Lichens are extremophilic symbiotic associations possessing phenomenal resistance to abiotic stress-factors. In this regard, melanization of thalli in response to UV is one of the mechanisms protecting lichens from excessive insolation. However, microstructure and biochemical properties of the melanized thalli are still poorly investigated. In the present study, morphological, nanomechanical, and physiological, and biochemical traits of naturally melanized thalli of the Cetraria islandica (L.) Ach. lichen were examined. In the upper cortex of its thallus, the nature of the pigment layer was verified using typical qualitative reactions for melanins. It was found that melanization leads to changes in microstructure of the upper cortex of the mycobiont, in particular, thickening of the cell walls and extension of the interhyphal space. The melanized and pale (nonmelanized) thalli were found to differ from each other in their nanomechanical properties, including the parameters of adhesion and rigidity. This implies the possible formation of complex associates of melanin with cell wall components in the melanized mycobiont. In addition, higher antioxidant activity and lower respiratory activity were found in the melanized thalli of C. islandica in comparison with the pale thalli. Presumably, the found modifications in the microstructure and nanomechanical, physiological, and biochemical properties of thalli occurring in the course of melanization make lichens more resistant to intense insolation.

Extremophile organisms can survive under extreme conditions through tolerance mechanisms. Such organisms include lichens, which are symbiotic associations of fungi and algae and/or cyanobacteria. Among other factors, the high stress tolerance of lichens can be attributed to their ability to synthesize a wide range of metabolites, including chemically diverse lipid compounds. Despite their obvious relevance, the biochemical mechanisms of stress tolerance in lichens that are mediated by changes in the lipid profile remain understudied. Peltigerous lichens constitute a separate division of lichens characterized by high growth rates and metabolic activity indices. Taking into account the temperate climate in which these lichens grow, it can be assumed that they may be highly sensitive to temperature fluctuations. These factors led to the choice of Peltigera canina, epigeic lichen, as the subject matter of this study. The present work examined stress-induced changes in the redox status of P. canina, as well as changes in its lipid composition at an elevated temperature (40 °С). The exposure of hydrated lichen thalli to an unfavorable temperature led to an increase in the level of hydrogen peroxide, phenoloxidase activity, and lipid peroxidation, which constitutes an important part of stress response in lichen. The stress-induced changes in the redox status of lichen thalli stimulated an increase in lipophilic antioxidant agents: a dramatic increase in the level of α-tocopherol and an increase in carotenoids, specifically β-carotene. Thus, the accumulation of lipophilic antioxidants constitutes an important part of the lipid-mediated stress response of P. canina to temperature elevation.

Peroxidases are widespread in animal and plant tissues, wherein they perform a variety of functions. Peroxidases have a broad specificity for substrates of various chemical structures. Along with hydrogen peroxide, phenolic compounds, and toxic compounds of aromatic nature, nitrogen-containing compounds are substrates for peroxidases. This work is devoted to the study of the role of wheat extracellular peroxidases in the metabolism of nitrogen-containing compounds. It has been shown that partially purified isozymes differing in peroxidase activity are involved in the metabolism of nitrogen-containing compounds. The formation of primary and secondary phenoxyl radicals during the combined oxidation of chlorogenic acid, nitrite, and H2O2 was demonstrated. With cooxidation with purified isoenzymes p˗coumaric acid and nitrite, the formation of 4˗hydroxy˗3˗nitrocinnamic acid was revealed. It is assumed that the same isoforms can participate both in the oxidation of nitrite with the formation of nitrophenol and in the reduction of nitrate. The participation of plant peroxidases in nitrogen metabolism can be represented as a set of reactions for the reduction and/or oxidation of nitrogen of different oxidation states with the formation of active intermediates.

