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.内质网相关降解（ER-associated degradation, ERAD）通路的膜蛋白客户蛋白，需经AAA-ATP酶（AAA-ATPase）p97介导从内质网膜逆转位，以进入蛋白酶体降解途径。在与p97直接结合前，已部分或完全跨膜的客户蛋白跨膜结构域（transmembrane domains, TMDs）需得到持续保护，以避免发生非特异性相互作用。目前尚不清楚客户蛋白的TMD如何从膜上无缝转运至p97。本研究鉴定出定位于内质网的TMUB1，其为一类TMD特异性护送酶。TMUB1可通过其跨膜区与胞质区，分别与膜内的客户蛋白TMD相结合，并结合暴露于膜外的约10至14个疏水残基序列。TMUB1的泛素样结构域（ubiquitin-like domain）可招募p97，后者可将结合于TMUB1的客户蛋白TMD牵拉至胞质中。破坏TMUB1的护送酶活性，会显著稳定内质网膜中原本仅瞬时存在的逆转位中间体，并损害客户蛋白的逆转位过程。因此，TMUB1通过守护客户蛋白TMD从膜至p97的转运过程，促进TMD的分离。