Yu. V. Venzhik

Expression of WRKY transcription factor and stress protein genes in wheat plants during cold hardening and ABA treatment

Changes in expression of WRKY transcription factor and stress protein genes (Wcor15, Wrab17, Wrab19, and Wcs120) were studied on wheat (Triticum aestivum L., cv. Moskovskaya 39) seedlings exposed to cold hardening for 7 days at 4°C. The high level of WRKY gene expression was noticed already in 15 min after the beginning of cold treatment, but the expression level lowered during longer treatments. Exogenous ABA (0.1 mM) suppressed the WRKY gene expression. The level of Wcor15 gene expression increased gradually, reaching the peak on the second day, and then decreased. Gene expression of Wrab17 remained elevated throughout the period of cold exposure (7 days), and expression of Wrab19 was promoted within the first two days. Exogenous ABA induced expression of Wcor15, Wrab17, and Wrab19 genes both at cold-hardening (4°C) and normal (22°C) temperatures. A significant increase in Wcs120 gene expression during cold hardening was ABA-independent. It is concluded that the increase of wheat plant resistance at the initial stage of cold hardening is related to gene expression of WRKY transcription factor and of stress proteins (Wcor15, Wrab17, Wrab19, and Wcs120), while the resistance increase during prolonged adaptation is related to gene expression of Wcor15 and Wrab17.

Structural and functional reorganization of the photosynthetic apparatus in adaptation to cold of wheat plants

In seedlings of a cold-resistant wheat variety, the dynamics was studied of the main structural-functional parameters of the photosynthetic apparatus (PSA) and of cold resistance of leaf cells in low-temperature plant adaptation. It has been established that a complex of structural-functional PSA changes takes place in seedling leaves under the influence of cold. As a result, as early as in the first hours of hardening, the formation of chloroplasts begins to occur in mesophyll cells of larger sizes and with a thylakoid system of the “sun type.” Owing to structural and functional readjustment (a change of content of pigments, stabilization of pigment-protein complexes, and enhancement of nonphotochemical quenching of excess energy) in the process of cold adaptation, the rate of photosynthesis stabilizes. It is suggested that the observed structural-functional PSA rearrangement is a necessary condition for formation of increased cold resistance of leaf cells; this, alongside with other physiological-biochemical changes occurring in parallel in cells and tissues of the plants, provides their survival under conditions of low temperatures.

Influence of lowered temperature on the resistance and functional activity of the photosynthetic apparatus of wheat plants

The dynamics of cold resistance and the activity of the photosynthetic apparatus (PSA) of wheat germs at 4°C were studied. It was shown that in the first hours of cold, a certain functional readjustment to the changed conditions takes place in the plant organism. A decrease in the activity of the PSA and cessation of the linear growth of the leaf are registered at this stage along with an increase in resistance, as well as an increase in the coefficient of non-photochemical quenching of the fluorescence of chlorophyll. In one to four days, when resistance reaches its maximum, photosynthesis and the rate of electron transport are stabilized, the chlorophyll content in the lightcollecting complex increases, and the growth recommences. The final stage of adaptation (days 4–7) is characterized not only by the steady level of resistance but also by new functional organization of the PSA, which allows the plants to endure the lowered temperature successfully.

Ultrastructure and functional activity of chloroplasts in wheat leaves under root chilling

The effects of root chilling (2xa0°C; during 1, 5xa0h, 1, 2, 4 and 7xa0days) on the ultrastructure, functional activity of chloroplasts and cold tolerance of leaf cells of wheat (Triticum aestivum L.) were studied. Results indicated that the area of the chloroplasts increased and the number of grana in the chloroplast decreased already within first hours of the experiment. On the 2nd–7th day of the cold treatment, the length of photosynthetic membranes in the chloroplasts increased owing to the membranes of thylakoids in grana. The number of chloroplasts per cell was increased by the end of the experiment. Reduction of electron transport rate and intensification of non-photochemical quenching of chlorophyll fluorescence were observed in the first hours of root chilling. The growth of the leaves slowed in the first day of the treatment and resumed on the second day. Leaf area in the root-chilled plants by the end of the experiment exceeded the initial values by 60xa0%. The significant rise in cold tolerance of leaf cells was detected after 24xa0h of root chilling. After 48xa0h of the treatment, the cold tolerance reached a maximum, and did not change thereafter. It is assumed that most of the observed structural and functional changes are adaptive, and meant to support the photosynthetic function and promote the cold tolerance of the plants.

The effect of low temperature on wheat roots causes rapid changes in chloroplast ultrastructure in wheat leaves

230 Under the natural conditions, the roots and abovee ground parts of plants are always exposed to different temperatures. For instance, the aerial part of a plant can be exposed to cooling or heating, whereas the root system remains under physiologically normal temperr ature and vice versa. Nevertheless, the plant organism responds to the situations of this kind as a highly intee grated system, where all the physiological and bioo chemical processes are strictly coordinated and linked. It is not surprising, therefore, that cooling the roots leads to changes in water metabolism [1], horr monal balance [2], and photosynthetic activity [3] of leaves, as well as to a higher expression of some genes in leaf cells and a higher cold resistance of leaves [4]. One can expect that some structural rearrangements underlie the above changes. However, we do not know about any published evidence supporting this suggess tion. This study has demonstrated that cooling the root system of the winter wheat leads to a whole set of ultraa structural changes in chloroplasts that promote cold resistance of leaves. Sevenndayyold seedlings of the winter wheat (Tritii cum aestivum L.), Moskovskaya variety, were grown using Knops nutrient solution (pH 6.2–6.4) in an artificial climate chamber at a temperature of 22°C, 60–70% relative humidity, 10 klx illumination, and 144h photoperiod. Seedling root system was exposed to cooling for 7 days in a specially designed device [5] at a temperature of 2°C, which is the optimal for conn ditioning this type of plants to the cold. The abovee ground part of seedlings remained under the temperaa ture of 22°C during the same period of time. The cold resistance of leaves was determined from the temperature (LT 50) causing death of 50% palisade cells in leaf cuttings that were exposed to 55min freezz ing in a thermoelectrical TZhRR02//20 microrefrigerr ator (Interm, Russia) [6]. To estimate cell viability, they were examined for cytoplasm coagulation and chloroplast destruction on a LOMO Micmedd2 light microscope (LOMO, Russia). Each experiment was at least thrice repeated with a sixfold biological replicaa tion. For the electron microscopic examination, we used cuttings from the middle of three to five leaf plates, which were fixed at a temperature of 0–4°C with gluu taraldehyde in phosphate buffer (pH 7.2) and then with OsO 4 (2% solution). The material dehydrated in a series of alcohols and acetones was stained with uraa nylacetate and then embedded …

