A Potential Endogenous Ligand of Annexin IV in the Exocrine Pancreas: CARBOHYDRATE STRUCTURE OF GP-2, A GLYCOSYLPHOSPHATIDYLINOSITOL-ANCHORED GLYCOPROTEIN OF ZYMOGEN GRANULE MEMBRANES

Abstract

We demonstrated previously that annexins IV, V, and VI, proteins of the calcium/phospholipid-binding annexin family, have glycosaminoglycan binding properties (Ishitsuka, R., Kojima, K., Utsumi, H., Ogawa, H., and Matsumoto, I. (1998) J. Biol. Chem. 273, 9935-9941). In this study, we investigated the endogenous ligands of annexin IV in the exocrine pancreas. Immunohistochemical study of bovine pancreas showed that annexin IV localized in the apical cytoplasmic region of pancreatic acinar cells where zymogen granules are concentrated. Because it is the major component of the zymogen granule membrane, the glycosylphosphatidylinositol-anchored glycoprotein GP-2 was suggested to play a role in apical sorting and secretion of zymogens. We isolated GP-2 from porcine pancreas extract and determined the structure of its N-linked oligosaccharides by two-dimensional mapping. The major carbohydrate structures of porcine GP-2 were trisialo-triantennary and tetrasialo-tetra-antennary complex-type oligosaccharides. Dot-blot assay showed that annexin IV interacts with GP-2 in the presence of calcium and that it recognizes the terminal sialic acid residues linked through alpha2-3 linkages to the carbohydrate of GP-2. Lectin blot assay showed that Maackia amurensis mitogen, a plant lectin specific for the trisaccharide sequence Sia(alpha)2-3Galbeta1-4GlcNAc of N-linked oligosaccharides, has strong affinity for GP-2. Thus, M. amurensis mitogen was used as a specific probe for GP-2 in the histochemical staining of the bovine pancreas. GP-2 was found to localize exclusively in the same apical cytoplasmic region of pancreatic acinar cells as annexin IV does. These results suggest that GP-2 is an endogenous ligand of annexin IV in the exocrine pancreas.