Polysialic acid in human neuroblastoma cells

Prokaryotic-derived probes that specifically detect α-2,8-linked polysialic acid (PSA) units on embryonic neural cell adhesion molecules (N-CAM) were used to show that membrane glycoproteins (GPs) from metastatic human neuroblastoma cells (CHP-134) also contain these unique carbohydrate moieties. This conclusion was based on the following evidence: (1) membranes from CHP-134 cells served as an exogenous acceptor of [14C]NeuNAc units in an E. coli K1 sialyltransferase (ST) assay. The bacterial ST is specific for the transfer of [14C]NeuNAc to exogenous acceptors containing at least 3 sialyl units (DP3); (2) in SDS-PAGE, the [14C]NeuNAc-labeled CHP-134 membranes showed a major peak of radioactivity that was polydisperse. N-CAM shows a similar Mr heterogeneity; (3) treatment of the high Mr CHP-134 product with Endo-N-acetylneuraminidase (Endo-N) released the [14C]NeuNAc label as a DP4. Endo-N is specific for hydrolysing α-2,8-linked PSA chains containing a minimum of 5 sialyl residues; (4) treatment of the DP4 with sialidase converted the label to [14C]NeuNAc, thus proving the tetramer contained sialic acid; (5) CHP-134 cells were labeled in vivo with [3H]GlcN. A glycopeptide fraction representing ca. 1% of the [3H]GlcN incorporated was isolated. Based on Endo-N sensitivity, this glycopeptide contained at least 15-20% of the [3H]GlcN label as PSA. Endo-N digestion of the [3H]-labeled glycopeptide released [3H]-DP4. These results suggest that the surface expression of PSA-containing GPs may be important in neuroblastoma metastasis