The sorting fate of a transmembrane protein in the secretory pathway is intrinsically linked to its sequence and topology. Single-pass transmembrane proteins can adopt a type I topology with a luminal N terminus or a type II topology with a cytosolic N terminus; however, the effect of topology on sorting has long remained unclear. Consensus sorting motifs have been identified in cytosolic and luminal domains, but sorting motifs within transmembrane domains (TMDs) are less understood, though equally important for protein localisation. In this study (Bian et al., 2024), Pierre Cosson and colleagues test TMD sequence requirements for targeting chimeric type I and type II proteins to the endoplasmic reticulum (ER), Golgi or cell surface. Previous studies have found that localisation is partly mediated by the length of the TMD, with long TMDs being targeted to the cell surface, intermediate TMDs being targeted to the Golgi and short TMDs being targeted to the ER. Here, the authors find that these requirements are more complex: type I chimeric proteins fail to efficiently localise to the Golgi even with intermediate TMDs. Furthermore, type II chimeric proteins with short TMDs localise efficiently to the Golgi and poorly to the ER. In both cases, proteins can be relocalised to the cell surface by increasing either the hydrophobicity or the length of the TMD. Overall, these findings suggest a nuanced interplay between TMD sequence and protein topology and provide important new insights into the sorting of single-pass transmembrane proteins.
