TY - JOUR T1 - Activity-based probe profiling of RNF12 E3 ubiquitin ligase function in Tonne-Kalscheuer syndrome JF - Life Science Alliance JO - Life Sci. Alliance DO - 10.26508/lsa.202101248 VL - 5 IS - 11 SP - e202101248 AU - Francisco Bustos AU - Sunil Mathur AU - Carmen Espejo-Serrano AU - Rachel Toth AU - C James Hastie AU - Satpal Virdee AU - Greg M Findlay Y1 - 2022/11/01 UR - https://www.life-science-alliance.org/content/5/11/e202101248.abstract N2 - Ubiquitylation enzymes are involved in all aspects of eukaryotic biology and are frequently disrupted in disease. One example is the E3 ubiquitin ligase RNF12/RLIM, which is mutated in the developmental disorder Tønne-Kalscheuer syndrome (TOKAS). RNF12 TOKAS variants largely disrupt catalytic E3 ubiquitin ligase activity, which presents a pressing need to develop approaches to assess the impact of variants on RNF12 activity in patients. Here, we use photocrosslinking activity-based probes (photoABPs) to monitor RNF12 RING E3 ubiquitin ligase activity in normal and pathogenic contexts. We demonstrate that photoABPs undergo UV-induced labelling of RNF12 that is consistent with its RING E3 ligase activity. Furthermore, photoABPs robustly report the impact of RNF12 TOKAS variants on E3 activity, including variants within the RING domain and distal non-RING regulatory elements. Finally, we show that this technology can be rapidly deployed in human pluripotent stem cells. In summary, photoABPs are versatile tools that can directly identify disruptions to RING E3 ubiquitin ligase activity in human disease, thereby providing new insight into pathogenic mechanisms. ER -