Ascorbate peroxidase (APX) is an important antioxidant enzyme responsible for the conversion of H2O2 to H2O and O-2. In this study, APX was studied in the widespread boreal cushion moss Dicranum scoparium. Native PAGE of crude extracts of moss thalli revealed the presence of APX isoforms in D. scoparium with a range of molecular masses. An APX complementary DNA (cDNA) gene of 771 -bp length was cloned and designated as DsAPX. The cloned coding domain sequence (CDS) encoded a 256 amino acid polypeptide, and the predicted protein product was calculated to have a molecular mass of 28.4 kDa with an isoelectric point of 5.7. Several highly conserved sites important for enzyme activity were predicted using bioinformatic tools. The DsAPX protein has similarities of 91%, 78%, 67% and 66% with APX homologs from Grimmia pilifera, Physcomitrella patens, Zea mays and Nicotiana tomentosiformis, respectively. The high homology with cytosolic APX from G. pilifera strongly suggests that the cloned DsAPX gene encodes a cytosolic APX. We studied the role of APX in the tolerance of this moss to abiotic stresses. Changes in both APX activity and DsAPX gene expression, estimated using reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR), showed that the gene was up-regulated in response to desiccation/rehydration, and expression was maintained during a heat stress of +50 degrees C. By contrast, DsAPX was down-regulated by a freezing (-20 degrees C) treatment. Results obtained extend our knowledge of the diversity of APX isoforms that occur in bryophytes and suggest that APX may be involved in the tolerance of D. scoparium to abiotic stresses.

Одним из ключевых факторов устойчивости лишайников к неблагоприятным условиям являются редокс-ферменты, которые участвуют в образовании меланина, защитного пигмента, синтезируемого микобионтом.В талломах лишайника Lobaria pulmonaria (L.) Hoffm.(Лобария легочная) с разной степенью меланизации выявлена активность лакказы, пероксидазы и тирозиназы.Определены изоэлектрические точки и молекулярные массы отдельных изоформ лакказы и тирозиназы.Изоферментный спектр этих ферментов в немеланизированных и меланизированных талломах лишайника L. pulmonaria не различался.Частичная очистка осажденных белков L. pulmonaria с помощью анионообменной хроматографии выявила пики активности лакказы и тирозиназы.Результаты 2Dэлектрофоретического разделения белков обнаружили две мажорные изоформы ферментов: 120 кДа с pI 6,6 и 60 кДа с pI 5,9, которые визуализировались при окрашивании гелей субстратами как лакказы, так и тирозиназы.Высказывается предположение, что в лишайнике L. pulmonaria L-DOPA не только является предшественником в реакции образования меланина с участием тирозиназы, но также может быть метаболизирован пероксидазами и лакказами.Обсуждается роль этих ферментов в синтезе меланина, а также генерации активных форм кислорода, участвующих в защите лишайников от патогенов и абиотических стрессоров.

In this paper, stress induced changes in the redox status of moss Dicranum scoparium Hedw.are discussed.It is shown that some peroxidase isoforms of D. scoparium can display both pro-and antioxidative properties.It is found that one peroxidase isoform of D. scoparium possesses both superoxide anion radical producing activity and superoxide dismutase-like activity.It is suggested that peroxidases from bryophytes, similarly to peroxidases from vascular plants, are among the main components of stress response and regulators of cellular redox status.

Министерство науки и высшего образования РФ Российская академия наук Общество физиологов растений России Правительство Республики Татарстан Федеральный исследовательский центр «Казанский научный центр Российской академии наук» Казанский институт биохимии и биофизики ФИЦ КазНЦ РАН Институт физиологии растений им.К.А

