Expression of nm23/NDP kinase proteins on the cell surface

Abstract

We determined whether proteins encoded by the nm23/nucleoside diphosphate (NDP) kinase gene, a potential metastasis-suppressor gene, are expressed on the cell surface. Monoclonal antibodies (mAb) specific for nm23-H1 or H2 proteins were prepared using the corresponding fusion proteins with glutathione S-transferase (GST) as immunogens. mAb H1-229 was specifically reactive with nm23-H1 protein, whereas mAb H2-439 was specific for nm23-H2 protein in immunoprecipitation and immunoblotting. mAb H1-229 was reactive with most human hematopoietic and some non-hematopoietic cell lines in flow cytometry. On the other hand, mAb H2-439 was reactive with only a limited number of cell lines. Based upon the surface expression of nm23/NDP kinase, cells were classified as nm23-H1+H2-, nm23-H1+H2+ or nm23-H1-H2-. No cell lines with nm23-H1-H2+ were found among those examined. The specificity of flow cytometry analysis was confirmed in the murine myeloma line NS-1 transfected with either the nm23-H1 or H2 genes. Both mAbs were reactive only to NS-1 transfected with the corresponding nm23 genes. Immunoprecipitation and SDS-PAGE analysis identified 20.5- and 18-kDa proteins with mAb H1-229 or H2-439, respectively, in cellular extracts of 125I-surface labeled NS-1 transfected with the corresponding genes. The presence of nm23/NDP kinase on the cell surface indicates an extracellular role for these proteins in addition to their reported intracellular functions.