Regulation of the nitrate reductase activity in Symbiodinium sp.

Supervisors: Jean C. Plumier, Stéphane Roberty (Univ. Liege).
Scleractinian corals, the organisms responsible for the coral reef formation, are symbiotic organisms living in symbiosis with the dinoflagellates of the genus Symbiodinium. The algae, thanks to the photosynthesis produced energy with photosynthetic products will be exchanged with the host against nutrients, allowing the reefs formation. These reefs are found in oligotrophic waters, an environment containing few nutrients, and thanks to the photosynthesis theses nutrients could be recycle, especially the nitrogen. The nitrogen is vital for Symbiodinium, allowing to the algae to grow and divided, but its metabolism is still poorly know, especially in Symbiodinium. An enzyme called the nitrate reductase is responsible of the conversion of nitrate into nitrite, its presence in the algae must be verified and the mechanisms of its regulation clarified. The aim of this master thesis was to study the nitrate reductase, in order to better understand how it was regulated. Algae growth rates, mean sizes and chlorophyll contents were measured in various nitrogen source media. The enzymatic activity of the enzyme was tried to be measured because it only happens once in 1977 by Crossland and Barnes without success then. Also, the effect of the nitrogen source composition of the medium was tested thanks to western-blot analysis. Our results shown that when the algae were raised in a nitrogen-free medium their growth rates were low in comparison to the growth rates of algae from medium containing ammonium or nitrate, and that the algae raised in a medium enriched in nitrate presented a high chlorophyll concentration. The enzymatic activity could not be measured due to a maladapted protocol and our results obtained by western blot analyses show that the enzyme was present in the algae depending of the nitrogen source present in the medium. When the algae were even placed in a medium containing both nitrate and ammonium the enzyme was absent which was the opposite for algae from medium containing nitrate. We conclude that when ammonium was present in the medium it was responsible of the regulation of the enzyme. The nitrate reductase seems to be answerable of the synthesis of the nitric oxide, a molecule involved in the bleaching phenomenon. Nowadays, coral reefs worldwide are threatened by bleaching events and the cellular mechanisms leading to this phenomenon are still misunderstood. Therefore, more studies of this enzyme must be further conducted.