New approaches to handle old compounds – Studying the ecological function and bioactivity of Microcystins exploiting bio-orthogonal chemistry

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Microcystins (MCs) are cyclic heptapeptides produced by different cyanobacterial genera, e.g. Microcystis. As major toxins produced during freshwater cyanobacterial blooms, they have a significant impact on the environment, and sometimes even threaten animal or human life. Due to their importance, MCs belong to the best studied cyanobacterial toxins. However, the biological role of MCs for the producing organisms as well as their multifactorial mode of action on eukaryotic cells are still not fully understood. The goal of this project is to contribute to the understanding of the biological functions and the mode of action of these important cyanotoxins. To reach this goal, we will work on answering the following questions: To which proteins bind MCs in MC producing cells of Microcystis? Where are MCs localized in the producing cells? To which proteins bind MCs after uptake into the eukaryotic cell besides the targets known to date?We will address these questions using our newly developed “clickable” MCs. These compounds, produced in situ in Microcystis, feature reactive functional groups that can be used for bio-orthogonal crosslinking techniques, so-called “click reactions“. We can covalently bind these MCs to labels such as fluorescent dyes or gold nanoparticles, and also attach them to polymers such as functionalized agarose beads. The major techniques that will be used in this project are pull-down proteomics to identify to which proteins MCs do bind, and electron microscopy to analyze the localization of MCs in the producing cells. The compounds can also be used for “target fishing” to identify binding partners in eukaryotic cells and for fluorescence microscopy to study MC uptake and distribution in cells. This will enable us to study the biological roles of MCs in greater detail than it has been possible in previous studies.