Cell Surface Glycopeptides from Human Intestinal Epithelial Cell Lines Derived from Normal Colon and Colon Adenocarcinomas1

The cell surface glycopeptides from an epithelial cell line (CCL 239) derived from normal human colon were compared with those from three cell lines (HCT-8R, HCT-15, and CaCo-2) derived independently from human colonie adenocarcinomas. Cells were incubated with o-[2-3H]mannose or L-[5,6-3H]fucose for 24 h and treated with trypsin to release cell surface compo nents which were then digested exhaustively with Pronase and fractionated on Bio-Gel P-6 before and after treatment with endoj3-A/-acetylglucosaminidase H. The most noticeable difference between the labeled glycopeptides from the tumor and CCL 239 cells was the presence in the former of an endo-|8-A/-acetylglucosaminidase H-resistant high molecular weight glycopeptide fraction which was eluted in the void volume of Bio-Gel P-6. This fraction was obtained with both labeled mannose and fucose as precursors. However, acid hydrolysis of this fraction obtained after incubation with [2-3H]mannose revealed that as much as 60-90% of the radioactivity was recovered as fucose. Analysis of the total glycopeptides (cell surface and cell pellet) obtained after incubation with [2-3H]mannose showed that from 40-45% of the radioactivity in the tumor cells and less than 10% of the radioactivity in the CCL 239 cells was recovered as fucose. After incubation of the HCT-8R cells with o-[1,6-3H]glucosamine and L-[1-14C]fucose, strong acid hydrolysis of the labeled glycopep tide fraction excluded from Bio-Gel P-6 produced 3H-labeled Nacetylglucosamine and W-acetylgalactosamine. This high molec ular weight glycopeptide fraction was susceptible to mild alkaline borohydride reduction, yielding a mixture of labeled oligosaccharides which contained W-acetylgalactosaminitol. Thus, the HCT8R cells are expressing fucosylated mucin-type glycoproteins on their surface.