Identification of salicylic acid (SA)- and pathogen-responsive ubiquitin-binding proteins in Arabidopsis thaliana

The ubiquitin-proteasome system (UPS) is a key regulatory mechanism in plant immunity. Target proteins are labelled with ubiquitin chains that form distinct topological structures to modulate their activity and stability. Here, we identified ubiquitin-binding proteins (UBPs) that specifically interact with M1-, K48-, and K63-linked ubiquitin chains in response to SA or infection with the bacterial pathogen Pseudomonas syringae pv. maculicola ES4326 (Psm). Four-week-old plants were sprayed with 0.5 mM SA or inoculated with Psm ES4326, samples were harvested after 24 hours post-treatment. For the M1- and K48-linked ubiquitin chain pulldown assays, monoubiquitin (Ub) was used as a control. For the K63-linked ubiquitin chain pulldown, empty agarose beads (Beads) served as the control. In total three independent biological repeats were collected.