Plant researcher Tobias Sieberer of the Max F. Perutz Laboratories of the University of Vienna works on signal transduction of hormones called strigolactones. Within his search for chemical substances to influence the activity of this pathway, he is establishing a high-throughput approach to test thousands of different chemical compounds. The project is funded by the Vienna Science and Technology Fund (WWTF).
Strigolactones are plant hormones, which were first discovered in crop plants during the infection with the parasitic plant Striga. Only plants which produce strigolactones were infected with the parasite resulting in a significant reduction of vigour. Moreover, this signalling pathway plays also an important role in plants to initiate symbiotic interactions with mycorrhiza fungi to enhance the absorption of nutrients from the soil.
The third hormonal effect known so far is an influence on shoot branching. If strigolactones are inactivated in experiments, the number of branches is increased. A means to manipulate these three known hormonal effects would have a strong potential for agricultural applications. Particularly in countries with food shortage parasite infection might be diminished. Moreover the rate of shoot branching is an important breeding trait, which affects the quantity and quality of crop harvest as well as the technical effort in cultivation techniques.
Searching for a substance with specific effect
"We want to find substances, which block or stimulate strigolactone effects to use it for different purposes", Tobias Sieberer describes the interdisciplinary project funded by the WWTF.
Sieberer and his project partners Gang Dong of the Max F. Perutz Laboratories and Gerhard Ecker of the Department for Medical Chemistry of the University of Vienna selected an innovative approach as yet not well established in academic research in Austria: Virtual and real high-throughput screening of a vast number of known small molecules. Gang Dong is a structural biologist and will reveal the 3D structure of known enzymes of the strigolatone biosynthesis pathway. The protein structures will then be compared to 3D structures of known small molecules in Gerhard Ecker’s virtual database. With this narrowed selection of inhibitor candidates functional experiments are carried out in the model plant Arabidopsis thaliana.
A second more unbiased approach is searching for substances with the direct help of the Arabidopsis plant. “The project allows the purchase of a highly diverse compound library containing over 30.000 molecule classes. We will grow the plants on one of the chemicals each under controlled laboratory conditions", explains the plant researcher. The lab strain of Arabidopsis comprises a reporter gene, which is turned on if the applied substance alters strigolactone levels – the plant starts to fluoresce.
Bacterium helps plant researchers
Also the bacterium Escherichia coli is utilized for the search of strigolactone regulators. An engineered laboratory strain produces beta-carotene, the dye also found in carrots, and thus colonies have an orange colour. With the help of molecular biological methods the researchers bring the proteins for strigolactone biosynthesis into the bacteria. If the proteins are active, the beta-carotene is recognized as substrate and will be degraded resulting in colourless bacterial colonies. Again the researchers then test for different chemical substances. Inhibitor molecules can easily be detected by the orange-coloured colonies as the degradation of beta-carotene is blocked and the dye will accumulate.
Interdisciplinary project important for basic and applied research
The project allows the establishment of the first academic compound screening facility in Austria. In pharmaceutical companies such libraries are routinely used for drug discovery. For scientists from public research institutes the use of such libraries is cost-intensive and results are subjected to complicate patent laws. "Our library will be open for collaboration with interested scientists from the Viennese area", Sieberer illustrates the possibility to use this library for research on additional model organisms. Results of this chemical genetics technique will support basic and applied research. For the strigolactone project this means that discovered inhibitors might be used to enlighten the basic mechanisms of biosynthesis and signalling of the hormone. But also in applied research this might lead to the development of directed shoot branching regulation or impact on the infection rate of plant parasites.
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