Cell-to-cell heterogeneity in the expression of carbohydrate-based epitopes of a mucin-type glycoprotein on the surface of human mammary carcinoma cells.
A large, O-linked glycoprotein, termed PAS-O, is a major differentiation antigen on the surface of normal lactating breast epithelia and is also found on the surface of many mammary tumors and mammary carcinoma cell lines. A characteristic feature of populations of tumor cells that express PAS-O is the cell-to-cell heterogeneity with respect to the presence or absence of the molecule. In this study, we used the human mammary carcinoma line 734B and a set of six monoclonal antibodies reactive with PAS-O to study the basis of this heterogeneity. Extensive Western blot analysis of antibody binding to PAS-O in milk fat globule membranes and in skim milk revealed that the antibodies all recognized different epitopes of PAS-O. Moreover, the epitopes were destroyed by periodic acid oxidation, demonstrating their oligosaccharide basis. All six monoclonal antibodies stained the 734B cells heterogeneously. In addition, five clones derived from the parent 734B population also exhibited heterogeneity in the expression of each of the epitopes. An analysis of staining of the 734B clones revealed that, in some cases, certain cells within the cloned population stained with one monoclonal antibody but not with another antibody. Significantly, though, when the 734B cells were treated with neuraminidase prior to antibody staining, most of the heterogeneity was eliminated, and all but one of the monoclonal antibodies stained 90-100% of the cells. This increase in cell staining was matched by an increase in PAS-O staining on Western blots. We conclude that heterogeneity in PAS-O expression on 734B cells is due partly to masking of epitopes by sialic acid and a variation (on a cell-to-cell basis) in the extent of PAS-O sialylation.
Moss
L
L
Cell and Molecular Biology Division, Lawrence Berkeley Laboratory, University of California, Berkeley 94720.
Greenwalt
D
D
Cullen
B
B
Dinh
N
N
Ranken
R
R
Parry
G
G
D000911:Antibodies, Monoclonal; D015153:Blotting, Western; D001943:Breast Neoplasms / Q000378:metabolism*; D002450:Cell Communication; D002460:Cell Line; D002999:Clone Cells; D000939:Epitopes / Q000096:biosynthesis*; D005260:Female; D006801:Humans; D008562:Membrane Glycoproteins / Q000096:biosynthesis*; D008854:Microscopy, Electron; D008858:Microscopy, Phase-Contrast; D008970:Molecular Weight; D018396:Mucin-1; D009077:Mucins / Q000096:biosynthesis*; D019158:N-Acetylneuraminic Acid; D012794:Sialic Acids / Q000032:analysis; D013499:Surface Properties
D000911:Antibodies, Monoclonal; D000939:Epitopes; D008562:Membrane Glycoproteins; D018396:Mucin-1; D009077:Mucins; D012794:Sialic Acids; D019158:N-Acetylneuraminic Acid