Golgi Localization of Carbohydrate Sulfotransferases Is a Determinant of L-selectin Ligand Biosynthesis*
- Departments of ‡Chemistry and ¶Molecular and Cell Biology and the ∥Howard Hughes Medical Institute, University of California, Berkeley, California 94720
- ** To whom correspondence should be addressed: Dept. of Chemistry, University of California, Berkeley, Berkeley, CA 94720. Tel.: 510-643-1682; Fax: 510-643-2628; E-mail: bertozzi{at}cchem.berkeley.edu.
Abstract
Sulfation of endothelial glycoproteins by the sulfotransferase GlcNAc6ST-2 is a regulatory modification that promotes binding of the leukocyte adhesion molecule L-selectin. GlcNAc6ST-2 is a member of a family of related enzymes that act on similar carbohydrate substrates in vitro but discrete glycoproteins in vivo. We demonstrate that GlcNAc6ST-1, -2, and -3 have distinct Golgi distributions, with GlcNAc6ST-1 confined to the trans-Golgi network, GlcNAc6ST-3 confined to the early secretory pathway, and GlcNAc6ST-2 distributed throughout the Golgi. Their localization was correlated with preferred activity on either N-linked or O-linked glycoproteins. A chimera comprising the localization domain of GlcNAc6ST-1 fused to the catalytic domain of GlcNAc6ST-2 was confined to the trans-Golgi network and adopted the substrate preference of GlcNAc6ST-1. We propose a model in which Golgi enzyme localization and competition orchestrate the biosynthesis of L-selectin ligands.
Footnotes
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↵1 The abbreviations used in this paper are: ER, endoplasmic reticulum; BFA, brefeldin A; BSA, bovine serum albumin; CGN, cis-Golgi network; CHO, Chinese hamster ovary; Core1-β3GlcNAcT, Core 1 GlcNAc transferase; Core2GlcNAcT-I, Core 2 GlcNAc transferase I; DAPI, 4,6-diamidino-2-phenylindole; ECFP, enhanced cyan fluorescent protein; ERGIC, endoplasmic reticulum Golgi intermediate compartment; EYFP, enhanced yellow fluorescent protein; FACS, fluorescence-activated cell sorter; FucTVII, fucosyltransferase VII; GFP, green fluorescent protein; GlcNAc6ST, GlcNAc-6-sulfotransferase; GlcNAcT-I, N-acetylglucosaminyltransferase I; HEV, high endothelial venule; LacNAc, N-acetyllactosamine; mAb, monoclonal antibody; MTOC, microtubule organizing center; PBS, phosphate-buffered saline; sLex, sialyl Lewis x; TGN, trans-Golgi network.
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This paper is dedicated to the memory of Frank Rusnak and to Jerry Mohrig on the occasion of his retirement.
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↵* This research was supported in part by National Institutes of Health Grant GM59907. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
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↵§ Supported by a National Science Foundation predoctoral fellowship.
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- Received May 12, 2003.
- Revision received July 8, 2003.
- The American Society for Biochemistry and Molecular Biology, Inc.
