Antigenic determinants shared by lysosomal proteins of Dictyostelium discoideum. Characterization using monoclonal antibodies and isolation of mutations affecting the determinant.
Polyclonal antisera raised against a single lysosomal enzyme from Dictyostelium discoideum will cross-react with other lysosomal proteins due to the presence of common post-translational modification antigens. We have now isolated hybridoma cell lines secreting monoclonal antibodies that also recognize a shared lysosomal determinant. Although the same proteins are recognized by the monoclonal and polyclonal antibodies, the actual determinants differ. Also, the polyclonal serum contains antibodies which recognize at least one determinant not recognized by the monoclonal antibodies. All of the antigenic proteins are minor cell proteins but represent a majority of the proteins secreted from the cells during axenic growth. Anode-directed nonequilibrium two-dimensional gel electrophoresis of cellular and secreted protein indicates that the antigenic proteins have extremely acidic pI values; moreover, most are both sulfated and phosphorylated. Several sulfated polysaccharides, including dextran sulfate, chondroitin sulfate, and heparin inhibit binding of the monoclonal but not the polyclonal antibodies, indicating that the shared determinant recognized by the monoclonal antibodies may involve sulfated oligosaccharides. As a step in analyzing the function of the post-translational modification of lysosomal enzymes, we have screened a mutagenized population of amoeba for those lacking the antigenic determinant recognized by one of the monoclonal antibodies. Five cell lines have been isolated that completely lack the determinant. They appear to fall into at least three different classes based upon physical properties of the enzymes. One strain lacks any detectable sulfation of glycoproteins. Several of the strains are capable of forming fruiting bodies indicating that the antigenic determinant plays no crucial role in morphogenesis under laboratory conditions.
D000911:Antibodies, Monoclonal; D004023:Dictyostelium / Q000032:analysis* / Q000201:enzymology; D004591:Electrophoresis, Polyacrylamide Gel; D000939:Epitopes / Q000032:analysis*; D005656:Fungal Proteins / Q000032:analysis* / Q000276:immunology; D006867:Hydrolases / Q000032:analysis*; D008247:Lysosomes / Q000032:analysis*; D009154:Mutation
D000911:Antibodies, Monoclonal; D000939:Epitopes; D005656:Fungal Proteins; D006867:Hydrolases