Effect of Root Heating on the Tolerance of Barley Leaf Cells and Ultrastructure of Chloroplasts and Mitochondria

In Nature, unfavorable factors affect frequently not the whole plant but its separate organs (or parts), which however influence functioning of its other organs (parts), which did not experience stress. For example, even short-term heating plant roots could improve leaf cell tolerance to elevated temperature [1‐3] due to various physiological and biochemical changes in the leaf cells [4‐6]. Such data indicate a capability of signal transduction from one plant organs (parts) to others reporting about temperature changes, although the mechanisms of this phenomenon are not completely deciphered and are still the object of discussion. To understand them, it is of importance to establish not only functional (physiological, biochemical, and molecular) but also possible structural changes arising in the plant cells under the local action of temperature. However, only few related studies [7, 8] are known, which do not permit of presenting a general pattern. In this connection, the objective of this work was to examine structural changes in chloroplasts and mitochondria occurring during the development of improved heat tolerance of leaf cells after the action of high hardening temperature on plant roots. Experiments were performed on barley ( Hordeum vulgare L., cv. Otra) seedlings grown in filter paper rolls on the Knop nutrient medium (pH 6.2‐6.4) in the chamber of the phytotron at 25 ° C, a relative humidity of 60‐70%, an illuminance about 10 klx, and a photoperiod of 14 h. The root system of 7-day-old seedlings was heated at 38 ° C for 1, 4, or 24 h in a specially constructed chamber [2] permitting the maintenance of a desired temperature only around the root system; seedling shoots were kept at 25 ° C. Exposure duration and temperature were chosen on the basis of previous studies [9]. The first leaves were used for the assessment of their heat tolerance and the analysis of the organelle ultrastructure. Leaf cell heat tolerance was assessed from a temperature (LT 50 ) killing 50% of palisade cells of excised leaf disks (0.3 cm 2 in area) after their 5-min heating in the water thermostat. A desired temperature in the thermostat was maintained with an accuracy of ± 0.1 K. Cell viability was assessed from the coagulation of the cytoplasm and chloroplast destruction using a light Mikmed-2 microscope ( × 40 objective, LOMO, St. Petersburg). To examine the ultrastructure of chloroplasts and mitochondria, leaf disks were fixed at 0‐4 ° C with 3% glutaraldehyde prepared in phosphate buffer (pH 7.4) and then postfixed in 2% OsO 4 . The material dehydrated in the series of ethanol concentrations and acetone was stained with uranyl acetate and embedded in the mixture of Epoxy resins. The sections of the leaf palisade parenchyma were prepared using an LKB-4800 ultramicrotome, contrasted with lead citrate, and examined with a JEM-100 electron microscope at magnification of 6000‐25000. The tables and figures present mean values from 2‐ 3 independent experiments performed in 5‐10 replicates and their standard errors. The values significant at P ≤ 0.05 are discussed.

Similarities and Differences in Wheat Plant Responses to Low Temperature and Cadmium

A comparative study was performed on the accumulation of biomass, dynamics of indicators of the activity of photosynthetic apparatus, and cold tolerance in the seedlings of frost-tolerance wheat varieties under the effect of a low hardening temperature and cadmium. It was found that the plant responses to the effects of the stressors studied are similar: an increase in cold tolerance of leaves, slowing rates of plant growth and photosynthesis, and increased non-photochemical fluorescence quenching were observed in both cases. At the same time, it was noted that the plant responses to the actions of low temperature and Cd have certain differences associated with the negative effect of Cd on growth and photosynthetic activity.

Features of the photosynthetic apparatus in winter and spring wheat plants with different cold tolerance

It has been shown that winter wheat differs significantly not only in cold tolerance but also in the nature of changes in functional organization of the photosynthetic apparatus. Such changes occur already in the first hours of low temperature action, minimizing the adverse effects of cold on plants. They are of an adaptive nature and, along with others, permit the plants of winter wheat to survive under cold conditions. Accounting for these specific features may be useful for breeding to create cold-resistant varieties, as well for assessing the prospects of their introduction to regions that are characterized by frequent and significant lowering of temperature with long-term nature included in the period of active plant vegetation.

Characteristics of expression of temperature-regulated genes in winter and spring wheat plants during cold adaptation

The characteristics of the expression of genes Wcs120, Wcor15, and Wrab17 during cold (4°C) adaptation, particularly slower changes in the expression of Wcs120 and Wcor15 in spring compared to winter wheat, are revealed in leaves of winter and spring wheat seedlings. It is concluded that the formation of increased cold tolerance both in winter and spring wheat is related to enhanced expression of the studied genes.