Increased expression of the polypeptide N-acetylgalactosaminyltransferase 6 (GALNT6), an O-glycosyltransferase, has been reported to play a crucial role in mammary carcinogenesis. Here, we demonstrate that GALNT6 O-glycosylates Glucose-regulated protein 78/Binding immunoglobulin protein (GRP78/Bip), a key regulator of the unfolded protein response (UPR), by adding N-acetylgalactosamine (GalNAc), thereby modulating its stability in breast cancer cells. Functional inhibition of either GALNT6 or GRP78/Bip suppressed the proliferation of the luminal-type breast cancer cell line, ZR-75-1, understanding their importance in tumor cell growth. We further found that GRP78/Bip, which is primarily localized in the endoplasmic reticulum (ER), is transported to the Golgi apparatus under the ER stress conditions, where it undergoes O-glycosylation at Thr203 by the Golgi-resident GALNT6. Substitution of Thr203 with alanine inhibited the binding of GRP78/Bip to IRE1, an ER stress sensor, suggesting that the O-glycosylation at Thr203 in GRP78/Bip facilitates sustained activation of the UPR. These findings define the GALNT6-GRP78/Bip axis as a novel mechanism driving persistent UPR activation and tumor cell adaptation to ER stress, offering a potential new therapeutic target for luminal-type breast cancers.

Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.