A proteoglycan undergoes different modifications en route to the apical and basolateral surfaces of Madin-Darby canine kidney cells

Abstract

We have grown polarized epithelial Madin-Darby canine kidney II (MDCK II) cells on filters in the presence of [35S]sulfate, [ 3H]glucosamine, or [35S]cysteine/[35S] methionine to study proteoglycan (PG) synthesis, sorting, and secretion to the apical and basolateral media. Whereas most of the [35S]sulfate label was recovered in basolateral PGs, the [3H]glucosamine label was predominantly incorporated into the glycosaminoglycan chains of apical PGs, indicating that basolateral PGs are more intensely sulfated than their apical counterparts. Expression of the PG serglycin with a green fluorescent protein tag (SG-GFP) in MDCK II cells produced a protein core secreted 85% apically, which was largely modified by chondroitin sulfate chains. Surprisingly, the 15% of secreted SG-GFP molecules recovered basolaterally were more heavily sulfated and displayed a different sulfation pattern than the apical counterpart. More detailed studies of the differential modification of apically and basolaterally secreted SG-GFP indicate that the protein cores have been designated to apical and basolateral transport platforms before pathway-specific, post-translational modifications have been completed. © 2005 by The American Society for Biochemistry and Molecular Biology, Inc.