Miren Alberdi

MANGANESE AS ESSENTIAL AND TOXIC ELEMENT FOR PLANTS: TRANSPORT, ACCUMULATION AND RESISTANCE MECHANISMS

Manganese is an essential element for plants, intervening in several metabolic processes, mainly in photosynthesis and as an enzyme antioxidant-cofactor. Nevertheless, an excess of this micronutrient is toxic for plants. Mn phytotoxicity is manifested in a reduction of biomass and photosynthesis, and biochemical disorders such as oxidative stress. Some studies on Mn toxicity and Mn translocation from soil to plant cells in Mn2+ form have demonstrated their importance under low pH and redox potential conditions in the soil. When Mn is inside the cells, mechanisms that can tolerate this toxicity are also observed, being important the compartmentalization of this metal in different organdies of shoot and leaf plant cells. A key role of antioxidative systems in plants in relation to high Mn amounts has also been reported as a defense mechanism. The purpose of this review is to show the role of Mn as an essential micronutrient and as a toxic element to higher plants as well as to their transport and tolerance mechanisms. The forms and dynamics of this element in soils and the importance of the acidity for this dynamic and availability for plants are also given.

Molecular and physiological strategies to increase aluminum resistance in plants

Aluminum (Al) toxicity is a primary limitation to plant growth on acid soils. Root meristems are the first site for toxic Al accumulation, and therefore inhibition of root elongation is the most evident physiological manifestation of Al toxicity. Plants may resist Al toxicity by avoidance (Al exclusion) and/or tolerance mechanisms (detoxification of Al inside the cells). The Al exclusion involves the exudation of organic acid anions from the root apices, whereas tolerance mechanisms comprise internal Al detoxification by organic acid anions and enhanced scavenging of free oxygen radicals. One of the most important advances in understanding the molecular events associated with the Al exclusion mechanism was the identification of the ALMT1 gene (Al-activated malate transporter) in Triticum aestivum root cells, which codes for a plasma membrane anion channel that allows efflux of organic acid anions, such as malate, citrate or oxalate. On the other hand, the scavenging of free radicals is dependent on the expression of genes involved in antioxidant defenses, such as peroxidases (e.g. in Arabidopsisthaliana and Nicotiana tabacum), catalases (e.g. in Capsicum annuum), and the gene WMnSOD1 from T. aestivum. However, other recent findings show that reactive oxygen species (ROS) induced stress may be due to acidic (low pH) conditions rather than to Al stress. In this review, we summarize recent findings regarding molecular and physiological mechanisms of Al toxicity and resistance in higher plants. Advances have been made in understanding some of the underlying strategies that plants use to cope with Al toxicity. Furthermore, we discuss the physiological and molecular responses to Al toxicity, including genes involved in Al resistance that have been identified and characterized in several plant species. The better understanding of these strategies and mechanisms is essential for improving plant performance in acidic, Al-toxic soils.

Excess manganese differentially inhibits photosystem I versus II in Arabidopsis thaliana

The effects of exposure to increasing manganese concentrations (50–1500 µM) from the start of the experiment on the functional performance of photosystem II (PSII) and photosystem I (PSI) and photosynthetic apparatus composition of Arabidopsis thaliana were compared. In agreement with earlier studies, excess Mn caused minimal changes in the PSII photochemical efficiency measured as Fv/Fm, although the characteristic peak temperature of the S2/3QB – charge recombinations was shifted to lower temperatures at the highest Mn concentration. SDS-PAGE and immunoblot analyses also did not exhibit any significant change in the relative abundance of PSII-associated polypeptides: PSII reaction centre protein D1, Lhcb1 (major light-harvesting protein of LHCII complex), and PsbO (OEC33, a 33kDa protein of the oxygen-evolving complex). In addition, the abundance of Rubisco also did not change with Mn treatments. However, plants grown under excess Mn exhibited increased susceptibility to PSII photoinhibition. In contrast, in vivo measurements of the redox transients of PSI reaction centre (P700) showed a considerable gradual decrease in the extent of P700 photooxidation (P700+) under increased Mn concentrations compared to control. This was accompanied by a slower rate of P700+ re-reduction indicating a downregulation of the PSI-dependent cyclic electron flow. The abundance of PSI reaction centre polypeptides (PsaA and PsaB) in plants under the highest Mn concentration was also significantly lower compared to the control. The results demonstrate for the first time that PSI is the major target of Mn toxicity within the photosynthetic apparatus of Arabidopsis plants. The possible involvement mechanisms of Mn toxicity targeting specifically PSI are discussed.

