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One of the most important issues facing health care is the prevention and treatment of different types of cancer. Among the most frequently encountered types of cancer leading to lethality are lung cancer, breast cancer and colon cancer. The search for new drugs for cancer cells treatment and clarifying the mechanisms of their anti-tumor effect is a scientific challenge and a necessary basis for new more effective methods of cancer treatment. A major role in tumor growth, invasion, angiogenesis and metastasis have different types of proteases whose activities are inhibited by some synthetic drugs. Protease inhibitors of plant origin have the potential to be an alternative or supplement to the treatment with synthetic drugs, but the mechanisms of their anti-tumor effects are poorly understood. In this review the current knowledge of plant protease inhibitors as tumor preventive and suppresive agents, and the perspectives of their use as antitumour drugs are discussed. [ABSTRACT FROM AUTHOR]

In order to evaluate the role of aminopeptidases (APs) in drought response and their potential as protein markers to distinguish between stress tolerant and sensitive varieties, various AP activities were studied in roots and leaves of winter wheat seedlings, subjected to severe but recoverable soil drought stress. Two varieties with contrasting drought tolerance - Yantar (drought tolerant) and Miziya (sensitive) were compared. Activity changes under severe water stress and subsequent recovery were related to changes in the pools of the major redox buffers ascorbate and glutathione, changes in protein profiles and total proteolysis in roots and leaves. Glutathione was responsive to drought both in roots and leaves, with increased total pool and transient rise in the oxidized form; stronger response in the roots of Yantar was observed. The sensitive variety had higher ascorbate content in leaves under stress. Severe drought led to reversible changes in protein profiles and increase in major protease bands in leaves but not in roots. AP activities were partly independent from the predominant endoprotease activities. Highest activities in roots were detected with substrates releasing terminal leucine, lysine and metionine. In stressed leaves AP activities toward most of the substrates increased under drought, without clear differences comparing varieties. Activities tested with Gly-pNA were raised in leaves only in recovery from stress. In roots, the tolerant variety Yantar presented increased AP activities under stress with most of the substrates used except Leu-pNA and Phe-pNA, whereas the sensitive variety Miziya had almost unchanged AP activities. Based on activity profile changes, at least two different AP enzymes should exist in wheat. It remains to be established which activities towards different substrates reflect distinct aminopeptidases. [ABSTRACT FROM AUTHOR]

Proteases and their specific inhibitors are ubiquitously distributed and play a key regulatory role in many biological processes. Gene expression and activity of certain proteases has been shown to increase in Triticum aestivum L. leaves under drought, with a major contribution of cysteine proteases, especially in sensitive wheat varieties. However, little is known about the stress response of protease inhibitors (PIs) and their role in the regulation of intracellular proteolysis. In this study the changes in transcript abundance of some protease inhibitors (belonging to cystatin and serpin classes) were evaluated by semi-quantitative RT-PCR in leaves and roots of winter wheat seedlings from two varieties with differing tolerance. The expression of two cysteine proteases in the same samples was also assessed. The expression of the studied genes was compared in the tolerant variety "Katya" and the more susceptible to water deprivation variety "Sadovo", applying severe but recoverable soil drought. Growth inhibition and stress related parameters confirmed the relatively higher drought sensitivity of variety "Sadovo". Serpin transcript abundance in control roots was higher than in the leaves. An opposite trend was documented for cystatins - the level of their expression was stronger in the non-treated leaves compared to roots. Drought stress inhibited PI expression in roots, while varying effects on the transcript levels were detected in the leaves of water deprived plants. The levels of the two cysteine protease transcripts under drought exhibited organ-specific response - they declined in roots, and increased in leaves. Further detailed studies using more sensitive methods are necessary to evaluate the potential of protease inhibitors as biochemical markers for drought tolerance. [ABSTRACT FROM AUTHOR]

Proteases and their specific inhibitors are ubiquitously distributed and play a key regulatory role in many biological processes. Gene expression and activity of certain proteases has been shown to increase in Triticum aestivum L. leaves under drought, with a major contribution of cysteine proteases, especially in sensitive wheat varieties. However, little is known about the stress response of protease inhibitors (PIs) and their role in the regulation of intracellular proteolysis. In this study the changes in transcript abundance of some protease inhibitors (belonging to cystatin and serpin classes) were evaluated by semi-quantitative RT-PCR in leaves and roots of winter wheat seedlings from two varieties with differing tolerance. The expression of two cysteine proteases in the same samples was also assessed. The expression of the studied genes was compared in the tolerant variety "Katya" and the more susceptible to water deprivation variety "Sadovo", applying severe but recoverable soil drought. Growth inhibition and stress related parameters confirmed the relatively higher drought sensitivity of variety "Sadovo". Serpin transcript abundance in control roots was higher than in the leaves. An opposite trend was documented for cystatins – the level of their expression was stronger in the non-treated leaves compared to roots. Drought stress inhibited PI expression in roots, while varying effects on the transcript levels were detected in the leaves of water deprived plants. The levels of the two cysteine protease transcripts under drought exhibited organ-specific response – they declined in roots, and increased in leaves. Further detailed studies using more sensitive methods are necessary to evaluate the potential of protease inhibitors as biochemical markers for drought tolerance. [ABSTRACT FROM AUTHOR]

