively. Meanwhile soluble sugar content displayed more weight to plant resistance by over-expressing AstEXPA1 gene, followed as proline content, REL, and MDA content. The results here would expand our understanding of the expansin roles and drive better insights into plant molecular breeding against stress.     

Proline and betaine provide protection to antioxidant and methylglyoxal detoxification systems during cold stress in <Emphasis Type="Italic">Camellia sinensis</Emphasis> (L.) O. Kuntze

The aim of this study was to monitor the influence of proline and betaine exposure on antioxidant and methylglyoxal (MG) detoxification system during cold stress in Camellia sinensis (L.) O. Kuntze. Cold stress enhanced MG and lipid peroxidation levels in tea bud (youngest topmost leaf). This increase was resisted upon the exposure of tea bud to proline and betaine. Exposure of tea bud with proline and betaine also help in maintaining thiol/disulfide ratio during cold stress. Proline exposure enhanced glutathione-S-transferase and glutathione reductase (GR) activity, while betaine exposure increased only GR activity during cold stress. Furthermore, effect of proline/betaine was studied on glyoxalase pathway enzymes that are involved in MG detoxification and comprise of two enzymes glyoxalase I and glyoxalase II. Both proline and betaine showed protective effect on glyoxalase I and activating effect on glyoxalase II during cold stress in tea bud. This investigation, therefore, suggest that proline and betaine might provide protection to cold stress in tea by regulating MG and lipid peroxidation formation as well as by activating or protecting some of antioxidant and glyoxalase pathway enzymes.     

Effects of Cytokinin and Nitrogen on Drought Tolerance of Creeping Bentgrass

Cytokinin (CK) is a vital plant hormone that controls many aspects of growth and development in plants. Nitrogen (N) is the indispensable macronutrient needed in plants and also one of the most important limiting factors for plant growth. This study was designed to investigate the simultaneous effects of CK and N on the visual turf quality and antioxidant metabolism of drought-stressed creeping bentgrass (Agrostis stolonifera L.). ‘PennA-4’ creeping bentgrass treated with trans-zeatin riboside at three rates of CK concentrations of 0, 10 and 100 μM (designated by CK0, 10, and 100) and two nitrogen rates with 2.5 and 7.5 kg N·ha^-1 every 15 days (designated by low and high N) in a complete factorial arrangement was grown under two soil moisture regimes: well-watered and drought stress. Exogenous CK improved turf quality and delayed leaf wilting under drought stress, especially under high N. The grasses treated with CK10 and CK100 had lower O₂^- production and H₂O₂ concentration than those without CK treatment. The CK100 treatment enhanced the activities of superoxide dismutase (SOD), ascorbate peroxidase (APX), catalase (CAT), and guaiacol peroxidase (POD) by 25%, 22%, 17% and 24%, respectively, relative to CK0. Moreover, the activity changes of the antioxidant enzyme isoforms were more significant under high N condition relative to low N condition. Our results demonstrated the beneficial impacts of CK and N on physiological reactions, especially antioxidant metabolism, and foliar application of CK at 10 or 100 μM plus 7.5 kg ha^-1 N biweekly may improve drought stress resistance of creeping bentgrass.     

Antioxidative responses in roots and shoots of creeping bentgrass under high temperature: effects of nitrogen and cytokinin

It has been previously reported that either nitrogen (N) or cytokinin (CK) applications can alleviate heat stress injury on creeping bentgrass, with some studies reporting enhanced antioxidant metabolism being related to stress protection. The objective of this research was to investigate the simultaneous effects of CK and N on the antioxidant enzyme activity and isoforms of heat stressed creeping bentgrass. 'L-93' creeping bentgrass treated with three rates of CK (trans-zeatin riboside, tZR, 0, 10 and 100μM, designated by CK0, 10, and 100) and two nitrogen rates (2.5 and 7.5kgNha(-1) biweekly, low and high N) in a complete factorial arrangement was maintained in a 38/28°C (day/night) growth chamber for 28d and then harvested. Grass grown at high N (averaged across CK rates) had higher O(2)(-) production, H(2)O(2) concentration, and malondialdehyde content in roots. The activities of superoxide dismutase (SOD), ascorbate peroxidase (APX), and guaiacol peroxidase (POD) in roots were enhanced 19%, 22%, and 24%, respectively, by high N relative to low N. Twenty-eight days of heat stress resulted in either the development of new isoforms or enhanced isoform intensities of SOD, APX, and POD in roots compared to plant responses prior to heat stress. However, no apparent differences were observed across treatments. Both SOD and POD showed different isoform patterns between roots and shoots, suggesting the function of these isoforms could be tissue specific. Interestingly, no CK effects on these antioxidant parameters were found in this experiment. These results demonstrate the impacts of N on antioxidant metabolism of creeping bentgrass under heat stress with some differences between roots and shoots, but no simultaneous impacts of CK and N.     