ANTIOXIDANT COMPOUNDS IN SKIN AND PULP OF FRUITS CHANGE AMONG GENOTYPES AND MATURITY STAGES IN HIGHBUSH BLUEBERRY (Vaccinium corymbosum L.) GROWN IN SOUTHERN CHILE

We evaluated the genotype and maturity effects on antioxidant activity and phenolic compounds of whole, skin and pulp fruits from three highbush blueberry cultivare (cv. Brigitta, cv. Bluegold and cv. Legacy) grown in southern Chile. Total antioxidant activity (TAA) in ripe fruits varied among the cultivare in the order Legacy > Brigitta > Bluegold. We found that TAA in unripe green and fully ripe fruits was high and similar between them, whereas the lowest levels were found in intermediate ripe fruits. The same trend was observed for fruit total phenolic content. This could be attributed to the higher concentrations of phenolic acids (mainly chlorogenic acid) and flavonols (mainly rutin) at immature fruit stages; whereas the high TAA in mature fruits could be explained by the elevated amounts of anthocyanin. All antioxidant compounds were mostly located in the skin. High amounts of delphinidin aglycone were found. HPLC-DAD/MS revealed that the main contents of skin anthocyanins are petunidin-3-glucoside and petunidin-3-arabinnoside followed by malvidin-3-galactoside. It is noticeable that highbush blueberry fruits grown in southern Chile have exceptionally higher antioxidant activity and anthocyanins contents compared with those cultivated in the northern hemisphere.

Non-structural carbohydrates in Deschampsia antarctica Desv. from South Shetland Islands, Maritime Antarctic

Abstract Deschampsia antarctica Desv. (Angiosperm: family Graminaceae) plants were collected from Robert Island, South Shetland Islands, Maritime Antarctic, during February, 1992 and January, 1993, and were extracted with 80% ethanol. Total soluble sugars were analyzed in leaves and roots by colorimetric and HPLC techniques. Compared with other gramineae, the levels of sucrose and fructans were higher. These substances reached their maximum levels by the end of summer. The levels of sucrose and fructans found in February, 1992 were twice the level found in January, 1993. We suggest that the unusually high accumulation of sucrose and fructans may be one of the protective mechanisms against low temperature that has allowed D. antarctica to grow in the Maritime Antarctic.

The role of photochemical quenching and antioxidants in photoprotection of Deschampsia antarctica

Deschampsia antarctica Desv. (Poaceae) is the only grass that grows in the maritime Antarctic. Constant low temperatures and episodes of high light are typical conditions during the growing season at this latitude. These factors enhance the formation of active oxygen species and may cause photoinhibition. Therefore, an efficient mechanism of energy dissipation and / or scavenging of reactive oxygen species (ROS) would contribute to survival in this harsh environment. In this paper, non-acclimated and cold-acclimated D. antarctica were subjected to high light and / or low temperature for 24 h. The contribution of non-photochemical dissipation of excitation light energy and the activities of detoxifying enzymes in the development of resistance to chilling induced photoinhibition were studied by monitoring PSII fluorescence, total soluble antioxidants, and pigments contents and measuring variations in activity of superoxide dismutase (SOD; EC 1.15.1.1), ascorbate peroxidase (APX; EC 1.11.1.11), and glutathione reductase (GR; EC 1.6.4.2). The photochemical efficiency of PSII, measured as Fv / F m, and the yield of PSII electron transport (ΦPSII) both decreased under high light and low temperatures. In contrast, photochemical quenching (qP) in both non-acclimated and cold-acclimated plants remained relatively constant (approximately 0.8) in high-light-treated plants. Unexpectedly, qP was lower (0.55) in cold-acclimated plants exposed to 4°C and low light intensity. Activity of SOD in cold-acclimated plants treated with high light at low temperature showed a sharp peak 2-4 h after the beginning of the experiment. In cold-acclimated plants APX remained high with all treatments. Activity of GR decreased in cold-acclimated plants. Compared with other plants, D. antarctica exhibited high levels of SOD and APX activity. Pigment analyses show that the xanthophyll cycle is operative in this plant. We propose that photochemical quenching and particularly the high level of antioxidants help D. antarctica to resist photoinhibitory conditions. The relatively high antioxidant capacity of D. antarctica may be a determinant for its survival in the harsh Antarctic environment.

