Abstract
Treatment with comparatively low doses of the 3-hydroxy-3-metylglutaryl coenzyme A (HMG-CoA) reductase inhibitor lovastatin caused growth arrest in both melanoma cells and Ewing's sarcoma (ES) cells. In melanoma cells growth inhibition was correlated with a drastic decrease in N-linked glycosylation of the insulin-like growth factor-1 receptor (IGF-1R), which in turn was followed by a decreased expression of IGF-1 binding sites at the cell surface. These data suggest that IGF-1R constitutes a link between HMG-CoA reductase, which catalyzes the conversion of HMG-CoA to mevalonate (MVA), and cell growth. This hypothesis was supported by the finding that addition of dolichyl phosphate, a product of MVA which functions as a carrier of activated oligosaccharides in glycoprotein assembly, stimulated IGF-1R expression and DNA synthesis in lovastatin-arrested cells. Inhibition of N-linked glycosylation, using tunicamycin, led to apoptotic cell death in both melanoma and ES cells. The same effect was obtained by an antibody ([alpha]IR-3) blocking the binding domain of IGF-1R These data therefore suggest that TM-induced cell death was due to a reduced plasma membrane expression of IGF-1R. As distinguished from melanoma cells, apoptosis of ES cells following ([alpha]IR-3-mediated blocking of IGF-1R was not preceded by growth arrest. Since inhibition of N-linked glycosylation caused both growth arrest and apoptosis in ES cells, it can be concluded that TM-induced growth inhibition is not mediated by the down-regulation of IGF-1R. in these cells. TM treatment also reduced the expression of the EWS/FLI-1 fusion protein, a gene product resulting from the ES-specific t(11;22) translocation. Using antisense oligonucleotides it was shown that the fusion protein is required for ES cell growth. The decreased expression of the fusion protein, which is not a glycoprotein, was due to a destabilization of de novo-synthesized proteins. The mechanisms by which N-linked glycosylation regulates the EWS/FLI-1 fusion protein are unknown but could involve the action of plasma membrane-bound glycoproteins. It was, however, confirmed that the IGF-1 pathway is not involved in this regulation.
