An E2-guided E3 Screen Identifies the RNF17-UBE2U Pair as Regulator of the Radiosensitivity, Immunodeficiency, Dysmorphic Features, and Learning Difficulties (RIDDLE) Syndrome Protein RNF168

Abstract Protein ubiquitinationhas emerged as a pivotal regulatory reaction that promotes cellular responses to DNA damage. With a goal to delineate the DNA damagesignal transduction cascade, we systematically analyzed the human E2 ubiquitin- and ubiquitin-like-conjugating enzymes for their ability to mobilize the DNA damagemarker 53BP1 onto ionizing radiation-induced DNA double strand breaks. An RNAi-based screen identified UBE2U as a candidate regulator of chromatin responses at double strand breaks. Further mining of the UBE2U interactomeuncovered its cognateE3 RNF17 as a novel factor that, via the radiosensitivity, immunodeficiency, dysmorphic features, and learning difficulties (RIDDLE) syndrome protein RNF168, enforces DNA damageresponses. Our screen allowed us to uncover new players in the mammalian DNA damageresponse and highlights the instrumental roles of ubiquitinmachineries in promoting cell responses to genotoxic stress.