Nitrate reductase (NR) and peroxidase (POX) are important enzymes involved in the metabolism of reactive oxygen (ROS) and nitrogen species in leaves of wheat (Triticum aestivum L.) seedlings. It has been confirmed that NR activity in wheat leaves depends on the light conditions and the presence of nitrates during the cultivation of the seedlings, and it is regulated by the molybdenum cofactor and phosphorylation. In the present study, confocal microscopy and EPR spectroscopy studies showed that the addition of nitrite, a product of NR, increased the level of nitric oxide (NO). This increase was prevented by the addition of sodium azide, an inhibitor of NR. The results suggest that in wheat leaves one of the key functions of NR is the formation of the signaling NO molecule. Cultivation of green plants under conditions of prolonged (4 days) darkness, a strong stress factor for photosynthesizing cells, decreased the activity of NR. Moreover, darkness induced significant elevation of the POX activity that was prevented by the addition of nitrate to the growth medium. It is proposed that the changes in light conditions result in the competition between nitrate- and ROS-metabolizing activities of POX in leaves, and a possible interaction between NR and POX controls the levels of NO and ROS in the leaf tissue.

Infiltration of wheat (Triticum aestivum L.) seedling leaves with excess of nitrate, nitrite, or the NO donor sodium nitroprusside leads to increase both in content of hydroperoxide and activity of peroxidase and decrease in superoxide dismutase (SOD) activity in the leaf apoplast. Polymorphism of extracellular peroxidases and the presence of Cu/Zn-SOD have been shown in apoplast. Using an ESR assay, a considerable increase in the level of NO following infiltration of leaf tissues with nitrite has been demonstrated. These data suggest development of both oxidative and nitrosative stresses in leaves exposed to high levels of nitrate or nitrite. A possible interplay of NO and reactive oxygen species in plant cells is discussed.

Установлено, что при инфильтрации в листья проростков пшеницы (Triticum aestivum L.) избытка нитратa и нитрита, а также донора NO - нитропруссида натрия - в апопласте увеличиваются содержание пероксида водорода и активность экстраклеточных пероксидаз, уменьшается активность супероксиддисмутазы (СОД). Выявлены полиморфизм экстраклеточных пероксидаз и наличие Cu/Zn-содержащей СОД в апопласте. Методом ЭПР показано значительное увеличение образования оксида азота в тканях листьев, обработанных нитритом. Предполагается, что повышенные концентрации нитрата и нитрита в инфильтрирующем растворе вызывают окислительный и нитрозильный стрессы. Обсуждается взаимодействие NO и активных форм кислорода в растительных клетках.

The effect of salicylic acid on the content of soluble proteins and individual polypeptides in Tatar buckwheat Fagopyrum tataricum calluses differing in ability for morphogenesis was studied. Changes in the protein composition of the calluses cultivated in the dark and in the light indicated the higher sensitivity of the non-morphogenic callus. Different response of callus cultures to salicylic acid and conditions of cultivation (light, darkness) is suggested to be associated with the antioxidant defense system, which is, in particular, characterized by the hydrogen peroxide content in the calluses. Salicylic acid increased the H2O2 content in non-morphogenic calluses more strongly than in morphogenic calluses, and the difference was more significant for the calluses cultivated in the light.

The content of soluble proteins and individual polypeptides was studied in calluses of buckwheat Fagopyrum tataricum (L.) Gaertn with different morphogenic potential. The morphogenic callus had a higher content of soluble proteins and cyclic pattern of changes in this index during passaging, which seems to be due to formation of proembryogenic cell complexes. Comparison of the protein patterns of the calluses demonstrated differences in composition and content of individual components. Morphogenic (35 and 73 kDa) and non-morphogenic callus-specific polypeptides (16 and 62 kDa) have been revealed.

The effect of exogenous abscisic acid and cAMP on synthesis of soluble proteins in wheat caryopses in drought has been studied. Both compounds affected the formation of the polypeptides whose synthesis was stimulated by dehydration: they increased the incorporation of the label into polypeptides of 13, 15, and 26 kD and decreased the incorporation of the label into polypeptides of 14, 64, and 77 kD. Abscisic acid and cAMP increased the level of the incorporation of [14C]leucine into the low-molecular-weight polypeptides of 12, 17, and 19 kD whose synthesis was suppressed by drought. These data suggest that the cyclic adenylate signal system is probably involved in the effect of abscisic acid on protein synthesis in drought.