TMUB1 promotes p97-mediated segregation of hydrophobic transmembrane domains from the endoplasmic reticulum membrane

Membrane protein clients of endoplasmic reticulum (ER)-associated degradation must be retrotranslocated from the ER membrane by the AAA-ATPase p97 for proteasomal degradation. Before direct engagement with p97, client transmembrane domains (TMDs) that have partially or fully crossed the membrane must be constantly shielded to avoid non-native interactions. How client TMDs are seamlessly escorted from the membrane to p97 is unknown. Here, we identified ER-resident TMUB1 as a TMD-specific escortase. TMUB1 interacts with the TMD of clients within the membrane and holds ~10-14 residues of hydrophobic sequence that is exposed out of membrane, using its transmembrane and cytosolic regions, respectively. The ubiquitin-like domain of TMUB1 recruits p97, which can pull client TMDs from bound TMUB1 into the cytosol. Disruption of TMUB1 escortase activity dramatically stabilizes otherwise transient retrotranslocating intermediates in the ER membrane and impairs client retrotranslocation. Thus, TMUB1 promotes TMD segregation by safeguarding TMD movement from the membrane to p97.