There are dramatic differences in processing of glycoprotein N- linked oligosaccharides (OLS) at different stages during development of D. discoideum. Recent studies with wild type and mutant strains have provided strong evidence for a role of two cell surface-derived OLS in intercellular recognition and cohesion during late aggregation. These OLS are not detectable at earlier and later developmental stages. The proposed research will establish the chemical structure of these glycans, using a combination of fast atom bombardment/mass spectrometry in collaboration with Dr. Catherine Costello at M.I.T. and using specific glycosidase treatments. These results will be extended to analysis of structure/function relations by specific degradative treatments to ascertain effects on cohesion inhibition by the OLS and by study of specific mutant strains conditionally defective in both OLS assembly and cohesion. The exact contribution of these OLS to cohesion, in comparison with known glycoproteins implicated in cohesion, will be tested by efforts to inhibit aggregation with combinations of glycans and other perturbing agents, including antibodies. Monoclonal antibodies will be generated by immunization with glycopeptides carrying the OLS, after covalent conjugation to a carrier protein such as BSA. These antibodies will also be used for identification of the in vivo glycoproteins which carry the OLS and for isolation of mutants lacking them and of revertants which have recovered them. %%% A large proportion of the proteins in higher organisms carry complex sugar molecules (oligosaccharides) as part of their structure. These oligosaccharides contribute to the functional properties of the proteins in ways that are not well characterized but are known to be important in cell-cell adhesion. Dr. Henderson has undertaken a structure/function analysis of oligosaccharides known to be involved in the aggregation of cells during development. She is using the simple organism, D. discoideum, as a model system
