UPTAKE AND INCORPORATION OF AN EPITOPE-TAGGED SIALIC-ACID DONOR INTO INTACT RAT-LIVER GOLGI COMPARTMENTS - FUNCTIONAL LOCALIZATION OF SIALYLTRANSFERASE OVERLAPS WITH BETA-GALACTOSYLTRANSFERASE BUT NOT WITH SIALIC-ACID O-ACETYLTRANSFERASE

The transfer of sialic acids (Sia) from CMP-sialic acid (CMP-Sia) to N -linked sugar chains is thought to occur as a final step in their bios ynthesis in the trans portion of the Golgi apparatus. In some cell typ es such Sia residues can have O-acetyl groups added to them. We demons trate here that rat hepatocytes express 9-O-acetylated Sias mainly at the plasma membranes of both apical (bile canalicular) and basolateral (sinusoidal) domains. Golgi fractions also contain 9-O-acetylated Sia s on similar N-linked glycoproteins, indicating that O-acetylation may take place in the Golgi. We show here that CMP-Sia-FITC (with a fluor escein group attached to the Sia) is taken up by isolated intact Golgi compartments. In these preparations, Sia-FITC is transferred to endog enous glycoprotein accepters and can be immunochemically detected in s itu. Addition of unlabeled UDP-Gal enhances Sia-FITC incorporation, in dicating a substantial overlap of beta-galactosyltransferase and sialy ltransferase machineries. Moreover, the same glycoproteins that incor porate Sia-FITC also accept [H-3]galactose from the donor UDP-[H-3]Gal . In contrast, we demonstrate with three different approaches (double- labeling, immunoelectron microscopy, and addition of a diffusible exog enous acceptor) that sialyltransferase and O-acetyltransferase machine ries are much more separated from one another. Thus, 9-O-acetylation o ccurs after the last point of Sia addition in the trans-Golgi network. Indeed, we show that 9-O-acetylated sialoglycoproteins are preferenti ally segregated into a subset of vesicular carriers that concentrate m embrane-bound, but not secretory, proteins.