Differential photosynthetic and survival responses to soil drought in two evergreen Nothofagus species

We asked if differences in distribution between Nothofagus nitida and N. dombeyi were associated with differences in drought tolerance. Survival, gas exchange and chlorophyll fluorescence were measured on seedlings subjected to a gradual drought. At a predawn leaf water potential (Ψm) of −2.7 MPa, survival of N. nitida was 50%, compared to 100% in N. dombeyi. Under well-watered conditions, the two species displayed similar stomatal conductance (gw) and transpiration (E), but net photosynthesis (A) and instantaneous water-use efficiency (WUEi) were slightly higher in N. nitida. A, E and gw declined in N. nitida along the gradual drought but increased in N. dombeyi at a Ψm between −1.5 and −2.5 MPa, and declined then drastically at a Ψm below < −2.5 MPa. As N. dombeyi was able to maintain A at higher levels despite declining gw, this species displayed significantly increased WUEi at Ψm below −2.5 MPa. Photochemical efficiency of PSII in the light (ΔF/Fmr) and photochemical quenching (qP) were always lower in N. nitida and along with the photochemical efficiency in the dark (Fv/Fm) they declined in both species. Non-photochemical quenching (NPQ) increased slowly in N. dombeyi along with the gradual drought, whilst it decreased in N. nitida. These results show that differences in drought tolerance are in agreement with sorting of Nothofagus species along moisture gradients in south-central Chile.RésuméNous nous sommes demandés si des différences de distribution entre Nothofagus nitida et N. dombeyi sont associées à des différences de tolérance à la sécheresse. La survie, les échanges gazeux et la fluorescence de la chlorophylle ont été mesurés sur de jeunes plants soumis à une sécheresse croissante. Lorsque le potentiel hydrique de base (Ψm) atteignait −2.7 MPa la survie était de 50 et de 100 % pour N. nitida, et N. dombeyi, respectivement. Dans des conditions d’alimentation hydrique suffisante, les deux espèces ont présenté des valeurs voisines de conductance stomatique (gw) et de transpiration (E) mais la photosynthèse nette (A) et l’effitience instantanée d’utilisation de l’eau (WUEi) étaient légèrement plus élevées pour N. nitida. A, E et gw ont diminué pour N. nitida au cours d’une sécheresse croissante mais ont légèrement augmenté pour N. dombeyi pour des valeurs de Ψm comprises entre −1.5 et −2.5 MPa, puis diminué fortement à des valeurs de Ψm inférieures à −2.5 MPa. Par conséquent, N. dombeyi a présenté des valeurs de WUEi plus élevée que N. nitida à des niveaux de Ψm inferieurs à −2.5 MPa. L’effitience photochimique du PSII à la lumière (ΔF/Fm) et le quenching photochimique (qp) étaient toujours inférieurs pour N. nitida. L’effitience photochimique à l’obscurité (Fv/Fm) ainsi que ΔFjFm ont diminué dans les deux espèces. Le quenching non-photochimique (NPQ) a légèrement augmenté pour N. dombeyi avec la sécheresse, alors qu’il diminuait pour N. nitida. Ces résultats montrent que des différences de tolérance à la sécheresse correspondent à la distribution d’espèces de Nothofagus le long de gradients d’humidité dans le centre sud du Chili.

Calcium sulfate ameliorates the effect of aluminum toxicity differentially in genotypes of highbush blueberry (Vaccinium corymbosum L.)

Abstract The effect of gypsum (CaSO 4 ) amendment in the reduction of Al phytotoxicity of blueberry cultivars differing in Al resistance (Legacy and Brigitta, Al-resistant and Bluegold, Al-sensitive) was studied in a Hoagland’s nutrient solution under acidic conditions for 2 weeks. Treatments were: Control (Hoagland solution), 2.5 mM CaSO 4 , 5 mM CaSO 4 , 100 µM Al (AlCl 3 ), 100 µM Al + 2.5 mM CaSO 4 , 100 µM Al + 5 mM CaSO 4 . Physiological, biochemical and chemical features of leaves and roots were determined to establish the amendment efficiency in the reduction of Al toxicity in these cultivars. Results showed that under Al toxicity the three investi-gated cultivars accumulated high Al concentrations in leaves and roots. These con-centrations decreased with CaSO 4 application. Statistically significant interactions among Al in leaves but not in roots ( p =0.719) and cultivars ( p <0.001), were found. The lowest Ca concentration was found in the most Al-sensitive cultivar (Bluegold) and the highest in the more Al-resistant cultivars (Legacy and Brigitta). Among the underlying processes affected by Al stress in these blueberry cultivars the most evident changes were exhibited by the Al-sensitive cultivar Bluegold, where the photosynthetic performance decreased showing a slight recovery in presence of gyp -sum amendment at the end of experiment. Instead, the more Al-resistant cultivar (Legacy) did not change its photosynthetic parameters in presence of the gypsum amendments during the treatment, whereas in Brigitta, only a slight recovery at the end of treatment was evidenced by the gypsum application. Thus, in relation to these parameters the gypsum amendment was efficient in complete recovery from the toxic Al effect in the Al-resistant cultivar Brigitta and a slight recovery of the toxic Al effect in the Al-sensitive cultivar Bluegold. Nonetheless, this amendment is a good alternative to ameliorate Al toxicity in Al-sensitive cultivars and additionally provides a good source of Ca and S.

