Phase separation-based visualization of protein-protein interactions and kinase activities in plants

Safi, A.; Smagghe, W.; Goncalves, A.; Xu, K.; Fernandez, A. I.; Cappe, B.; Riquet, F.; Mylle, E.; Van Damme, D.; Geelen, D.; De Jaeger, G.; Beeckman, T.; Van Leene, J.; Vanneste, S.

doi:10.1101/2022.09.06.506782

bioRxiv · 10.1101/2022.09.06.506782

Phase separation-based visualization of protein-protein interactions and kinase activities in plants

Safi, A.·Smagghe, W.·Goncalves, A.·Xu, K.·Fernandez, A. I.·Cappe, B.·Riquet, F.·Mylle, E.·Van Damme, D.·Geelen, D.·De Jaeger, G.·Beeckman, T.·Van Leene, J.·Vanneste, S.

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Abstract

Protein activities depend heavily on protein complex formation and dynamic post-translational modifications, such as phosphorylation. Their dynamic nature is notoriously difficult to monitor in planta at cellular resolution, often requiring extensive optimization and high-end microscopy. Here, we generated and exploited the SYnthetic Multivalency in PLants (SYMPL)-vector set to study protein-protein interactions (PPIs) and kinase activities in planta based on phase separation. This technology enabled easy detection of inducible, binary and ternary protein-protein interactions among cytoplasmic, nuclear and plasma membrane proteins in plant cells via a robust image-based readout. Moreover, we applied the SYMPL toolbox to develop an in vivo reporter for SnRK1 kinase activity, allowing us to visualize tissue-specific, dynamic SnRK1 activation upon energy deprivation in stable transgenic Arabidopsis plants. The applications of the SYMPL cloning toolbox lay the foundation for the exploration of PPIs, phosphorylation and other post-translational modifications with unprecedented ease and sensitivity.

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Safi, A., Smagghe, W., Goncalves, A., Xu, K., Fernandez, A. I., Cappe, B., Riquet, F., Mylle, E., Van Damme, D., Geelen, D., De Jaeger, G., Beeckman, T., Van Leene, J., Vanneste, S.. 2022-09-08. Phase separation-based visualization of protein-protein interactions and kinase activities in plants. https://doi.org/10.1101/2022.09.06.506782

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