Targeted protein degradation through chemical hijacking of E3 ubiquitin ligases is an emerging concept in precision medicine. The ubiquitin code is a critical determinant of the fate of substrates. Although two E3s, CRL2^VHL and CRL4^CRBN, frequently assemble with proteolysis-targeting chimeras (PROTACs) to attach lysine-48 (K48)-linked ubiquitin chains, the diversity of the ubiquitin code used for chemically induced degradation is largely unknown. Here we show that the efficacy of cIAP1-targeting degraders depends on the K63-specific E2 enzyme UBE2N. UBE2N promotes degradation of cIAP1 induced by cIAP1 ligands and subsequent cancer cell apoptosis. Mechanistically, UBE2N-catalyzed K63-linked ubiquitin chains facilitate assembly of highly complex K48/K63 and K11/K48 branched ubiquitin chains, thereby recruiting p97/VCP, UCH37 and the proteasome. Degradation of neo-substrates directed by cIAP1-recruiting PROTACs also depends on UBE2N. These results reveal an unexpected role for K63-linked ubiquitin chains and UBE2N in degrader-induced proteasomal degradation and demonstrate the diversity of the ubiquitin code used for chemical hijacking.

通过化学劫持 E3 泛素连接酶进行靶向蛋白质降解是精准医学中的一个新兴概念。泛素代码是底物命运的关键决定因素。尽管两个 E3 CRL2 ^VHL和 CRL4 ^CRBN经常与蛋白水解靶向嵌合体 (PROTAC) 组装以附着赖氨酸 48 (K48) 连接的泛素链，但用于化学诱导降解的泛素代码的多样性在很大程度上是未知的。在这里，我们表明 cIAP1 靶向降解剂的功效取决于 K63 特异性 E2 酶 UBE2N。UBE2N 促进 cIAP1 配体诱导的 cIAP1 降解以及随后的癌细胞凋亡。从机制上讲，UBE2N 催化的 K63 连接的泛素链促进高度复杂的 K48/K63 和 K11/K48 分支泛素链的组装，从而招募 p97/VCP、UCH37 和蛋白酶体。cIAP1 招募 PROTAC 指导的新底物降解也取决于 UBE2N。这些结果揭示了 K63 连接的泛素链和 UBE2N 在降解剂诱导的蛋白酶体降解中的意外作用，并证明了用于化学劫持的泛素代码的多样性。