Abstract
Sphingomyelin synthase (SMS) is the key enzyme for cross-talk between bioactive sphingolipids and glycerolipids. In mammals, SMS consists of two isoforms: SMS1 is localized in the Golgi apparatus, whereas SMS2 is localized in both the Golgi and plasma membranes. SMS2 seems to exert cellular functions through protein-protein interactions; however, the existence and functions of quaternary structures of SMS1 and SMS2 remain unclear. Here we demonstrate that both SMS1 and SMS2 form homodimers. The SMSs have six membrane-spanning domains, and the N and C termini of both proteins face the cytosolic side of the Golgi apparatus. Chemical cross-linking and bimolecular fluorescence complementation revealed that the N- and/or C-terminal tails of the SMSs were in close proximity to those of the other SMS in the homodimer. Homodimer formation was significantly decreased by C-terminal truncations, SMS1-ΔC22 and SMS2-ΔC30, indicating that the C-terminal tails of the SMSs are primarily responsible for homodimer formation. Moreover, immunoprecipitation using deletion mutants revealed that the C-terminal tail of SMS2 mainly interacted with the C-terminal tail of its homodimer partner, whereas the C-terminal tail of SMS1 mainly interacted with a site other than the C-terminal tail of its homodimer partner. Interestingly, homodimer formation occurred in the endoplasmic reticulum (ER) membrane before trafficking to the Golgi apparatus. Reduced homodimerization caused by C-terminal truncations of SMSs significantly reduced ER-to-Golgi transport. Our findings suggest that the C-terminal tails of SMSs are involved in homodimer formation, which is required for efficient transport from the ER.