It’s Time for Some “Site”-Seeing: Novel Tools to Monitor the Ubiquitin Landscape in Arabidopsis thaliana

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Abstract
In the postgenomic era, it is increasingly apparent that the one gene-one function model is not sufficiently broad to fully understand the molecular mechanisms at play within a cell. Numerous levels of complexity, such as protein-protein interactions and posttranslational modifications, were shown to be essential in determining the role, localization, and activity of a protein. A frequently studied modification is ubiquitination, the covalent binding of the small protein modifier ubiquitin to a target protein. Multiple reports provide useful insights into the plant 'ubiquitinome', but mostly at the protein level without comprehensive site identification. Here, we implemented a new technology, ubiquitin combined fractional diagonal chromatography (COFRADIC), for proteome-wide ubiquitination site mapping on Arabidopsis thaliana cell cultures. We identified 3,009 sites on 1,607 proteins, thereby greatly increasing the number of known ubiquitination sites in this model plant. Finally, the Ubiquitination Site tool provided gives access to the obtained ubiquitination sites, not only to consult the ubiquitination status of a given protein, but also to conduct intricate experiments aiming to study the roles of specific ubiquitination events. Together with the antibodies recognizing the ubiquitin remnant motif, ubiquitin COFRADIC represents a powerful tool to resolve the ubiquitination maps of numerous cellular processes in plants.

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