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Binding of Monosialoganglioside by Murine Thymus Cells in Vitro¹

Department of Biochemistry, University of Health Sciences/The Chicago Medical School, Chicago, Illinois 60612

Binding of tritium-labeled monosialoganglioside (G_M1) by intact thymocytes of young AKR/J mice was studied in vitro. The binding was dependent on time, temperature, concentration of cells, and ganglioside, suggesting that the cells possess a finite number of G_M1-binding sites. Binding, which could be observed as early as 15 sec, was very rapid, reaching a plateau in about 5 min, followed by a slow and steady increment up to 4 hr of incubation. Cellular binding of G_M1 appeared to be tight with very little exchange with exogenous gangliosides. G_M1 was almost entirely recovered intact 30 min after it was bound by thymus cells. The structural components of G_M1 (galactose, glucose, N-acetylneuraminic acid, mixed ceramides, and lactosylceramide) did not compete with G_M1 for cellular binding. The relative specificity of the G_M1-binding site was revealed by the lack of competition by lactosylceramide and very weak competition by disialoganglioside and trisialoganglioside that appeared to be due to a G_M1 contamination in these preparations. Further characterization of the G_M1-thymocyte interaction indicated that it was a Ca²⁺-dependent process, since chelating agents ethyleneglycolbis(ß-aminoethyl ether)-N,N'-tetraacetic acid and ethylenediaminetetraacetic acid inhibited the binding up to 75 to 80%, and the inhibition by ethyleneglycolbis(ß-aminoethyl ether)-N,N'-tetraacetic acid was reversed by the addition of excess of Ca²⁺. Also, there appeared to be no overlap of the G_M1-binding site with the binding site for concanavalin A. Finally, a comparison of the amount of G_M1 bound by leukemic and nonleukemic thymus cells revealed that the tumor cells have a higher capacity to bind G_M1 molecules than do normal thymus cells on a per cell basis under several experimental conditions.

¹ Supported by USPHS Grant CA 21631 from the National Cancer Institute.

² To whom requests for reprints should be addressed.

Received 11/14/79. Accepted 5/12/80.