Identification of exposed surface glycoproteins in undifferentiated and differentiated mouse N-18 neuroblastoma cells.
A simple method is described that permitted rapid isolation of plasma membranes from mouse N-18 neuroblastoma cells. The purified plasma membranes gave a 10-fold increase in the specific activity of incorporated [3H]fucose over that of the cell homogenate. The specific activities of two other membrane markers, 5'-nucleotidase and alkaline phosphatase, increased 11-fold and 15-fold, respectively. Metabolic labeling with [3H]fucose identified a major fucosyl glycoprotein with apparent molecular weight of 92 000. Three surface labeling methods together with SDS-polyacrylamide gel electrophoresis and fluorography were used to characterize and compare the surface glycoproteins of undifferentiated and differentiated N-18 cells. The galactose oxidase/NaB3H4 method labeled two major galactoproteins (Mr = 52 000, 42 000) in both undifferentiated and differentiated cells. The neuraminidase/galactose oxidase/NaB3H4 method revealed many sialylgalactoproteins. Among them, the 220-kdalton, 150-kdalton and 130-kdalton bands were at least 100% more prominently labeled in the differentiated cells whereas the 76-kdalton and 72-kdalton bands were less prominently labeled in the differentiated cells when compared to their undifferentiated counterparts. The prominently iodinated protein bands in the undifferentiated cells had apparent molecular weights of 130 000, 92 000, 76 000 and 72 000 as compared to 150-, 130-, 92- and 76-kdalton bands in the differentiated cells. The labeling data obtained will enable us to further study the changes of these identified surface glycoproteins, both quantitatively and topologically, during the differentiation of neuroblastoma cells.
D000818:Animals; D002454:Cell Differentiation; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D005643:Fucose; D006023:Glycoproteins / Q000096:biosynthesis*; D008565:Membrane Proteins / Q000096:biosynthesis*; D051379:Mice; D008970:Molecular Weight; D009447:Neuroblastoma / Q000503:physiopathology*; D014316:Tritium
D006023:Glycoproteins; D008565:Membrane Proteins; D014316:Tritium; D005643:Fucose