Recurrent drought periods of varying duration often cause extensive crop damage and affect wheat production in Southern Europe. This study compares biochemical and ultrastructural responses of four wheat ( Triticum aestivum L.) cultivars to long-term field drought, and their contribution to final grain yield. Gel electrophoresis and immunoblotting analyses combined with transmission electron microscopy and grain yield evaluation were employed to assess drought susceptibility of the wheat cultivars. Two of them behaved as drought-tolerant, the other two presented as drought sensitive. Enhanced degradation of Rubisco large subunit (RLS), Rubisco small subunit (RSS) and Rubisco activase (RA) accompanied by an increased protease activity and reduced levels of heat shock proteins (HSP70) and dehydrins (DHNs) were associated with drought sensitivity. Drought tolerance coincided with relatively stable or increased HSP70 and DHN contents, and unchanged/higher levels of RLS, RSS and RA. Sensitive cultivars were more vulnerable to ultrastructural damages, showing obvious degradation of chloroplast membrane systems and depletion of leaf starch reserves. These drought responses affected yield potential, as tolerant cultivars gave higher yield under intense drought. Thus, our results provide additional insights into the complexity of plant drought responses, identifying multiple interacting traits that may serve as indirect selection criteria for wheat drought tolerance. [ABSTRACT FROM AUTHOR]

Three varieties (cv. Pobeda, Katya and Sadovo) of winter wheat ( Triticum aestivum), differing in their agronomic characteristics, were analysed during progressive soil water stress and recovery at early vegetation stages. Changes in abscisic acid content, SDS–PAGE and immunoblot profiles of proteins that remained soluble upon heating were monitored. Initially higher ABA content in control Pobeda and Katya corresponded to earlier expression of the studied late embryogenesis abundant (LEA) proteins. A combination of higher ABA content, early immunodetection of dehydrins, and a significant increase of WZY2 transcript levels were observed in drought-stressed leaves of the tolerant variety Katya. One-step RT-PCR analyses of some acidic dehydrin genes ( WCOR410b, TADHN) documented their relatively constant high expression levels in leaves under drought stress during early vegetative development. Neutral WZY2 dehydrin, TaLEA2 and TaLEA3 transcripts accumulated gradually with increasing water deficit. Delayed expression of TaLEA2 and TaLEA3 genes was found in the least drought-tolerant wheat, Sadovo. The expression profile of WZY2 revealed two distinct and separate bands, suggesting alternative splicing, which altered as water stress increased. [ABSTRACT FROM AUTHOR]

Transformed tobacco plants expressing a rice cysteine proteinase inhibitor (OC-I) and non-transformed plants were grown in a controlled environment and subjected to various stresses. Two-month-old transformed and non-transformed plants were exposed for 5 days to drought conditions by withholding watering. High temperature (40 °C) was applied additionally at day 6th for 5 h either individually or in combination with drought. All stress treatments were applied under low (150 μmol m−2 s−1 PPFD) and high light intensity (HL) of 1000 μmol m−2 s−1 PPFD to determine if OC-I expression might provide protection under combination of stresses usually existing in nature. Drought stress led to diminution in leaf relative water content, photosynthesis inhibition, decrease in chlorophyll content and accumulation of malondialdehyde and proline. Heat stress alone did not affect the plants significantly, but intensified the effect of drought stress. HL intensity further increased the proline content. OC-I transformed plants grown under low light intensity had significantly higher total superoxide dismutase and guaiacol peroxidase activities as well as their isoforms than non-transformed control plants under non-stress and stress conditions. Catalase activity was not highly affected by OC-I expression. Results indicate that OC-I expression in tobacco plants provides protection of the antioxidative enzymes superoxide dismutase and guaiacol peroxidise under both non-stress and stress conditions. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

Intact chloroplasts were isolated from dark-senescing primary barley ( Hordeum vulgare L.) leaves in order to study selective ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBPCO) degradation by the stromal and membrane fractions. RuBPCO specific degradation was estimated and characterised applying sensitive avidin-biotin ELISA method with non-modified or oxidatively modified biotinylated RuBPCO (BR) as substrates. Distinct proteolytic activities were detected. They differed in ATP and divalent metal ion dependence, protease inhibitory profile, and dynamics in the time-course of dark-induced senescence. The results supported involvement of ATP- and metal ion-dependent serine type proteolytic activity against non-modified BR early in induced senescence and appearance of ATP-independent activity at later stage. Active oxygen-modified BR was degraded by ATP-independent serine-type protease probably containing essential SH-groups and requiring divalent metal ions. [ABSTRACT FROM AUTHOR]

Limited proteolysis of barley ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBPCO) was effected by treatments with trypsin and Staphylococcus aureus strain V8 protease. Treatment of native RuBPCO with proteases resulted in the degradation of the large subunit (LS) of the enzyme. Trypsin cleaved three fragments from the LS but the S. aureus strain V8 protease cleaved only one. The small subunit (SS) was not affected. In the presence of 0.5 % sodium dodecyl sulfate, RuBPCO degraded into several fragments; some of them were fairly stable. Monoclonal antibodies (Mabs) against barley RuBPCO were applied in immunoblotting analysis to distinguish which of the fragments were recognized. All the Mabs recognized the fragments with molecular masses close to those of the LS. Differences among Mabs were observed in the fragments with low molecular mass. [ABSTRACT FROM AUTHOR]