Polypeptide N-acetylgalactosaminyltransferase 18 retains in endoplasmic reticulum depending on its luminal regions interacting with ER resident UGGT1, PLOD3 and LPCAT1.

Mucin-type O-glycosylation is initiated by the polypeptide: N-acetylgalactosaminyltransferase (ppGalNAc-T) family of enzymes, which consists of 20 members in humans. Among them, unlike other ppGalNAc-Ts located in Golgi apparatus, ppGalNAc-T18 distributes primarily in the endoplasmic reticulum (ER) and non-catalytically regulates ER homeostasis and O-glycosylation. Here, we report the mechanism for ppGalNAc-T18 ER localization and the function of each structural domain of ppGalNAc-T18. By using ppGalNAc-T18 truncation mutants, we revealed that the luminal stem region and catalytic domain of ppGalNAc-T18 are essential for ER localization, whereas the lectin domain and N-glycosylation of ppGalNAc-T18 are not required. In the absence of the luminal region (i.e., stem region, catalytic and lectin domains), the conserved Golgi retention motif RKTK within the cytoplasmic tail combined with the transmembrane domain ensure ER export and Golgi retention, as observed for other Golgi resident ppGalNAc-Ts. Results from coimmunoprecipitation assays showed that the luminal region interacts with ER resident proteins UGGT1, PLOD3 and LPCAT1. Furthermore, flow cytometry analysis showed that the entire luminal region is required for the non-catalytic O-GalNAc glycosylation activity of ppGalNAc-T18. The findings reveal a novel subcellular localization mechanism of ppGalNAc-Ts and provide a foundation to further characterize the function of ppGalNAc-T18 in the ER.