V. V. Talanova

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.

Cold-responsive COR/LEA genes participate in the response of wheat plants to heavy metals stress

28 Under natural conditions, plants are often exposed to the action of two or more stress factors, such as a high temperature and water deficiency, or unfavorable temperature and salinization or heavy metals stress. However, the mechanisms of a combined (simultaa neous or sequential) action of different stress factors on plants still remain poorly studied. It is known that COR/LEA (cold responsive/late embryogenesis abunn dant) genes [1, 2], which encode COR/LEA proteins participating in both cold resistance improvement and protection of cells against dehydration, play an imporr tant role in the lowwtemperature adaptation of plants [3]. Moreover, the expression of many of the LEA genes and the synthesis of the corresponding proteins in plants are enhanced under the influence of the drought [4] and salinization [5]. At the same time, there is no information concerning the possible particc ipation of COR/LEA genes in the plant response to heavy metals stress. It is known that abscisic acid (ABA), being a phytohormone, participates in the regg ulation of the expression of many of the COR genes [2]. For example, ABAAdependent pathways of the loww temperature signal transduction and regulation of COR genes (including the WRAB15 and WRAB18 genes) were revealed in wheat [2]. Taking into account this fact, we studied the changes in the level of express sion of WRAB15 and WRAB18 genes induced by a low positive (hardening) temperature and cadmium, one of the most toxic heavy metals, and also by the combii nation of these two stress factors. Our study was the first to demonstrate a positive correlation between the level of expression of the coldd responsive WRAB15 and WRAB18 genes in leaves of wheat seedlings and the resistance of these seedlings to low temperatures, cadmium, and to the combination of these stress factors. Experiments were conducted with winter wheat (Triticum aestivum L.) seedlings of the Moskovskaya 39 cultivar. The seedlings were grown in filter paper rolls moistened with Knops solution supplemented with microelements (pH 6.2), in an environmental chamber at 22°C, a relative humidity of 60–70%, illuu mination of 10 klx, and photoperiod of 14 h. Onee weekkold seedlings were exposed to a low positive temm perature (4°C) or cadmium sulfate (100 µM) or the combination of both factors for 7 days, with other conn ditions remaining unchanged. Cold resistance of seedlings was assessed by the temperature causing the death of 50% of palisade cells in leaves (LT 50) …

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.

Effect of abscisic acid on the resistance of cucumber seedlings to combined exposure to high temperature and chloride

The effect of abscisic acid (ABA) on heat and salt resistance of cucumber seedlings exposed (consecutively or simultaneously) to high temperature and chloride was studied. Exogenous ABA proved to additionally increase the heat and salt resistance after both consecutive and simultaneous exposure of cucumber seedlings to 38∞N and NaCl. The involvement of this hormone in the common (nonspecific) mechanisms increasing plant resistance to the studied environmental factors is concluded.

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.

Effect of Stress Factors on Expression of the Gene Encoding a CBF Transcription Factor in Cucumber Plants

419 Plant responses to various stress factors differing in their nature are related to the complex of similar changes in their cells, including derepression of definite genes [1, 2]. For example, expression of the family of transcription factors CBF/DREB (C-repeated binding factor/dehydration response elements binding protein) in arabidopsis plants is induced by dehydration [3, 4], low temperature [3, 4], and cadmium [5]. In its turn, CBF transcription factors activate expression of some stress-protein genes and thus could be involved in the improving of plant tolerance to unfavorable environmental factors [6, 7]. In this connection, we studied the expression of the CBF transcription factor in cucumber plants under the effects of low and high hardening temperatures, NaCl, and also of the stress hormone, abscisic acid (ABA).

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 …

Effects of abscisic acid treatment on the expression of cysteine proteinase gene and enzyme inhibitor during wheat cold adaptation

The effects of exogenous ABA on the expression of the cysteine proteinase (CP) gene and its inhibitor cystatin (WC3) in the leaves of winter wheat (Triticum aestivum L.) seedlings that were hardened for 7 days at 5°C were studied. Seedling cold-hardiness was judged by the temperature causing death of 50% of palisade cells (LT50) following 5-min freezing at the temperatures from −5 to −10°C. In the initial period of cooling, when seedling cold tolerance increased, the CP and WC3 gene expression increased considerably, but at the highest cold-hardiness the transcript levels of these genes returned to the control level. When ABA was applied exogenously, the CP gene expression grew by the order of magnitude already in 1–5 h from the onset of low temperature treatment and then abruptly dropped, but it was further maintained at a high level. In cold- and ABA-treated seedlings, the WC3 gene expression initially decreased, but it grew considerably in 3–7 days. It is concluded that the development of wheat cold-hardiness is related to ABA-regulated changes in the CP and WC3 gene expression.

Effect of low-temperature hardening on activities of proteolytic enzymes and their inhibitors in the leaves of wheat and cucumber seedlings

On seedlings of winter wheat (Triticum aestivum L.) and cucumber (Cucumis sativus L.), the dynamics of cysteine and serine trypsin-like proteinases and also trypsin inhibitors at cold hardening (5°C for wheat and 10°C for cucumber) was studied. Activation of proteinases and inhibitors coincided in time or preceded an increased tolerance in wheat and cucumber seedlings in the early period of their hardening. After attaining the highest wheat tolerance, activity amidases reduced, whereas the increased activity levels of cysteine proteinases and trypsin inhibitors was maintained during the entire period of hardening. In cucumber, in these period activities of amidases and trypsin inhibitors reduced, whereas the activity of cysteine proteinases was maintained at the level close to the initial one. It is suggested that cysteine proteinases, amidases, and trypsin inhibitors are involved in plant adaptation to cold.

The abscisic acid influence on the gene expression of pro- and anti-apoptotic proteins in the leaves of cucumber plants at low temperatures

We studied the effect of abscisic acid (ABA) on the expression of genes encoding the functional homologues of proapoptotic (CsBAX) and antiapoptotic (CsBI-1) proteins in cucumber leaves at low temperatures. The exposure to a hardening temperature of 12°C caused a slight increase in the level of CsBAX gene transcript and a sharp increase in the CsBI-1 gene transcript. At a damaging temperature of 4°C, the content of the CsBAX gene mRNA significantly increased, whereas the content of the CsBI-1 gene mRNA remained virtually unchanged. The treatment of seedlings with ABA reduced the release of electrolytes from cells and both 12 and 4°C, i.e., increased the cold resistance and simultaneously decreased the level of CsBAX gene transcripts, but drastically increased the content of CsBI-1 gene mRNA.