Sarah Gabelmann (Rheinland-Pfälzische Technische Universität Kaiserslautern-Landau, RPTU)
Sarah Gabelmann's Master thesis was awarded with the Prize for the Best Plant Science Master Thesis, which was carried out at Rheinland-Pfälzische Technische Universität Kaiserslautern-Landau in the year 2024 with the title:
Mapping of the nuclear proteome in Chlamydomonas reinhardtii using TurboID-mediated proximity labeling
For the first time, a proximity labeling approach was used to characterize the nuclear proteome of the green alga Chlamydomonas reinhardtii under different conditions, leading to the identification of proteins potentially involved in the stress response to increased H2O2 production.
Nuclear proteins like transcription factors and other transcriptional regulators play an essential role in the regulation of gene expression, a complex process which controls the activities of the cell and allows it to acclimate to external stimuli and stress conditions. To analyze the protein composition of the nucleus under different conditions, a proximity labeling method that has only recently been established in Chlamydomonas reinhardtii was employed. This method utilizes TurboID, a variant of the Escherichia coli biotin ligase BirA, to activate biotin which then covalently attaches to amines of close-by proteins, allowing for the capturing of protein–protein interactions and mapping of proteomes in the living cell. Constructs driving the production of a nuclear-localized TurboID bait and a cytosolic TurboID control bait were generated and transformed into C. reinhardtii. After the functionality of the baits in the two compartments was confirmed by immunoblot analysis, in vivo proximity labeling experiments with externally added biotin were carried out. Mass spectrometry analysis of the purified biotinylated protein fraction revealed 852, 1,160 and 835 proteins significantly enriched for the nuclear bait under continuous light, heat stress and elevated H2O2 levels triggered by paraquat treatment, respectively. Furthermore, when the paraquat treatment was compared to the continuous light control condition, 20 proteins were found to be significantly enriched in the nucleus under increased H2O2 levels. These proteins might be involved in the response to oxidative stress, although further analysis is required to specify their role in the nucleus.
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Sarah Gabelmann conducted this work in the Biology department in the working group Molecular Biotechnology and Systems Biology of Prof. Dr. Michael Schroda.
