Cell-cycle dependence of a ganglioside glycosyltransferase activity and its inhibition by enkephalin in a neurotumor cell line

Abstract: Rat glioma mouse neuroblastoma hybrid neurotumor cells (NG108-15), synchronized by amino acid deprivation, showed a cell-cycle-dependent peak of activity of a ganglioside N-acetylgalactosaminyl transferase 14-24 h following release from the cell cycle block (S/G₂ phase). Maximal expression of two typical lysosomal hydrolases, N-acetyl-β-hexosaminidase and β-galactosidase, occurred between 18 and 21 h following release (S phase), declining to G₁ phase levels during the peak of N-acetylgalactosamine (GalNAc) transferase activity. In addition, glycosyltransferase activity in G₂ phase cells showed an increase in apparent V_max (suggesting the presence of more enzyme/mg of cell protein) and apparent binding affinity for uridine diphosphate N-acetylgalactosamine (UDP-GalNAc) (32 versus 14 M) when compared to transferase activity in the G₁ phase. However, the opioid peptide enkephalin D-Ala², o-Leu⁵], which inhibits ganglioside GalNAc transferase activity in unsynchronized NG108-15 cultures, was much more inhibitory in whole cells 8 h after release from the cell cycle block (G₁ phase) than in cells 20 h after release (G, phase), with 50% inhibition occurring at 2 ± 10^-9M and 2 ± 10^-7M, respectively. These results suggest that the GalNAc transferase activity is regulated in more than one way during the cell cycle, since both V_max and K_m changes are observed, and that the cyclic AMP-dependent mechanism by which opiates reduce transferase activity is receptor mediated and cell cycle dependent.