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03.03.2009

Protein communicates information about stress

During evolution, plants had to learn to cope with stress factors such as drought or high salt concentrations in soil. Biologists working with Professor Dr. Ralf Reski and Dr. Wolfgang Frank at the University of Freiburg, in cooperation with British researchers, have discovered a protein involved in the transduction of information about environmental stress to the nucleus. It appears that this protein has an important role to play in the plant’s ability to recognise stress and initiate counter measures.

As plants moved from the sea to the land about 450 million years ago, they were confronted with numerous problems. They had to find ways to survive dry periods as well as systems that enabled them to grow where salt concentrations in the soil were high. Consequently, the first green plants developed a system of interconnected molecules that perceived environmental stress, information that they then transferred to the nucleus where genes were activated and induced counter reactions. “Seed plants, including the majority of crops, have lost this system over time,” said Professor Dr. Ralf Reski, head of the Department of Plant Biotechnology at the University of Freiburg. “Seed plants no longer require these systems since they have developed specific tissue types to get rid of salt or to store water.” In addition, humans have bred the particular plant varieties that they require under conditions that obviated the plants’ need to protect themselves against environmental stress. Genes evolved that no longer encoded for such stress tolerance systems. This is especially fatal in the present climate as “the effects of global climate change and intensive agriculture, leading to droughts and the salinisation of vast farmland areas, amongst other things, are becoming more and more apparent,” said Reski. “And this will only become worse in the future.”

A “communicative” plant

The moss Physcomitrella patens is used by researchers in the Department of Plant Biotechnology at the University of Freiburg as model organism for the investigation of mechanisms that protect themselves in stressful situations
The moss Physcomitrella patens is used by researchers in the Department of Plant Biotechnology at the University of Freiburg as model organism for the investigation of mechanisms that protect themselves in stressful situations  (© Prof. Dr. Ralf Reski)

Is it possible to "teach" plants how to deal with stress? If this is the case, the moss Physcomitrella patens may well be a good teacher. The moss, the model organism used by Professor Reski and his team, does not have so many specialist tissue types as seed plants. Mosses were the first green organisms to colonise the land. They are so-called generalists, meaning that they are able to grow reasonably well in any environment. "We found out that the moss Physcomitrella patens is able to effectively protect itself against abiotic stress, for example drought and high salt concentrations," said Dr. Wolfgang Frank, head of a group of researchers in Reski's department. All moss cells can detect changes in salt concentration and pass this information on to their nuclei and other cells. Their latest investigations have provided the researchers with greater insights into the signalling pathways that enable the moss to pass on information about stress.

“It has been known for a while that calcium ions play an important part in signal transduction,” said Frank. “Levels of calcium ions can change very quickly within the cells and these rhythmic changes may occur in particular patterns that encode this information.” Calcium ion signals can be propagated as waves of Ca2+ release and uptake and specific information is encoded in the amplitude and frequency of these oscillations within the cytosol. Amplitude and frequency depend on salt levels, drought or other stress factors. "This is a kind of language," said Reski. "The moss cells perceive external stimuli, which are then translated into different calcium wave patterns." This information is transferred to the nuclei where it induces different genetic programmes. It has also been known for a while that molecular pumps in the cell wall and in the membranes of storage organs in the cytoplasm regularly pump calcium ions into the cell in stressful situations. But what causes the calcium concentrations to decrease again?

A pump and a storage organ

“In order to find factors related to these processes, we attempted to find out which moss genes were switched on very quickly following the exposure to stress,” explained Frank. The DNA sequence of one of these genes was known. This particular gene encoded an ion pump. Frank and his team found that when they switched off the gene the moss plants lost their tolerance to high salt concentrations. If the researchers used dyes which bound to calcium and whose brightness depended on the concentration of calcium, the intensity of the dye did not oscillate. This suggested that the calcium concentration did neither decrease nor increase. In such mutants, the calcium concentration initially increased, and then stayed at the elevated level (calcium “spike”). A protein that was central to the decrease of calcium in the cell therefore had to exist.
Wild-type plants can tolerate high salt concentrations. Mutant moss plants which had the calcium ATPase switched off lose their green colour and die.
Wild-type plants can tolerate high salt concentrations. Mutant moss plants which had the calcium ATPase switched off lose their green colour and die. (© Prof. Dr. Ralf Reski)
By coupling the calcium ATPase gene to the GFP gene (green fluorescent protein), the researchers identified the position of the ion pump in the moss cell. Chloroplasts (red), membranes of the small ATPase-containing vacuoles (green).
By coupling the calcium ATPase gene to the GFP gene (green fluorescent protein), the researchers identified the position of the ion pump in the moss cell. Chloroplasts (red), membranes of the small ATPase-containing vacuoles (green). (© Prof. Dr. Ralf Reski)

"The gene we found encodes a calcium ATPase enzyme," said Frank. "This protein is located on the membranes of small vacuoles and pumps excess calcium ions from the cytoplasm into the small vacuoles - the storage tanks - of the moss cells. Vacuoles are small cell organelles that are present in each moss cell. Moss cells also have a big vacuole. The function of these storage tanks was previously unknown. The researchers were able to show that the calcium ATPases are located between the storage vacuoles and the cytoplasm by coupling the ATPase gene with a GFP gene. The cells then produced a fusion protein from these two proteins. Under the microscope, the membranes of the vacuoles fluoresced and revealed the position of the ATPase.

"Our work shows for the first time how calcium patterns in the cell develop, in particular the low calcium concentrations," said Reski. The researchers at the University of Freiburg now have a better understanding of how mosses transmit signals relating to environmental stress. Seed plants such as rice may also have a rudimentary "stress signalling language". The researchers now hope to put their knowledge to good use on rice and other crops, with the aim of making them less susceptible to high salt concentrations.

A contribution from:
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mn - 03.03.2009
© BIOPRO Baden-Württemberg GmbH

Further information:

Professor Dr. Ralf Reski
Department of Plant Biotechnology
Faculty of Biology
University of Freiburg
Schänzlestrasse 1
79104 Freiburg
Tel.: +49-(0)761/203-6969
Fax: +49-(0)761/203-6967
E-Mail: ralf.reski(at)biologie.uni-freiburg.de

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