HERC3 E3 ligase provides an ERAD branch eliminating select membrane proteins

Aberrant proteins located in the endoplasmic reticulum (ER) undergo rapid ubiquitination by multiple ubiquitin (Ub) E3 ligases and are retrotranslocated to the cytosol as part of the ER-associated degradation (ERAD). Despite several ERAD branches involving different Ub E3 ligases, each with distinct substrate specificity, the molecular machinery responsible for these ERAD branches in mammalian cells remains not fully understood. In this study, we have discovered a cytosolic Ub ligase called HERC3, which fulfills a distinct role in facilitating the ERAD of select polytopic membrane proteins. Using a series of multiplex knockdown/knockout experiments, we have demonstrated that HERC3 functions independently of the ER-embedded ubiquitin ligases RNF5 and RNF185 (RNF5/185) to facilitate the ubiquitination, retrotranslocation, and ERAD of misfolded CFTR. Furthermore, HERC3 collaborates with RNF5/185 to enhance the association of UBQLN proteins, thereby augmenting the retrotranslocation and ERAD of misfolded CFTR. While RNF5/185 participates in the ERAD process of both misfolded ABCB1 and CFTR, HERC3 specifically promotes the ERAD of CFTR, likely due to its ability to interact with the less hydrophobic membrane-spanning domains of CFTR. HERC3 may detect exposed transmembrane domains on the cytoplasmic surface of the ER, thereby facilitating the recruitment of UBQLN and subsequently accelerating the ERAD of select polytopic membrane proteins.