Thermotolerance and antioxidant systems in Agrostis stolonifera: involvement of salicylic acid, abscisic acid, calcium, hydrogen peroxide, and ethylene

This study investigated whether pre-treating plants with specific putative signaling components and heat acclimation would induce tolerance of a cool-season grass, creeping bentgrass (Agrostis stolonifera var. palustris), to subsequent heat stress and whether thermotolerance induction of those pretreatments was associated with the regulation of antioxidant regenerating enzymes. The treatments included foliar application of salicylic acid (SA), abscisic acid (ABA), calcium chloride (CaCl2), hydrogen peroxide (H2O2), 1-aminocyclopropane-1-carboxylic acid (ACC, a precursor of ethylene prior to the exposure of plants to heat stress (35 degrees C) in a growth chamber. Physiological measurements including turf quality, leaf photosynthetic rate, and levels of oxidative damage demonstrated that all treatments increased heat tolerance. The better heat tolerance for pre-treated plants as compared to controls was related to the protection of oxidative damage under heat stress. APX activity increased over the first 2 days and 5 days of heating for ACC and CaCl2 respectively, but for only 12 h for H2O2. SA and ABA pre-treatments had no effects on APX activity earlier, but maintained APX activity at a significantly higher level than in controls after 24 h of heating. SA and ABA pre-treatments had no effects on POX activity. ACC treatment significantly increased POX activity. Pre-treatment with CaCl2, H2O2, and HA reduced POX activity, particularly during the later phase of heating. Plants treated with SA, CaCl2, H2O2 and HA had lower CAT activity than their control plants prior to heating and within 48 h of heat stress. ABA and ACC pre-treatments maintained higher CAT activity than the controls after 48 h of heating. ACC, CaCl2, or HA pre-treatments increased SOD activity only before 5 days of heat stress. SA and ABA pre-treatments had less effect on APX activity earlier under heat stress. These results suggest that specific groups of potential signaling molecules may induce tolerance of creeping bentgrass to heat stress by reducing oxidative damage.     

Overexpression of an Arabidopsis β-glucosidase gene enhances drought resistance with dwarf phenotype in creeping bentgrass

An Arabidopsis β-glucosidase, AtBG1 is known to hydrolyze glucose-conjugated, biologically inactive abscisic acid (ABA) to produce active ABA, which increases the level of ABA in plants. Since an increase of ABA in plants confers tolerance against abiotic stress such as drought, we introduced the pCAMBIA3301 vector harboring the AtBG1 gene into creeping bentgrass through Agrobacterium-mediated transformation. After transformation, putative transgenic plants were selected using the BASTA resistance assay at a concentration of 0.8?%. Genomic integration of the AtBG1 gene was confirmed by genomic PCR and Southern blot analysis, and gene expression was validated by Northern blot and Western blot analyses. Interestingly, the transgenic bentgrass plants overexpressing AtBG1 had a dwarf phenotype with reduced growth rates when compared to wild-type creeping bentgrass. In addition, the transgenic plants accumulated higher ABA levels and displayed enhanced drought tolerance. These results suggest that the expression of AtBG1 in plants induces the accumulation of higher ABA levels, which results in the formation of dwarf creeping bentgrass and enhances the survival in water-limiting environments. Key message We used an Arabidopsis β-glucosidase AtBG1 to engineer a crop with elevated active ABA levels, and developed transgenic creeping bentgrass with enhanced drought tolerance and dwarf phenotype.     

Functional significance of the pentose phosphate pathway and glutathione reductase in the antioxidant defenses of human sperm

Glutathione peroxidase is one of the principal antioxidant defense enzymes in human spermatozoa, but it requires oxidized glutathione to be reduced by glutathione reductase using NADPH generated in the pentose phosphate pathway. We investigated whether flux through the pentose phosphate pathway would increase in response to oxidative stress and whether glutathione reductase was required to protect sperm from oxidative damage. Isotopic measurements of the pentose phosphate pathway and glycolytic flux, thiobarbituric acid assay of malondialdehyde for lipid peroxidation, and computer-assisted sperm analysis for sperm motility were assessed in a group of normal, healthy semen donors. Applying moderate oxidative stress to human spermatozoa by adding cumene hydroperoxide, H(2)O(2), or xanthine plus xanthine oxidase or by promoting lipid peroxidation with ascorbate increased flux through the pentose phosphate pathway without changing the glycolytic rate. However, adding higher concentrations of oxidants inhibited both the pentose phosphate pathway and glycolytic flux. At concentrations of 50 microg/ml or greater, the glutathione reductase-inhibitor 1,3-bis-(2-chloroethyl) 1-nitrosourea decreased flux through the pentose phosphate pathway and blocked the response to cumene hydroperoxide. It also increased lipid peroxidation and impaired the survival of motility in sperm incubated under 95% O(2). These data show that the pentose phosphate pathway in human spermatozoa can respond dynamically to oxidative stress and that inhibiting glutathione reductase impairs the ability of sperm to resist lipid peroxidation. We conclude that the glutathione peroxidase-glutathione reductase-pentose phosphate pathway system is functional and provides an effective antioxidant defense in normal human spermatozoa.     

The pentose phosphate pathway: an antioxidant defense and a crossroad in tumor cell fate

The pentose phosphate pathway, one of the main antioxidant cellular defense systems, has been related for a long time almost exclusively to its role as a provider of reducing power and ribose phosphate to the cell. In addition to this "traditional" correlation, in the past years multiple roles have emerged for this metabolic cascade, involving the cell cycle, apoptosis, differentiation, motility, angiogenesis, and the response to anti-tumor therapy. These findings make the pentose phosphate pathway a very interesting target in tumor cells. This review summarizes the latest discoveries relating the activity of the pentose phosphate pathway to various aspects of tumor metabolism, such as cell proliferation and death, tissue invasion, angiogenesis, and resistance to therapy, and discusses the possibility that drugs modulating the pathway could be used as potential tools in tumor therapy.