Ectomycorrhizal fungi enhance nitrogen and phosphorus nutrition of Nothofagus dombeyi under drought conditions by regulating assimilative enzyme activities

Drought stress conditions (DC) reduce plant growth and nutrition, restraining the sustainable reestablishment of Nothofagus dombeyi in temperate south Chilean forest ecosystems. Ectomycorrhizal symbioses have been documented to enhance plant nitrogen (N) and phosphorus (P) uptake under drought, but the regulation of involved assimilative enzymes remains unclear. We studied 1-year-old N. dombeyi (Mirb.) Oerst. plants in association with the ectomycorrhizal fungi Pisolithus tinctorius (Pers.) Coker & Couch. and Descolea antartica Sing. In greenhouse experiments, shoot and root dry weights, mycorrhizal colonization, foliar N and P concentrations, and root enzyme activities [glutamate synthase (glutamine oxoglutarate aminotransferase (GOGAT), EC 1.4.1.13-14), glutamine synthetase (GS, EC 6.3.1.2), glutamate dehydrogenase (GDH, EC 1.4.1.2-4), nitrate reductase (NR, EC 1.6.6.1), and acid phosphomonoesterase (PME, EC 3.1.3.1-2)] were determined as a function of soil-water content. Inoculation of N. dombeyi with P. tinctorius and D. antartica significantly stimulated plant growth and increased plant foliar N and P concentrations, especially under DC. Ectomycorrhizal inoculation increased the activity of all studied enzymes relative to non-mycorrhizal plants under drought. We speculate that GDH is a key enzyme involved in the enhancement of ectomycorrhizal carbon (C) availability by fuelling the tricarboxylic acid (TCA) cycle under conditions of drought-induced carbon deficit. All studied assimilative enzymes of the ectomycorrhizal associations, involved in C, N, and P transfers, are closely interlinked and interdependent. The up-regulation of assimilative enzyme activities by ectomycorrhizal fungal root colonizers acts as a functional mechanism to increase seedling endurance to drought. We insist upon incorporating ectomycorrhizal inoculation in existing Chilean afforestation programs.

Identification and characterization of three novel cold acclimation-responsive genes from the extremophile hair grass Deschampsia antarctica Desv.

Deschampsia antarctica Desv. is the only monocot that thrives in the harsh conditions of the Antarctic Peninsula and represents an invaluable resource for the identification of genes associated with freezing tolerance. In order to identify genes regulated by low temperature, we have initiated a detailed analysis of its gene expression. Preliminary 2-D gels of in vivo-labeled leaf proteins showed qualitative and quantitative differences between cold-acclimated and non-acclimated plants, suggesting differential gene expression. Similarly, cold-acclimation-related transcripts were screened by a differential display method. Of the 38 cDNAs initially identified, three cDNA clones were characterized for their protein encoding, expression pattern, response to several stresses, and for their tissue-specific expression. Northern blot analysis of DaGrx, DaRub1, and DaPyk1 encoding a glutaredoxin, a related-to-ubiquitin protein, and a pyruvate kinase-like protein, respectively, showed a distinct regulation pattern during the cold-acclimation process, and in some cases, their cold response seemed to be tissue specific. All three transcripts seem to be responsive to water stress as their levels were up-regulated with polyethyleneglycol treatment. DaRUB1 and DaPyk1 expression was up-regulated in leaf and crown, but down-regulated in roots from cold-acclimated plants. The significance of these results during the cold-acclimation process will be discussed.