Three glutamic acid residues in the cytoplasmic N-terminal tail of long-form GlcAT-P define Golgi-to-ER trafficking.

Abstract

Glucuronyltransferase GlcAT-P is a rate-limiting enzyme involved in the biosynthesis of the Human Natural Killer-1 carbohydrate and is essential for acquiring higher brain functions. Alternative splicing produces two isoforms, short-form GlcAT-P (sGlcAT-P) and long-form GlcAT-P (lGlcAT-P), which share identical peptide sequences except for an additional 13 amino acids (AA) in the cytoplasmic N-terminal tail of lGlcAT-P. Although sGlcAT-P localizes to the Golgi apparatus (GA), where many glycosyltransferases reside, lGlcAT-P is distributed in both the GA and endoplasmic reticulum (ER). However, the mechanisms responsible for this distinct intracellular distribution remain poorly understood. In this study, we explored the role of the 13 AA in the cytoplasmic N-tail of lGlcAT-P in trafficking between the GA and the ER using the Retention Using Selective Hooks system. Our findings revealed that lGlcAT-P undergoes enhanced retrograde trafficking from the GA to the ER, whereas its anterograde trafficking from the ER to the GA remains largely unaffected. In addition, three glutamic acid residues within the 13 AA of lGlcAT-P were identified as crucial for promoting retrograde trafficking. These results suggest that the ER distribution of lGlcAT-P is primarily governed by Golgi-to-ER trafficking regulated by specific sequences in its cytoplasmic N-tail.