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The leaf protein pattern from drought-tolerant and drought-sensitive wheat varieties subjected to severe soil drought but with the possibility for recover from stress was studied by two-dimensional polyacrylamide gel electrophoresis (2D-PAGE). The spots representing Rubisco large subunit (RLS) were identified using polyclonal antibodies against Rubisco and immunoblotting. Some qualitative and quantitative differences in the 2D-PAGE protein map of wheat varieties were revealed under drought conditions. Three days recovery of wheat plants were not enough for restoring RLS quantity to the level of controls after 7 days drought, especially in the drought-sensitive variety Miziya. There are contradictory data in the literature concerning increased or diminished RLS level in drought stressed plants. A comparison of RLS after SDS-PAGE and 2D-PAGE was made. The revealed protein pattern depended on the presence or absence of protease inhibitors in the extraction buffer, on the procedure of extraction, and on the degree of stress.

The antioxidative protection in leaves of four winter wheat (Triticum aestivum L.) varieties with different field drought resistance was studied under severe recoverable soil drought at seedling stage by withholding irrigation for 7 days (57-59% leaf water deficit) followed by rewatering. A 3-fold raise in electrolyte leakage and a sharp increase in proline accumulation corresponded to drought severity. Hydrogen peroxide content and catalase (CAT) activity were maintained low under stress. Peroxidase (GPX) activity increased, whereas superoxide dismutase (SOD) activity only slightly changed. The content of ascorbate and low-molecular thiols diminished under severe drought and was restored in recovery. Malondialdehyde level was not changed significantly in drought-treated plants but raised after re-watering. In recovery CAT activity became significantly higher whereas GPX activity diminished. Three isoforms of SOD, one of catalase and three of GPX were revealed. Proline accumulation had a predominant role in drought response. As for varieties, drought sensitivity or tolerance was not necessarily correlated with differences in the antioxidative response at early vegetative stage.

Four wheat varieties differing in their drought tolerance were subjected to severe but recoverable water stress at seedling stage. Growth parameters, leaf water deficit (WD) and electrolyte leakage (EL) were used to evaluate the stress intensity and the extent of recovery. The physiological response of the varieties was quite similar under severe drought. Leaf protein patterns and levels of some individual proteins relevant to ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) maintenance were studied in control, stressed and recovering plants by electrophoresis and immunoblotting. The bands representing Rubisco large subunit (RLS), N-and C-terminus of RLS, Rubisco activase (RA) and Rubisco binding protein (RBP, cpn 60), as well as the chaperone and proteolytic subunits of the Clp protease complex were identified using polyclonal antibodies. Under drought conditions RLS, Clp proteases and especially RBP were enhanced, whereas the RA band was only slightly affected. The drought tolerant varieties had higher RBP content in the controls and drought treated plants. Its concentration could be a potential marker for drought tolerance.

Changes in chlorophyll content, ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) binding protein (RBP), Rubisco activase (RA), Rubisco large (LS) and small (SS) subunits, and electrolyte leakage were investigated in wheat leaf segments during heat stress (HS) for 1 h and for 24 h at 40 °C in darkness or in light, as well as after recovery from heat stress (HSR) for 24 h at 25 °C in light. The 24-h HS treatment in darkness decreased irreversibly photosynthetic pigments, soluble proteins, RBP, RA, Rubisco LS and SS. An increase in RA and RBP protein contents was observed under 24-h HS and HSR in light. This increase was in accordance with their role as chaperones and the function of RBP as a heat shock protein.Additional key words: high temperature stress, Triticum aestivum L., immunoblotting.

The main objective of the present work was to examine leaf respiratory responses to dehydration and subsequent recovery in three varieties of winter wheat (Triticum aestivum L.) known to differ in their level of drought tolerance. Under dehydration, both total respiration and salicylhydroxamic acid (SHAM)-resistant cytochrome (Cyt) pathway respiration by leaf segments decreased significantly compared with well-watered plants. This decrease was more pronounced in the drought-sensitive Sadovo and Prelom genotypes. In contrast, the KCN-resistant SHAM-sensitive alternative (Alt) pathway became increasingly engaged, and accounted for about 80% of the total respiration. In the drought-tolerant Katya variety, increased contribution of the Alt pathway was accompanied by a slight decrease in Cyt pathway activity. Respiration of isolated leaf mitochondria also showed a variety-specific drought response. Mitochondria from drought-sensitive genotypes had low oxidative phosphorylation efficiency after dehydration and rewatering, whereas the drought-tolerant Katya mitochondria showed higher phosphorylation rates. Morphometric analysis of leaf ultrastructure revealed that mitochondria occupied approximately 7% of the cell area in control plants. Under dehydration, in the drought-sensitive varieties this area was reduced to about 2.0%, whereas in Katya it was around 6.0%. The results are discussed in terms of possible mechanisms underlying variety-specific mitochondrial responses to dehydration.

Field drought studies were performed in order to assess oxidative stress, proteolytic activity and yield loss under natural stress conditions. Flag leaves of two drought-tolerant (Yantar and Zlatitsa) and two drought-sensitive (Miziya and Dobrudjanka) winter wheat varieties were analyzed. Stress intensity was assessed by relative electrolyte leakage and proline accumulation. Senescence progression was followed by loss of chlorophyll and protein. Lipid peroxidation, H 2 O 2 content, activities of superoxide dismutase (SOD), catalase (CAT), and non-specific peroxidase (GPX) isoforms, as well as proteolytic activities were analyzed from heading throughout grain filling. Weakening of membrane integrity and oxidative damage to lipids were more pronounced in the sensitive varieties under field drought. The activities of Fe-and Cu/Zn SOD isoforms decreased in the controls, but remained high in drought-treated plants. The activities of