department of biotechnology
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Plant response to drought

In the book Droughts: New Research (NOVA Publishers, 2012) the open access chapter The Response of Plants to Drought Stress: The Role of Dehydrins, Chaperones, Proteases and Protease Inhibitors in Maintaining Cellular Protein Function, reviews the proteins known to be involved in the response of plants to drought stress.

Droughts

Drought, already a threat in Europe, is a stress that has a major effect on the growth, development and, ultimately, life of plants. It not only reduces the yield of affected crops but has also an impact on biodiversity. During evolution plants have developed both physiological and biochemical responses to promote their survival under stress. Understanding these processes is fundamental to improving crop production and for retaining plant resources. Our current knowledge suggests that a number of mechanisms exist by which plants can, within different limits, tolerate drought and recover from its effects. Differential gene expression is among the prerequisites for plant success but ultimately the level of active proteins is decisive. Therefore, besides analysing plant gene expression under different conditions and detecting genes up or down regulated by drought, it is equally important to identify proteins whose levels change.  One consequence of changes in gene expression could be a change in proteolytic activities, which will affect protein breakdown, known to be essential for adaptation to environmental conditions.
Our group is primarily focusing on peptidases, enzymes which cleave peptide bonds in proteins. They can be involved positively in modifying protein content during conditions that require change of metabolic status or removal of damaged proteins, but also negatively in uncontrolled proteolysis resulting from disruption of cellular compartments by water stress. We are investigating two types of plants. One is an important crop, common bean (Phaseolus vulgaris L.), nutritionally the most important legume and, compared to other grain legumes, relatively sensitive to water deficit. It is characterised by a high content of proteins containing essential amino acids. Moreover, it has a low glycemic index and contains very small amounts of fats, no gluten, saturated fatty acids or cholesterol, but significant amounts of antioxidants, which is useful in decreasing the risk of various diseases and has a positive influence on people with a weakened immune system. All this makes common bean an important component of quality diets with great potential as a healthy replacement for meat, and emphasises the importance of effective measures for maintaining and improving its yield and quality. In order to better understand the roles of different components in response of common bean to drought we have [identified and subsequently] chosen for our studies several cultivars of European origin which exhibit different degrees of sensitivity to drought.

 The second plant that we are studying is ramonda (Ramonda serbica Panč.) from the family Gesneriaceae, a rare resurrection flowering plant of the northern hemisphere, which grows as an endemic relict in gorges of the Balkan Peninsula. It belongs to the group of plants that have the unique ability to survive periods of drought in anabiosis, which is characterised by almost total dehydration of cells in their vegetative tissues such as leaves. On rehydration they revive and resume normal functioning, whence the name resurrection or desiccation tolerant plants. They are an important model system, not only for elucidating molecular mechanisms that form the basis of cellular tolerance of water deficit, but also for studying the control of protein breakdown and thus the role of plant peptidases and regulation of their activities during environmental stress conditions. For potentially useful comparisons we are also studying african violet (Saintpaulia ionantha H. Wendl.) which belongs to the same family as ramonda but, although it is remarkably resistant to drought , eventually dies and is thus not desiccation tolerant.

One of our aims is to identify, isolate and characterise peptidases involved in the response of common bean to drought, which are characterised by stress induced changes of their activities that differ in different cultivars. Similarly, we are interested in those peptidases whose activities change in ways that differ in ramonda and african violet. We have adopted this approach in order to select peptidases which may be connected to greater tolerance to drought in the case of bean cultivars and to the specific response of desiccation tolerant plants. is A quantitative zymographic technique, developed in our laboratory, has proved decisive in enabling such peptidases to be selected. . It enables the substrate specificities, susceptibility to specific peptidase inhibitors, relative quantities/activities of individual enzymes and pH profiles of active proteolytic enzymes to be determined in unfractionated leaf tissue extracts.
We have been among the first to show that serine peptidases are involved in the plant response to drought, some of them being related to tolerance to this stress. Results indicate their complex and probably specific roles. So far we have identified and characterized in more detail three serine endopeptidases and five serine aminopeptidases from the Slovene common bean cultivar Zorin, which is moderately tolerant to drought. Their relative activities depend on leaf age and change under the influence of water stress. In general there is very little known about plant serine aminopeptidases and even less about their role, therefore we shall further study their occurrence and function.
In collaboration with the Agricultural Institute of Slovenia we have studied changes in gene expression at different levels of dehydration of common bean. Using differential display reverse transcriptase PCR (DDRT-PCR), confirmed by quantitative real-time PCR, 16 transcripts have been identified whose levels were changed significantly. Five of the genes so identified have not previously been reported as being influenced by drought.

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Selected publications:
  • Hieng, B., Ugrinovič, K., Šuštar-Vozlič, J. in Kidrič, M. (2004). "Different classes of proteases are involved in the response to drought of Phaseolus vulgaris L. cultivars differing in sensitivity." Journal of Plant Physiology 161(5): 519-530.
  • Kavar, T., Maras, M., Kidrič, M., Šuštar-Vozlič, J. in Meglič, V. (2008). "Identification of genes involved in the response of leaves of Phaseolus vulgaris to drought stress." Molecular Breeding 21(2): 159-172.
  • Budič, M., Kidrič, M., Meglič, V. in Cigić, B. (2009). "A quantitative technique for determining proteases and their substrate specificities and pH optima in crude enzyme extracts." Analytical Biochemistry 388(1): 56-62.
    Contact:
    Marjetka Kidrič
    trak ang