Probing kinase proximity interactome modulation upon drug treatment by miniTurboID proximity labelling

In a systematic effort to profile kinase inhibitors, which also act as degraders, we identified multiple hit compounds. These were subjected to detailed mechanism of action elucidation including analysis of the perturbed interactome and its effect on degradation. For our first example, LYN, Src inhibitor 3 showed rapid degradation at nanomolar potencies. With BioID we could identify that two E3 ligases CBL and CBL-B are both required for this degradation mechanism, which we validated orthogonally by CRISPR/Cas9 k.o. For the second example, we identified TAK285 to degrade BLK in a γ-secretase dependent manner. We thus performed a two-pronged BioID approach using both BLK as the bait as well as APH1A, a subunit of the γ-secretase complex. We could identify an interaction with multiple components involved in γ-secretase processing for BLK, namely NCTSN and ADAM10. As a third example, we investigated the proximity interactome of RIPK2, upon inhibitor/degrader treatment.