Role of dolichyl phosphate, N-linked glycosylation and cell membrane expression of insulin-like growth factor-1 receptor in maintenance of malignant cell growth

Abstract: ROLE OF DOLICHYL PHOSPHATE, N-LINKED GLYCOSYLATION AND CELLMEMBRANE EXPRESSION OF INSULIN-LIKE GROWTH FACTOR-1 RECEPTOR IN MAINTENANCE OF MALIGNANTCELL GROWTH Several studies have demonstrated that mevalonate (MVA) synthesis is requiredfor cell proliferation. The mechanisms underlying MVA-regulated cell growth are stillnot fully understood. One conceivable mechanism for MVA-dependent cell growth isN-linked glycosylation of growth regulatory proteins. It has for example been proposedthat MVA may be important for N-linked glycosylation and function of the insulin-likegrowth factor 1 receptor (IGF-lR). A reduced rate of N-linked glycosylation, inducedby tunicamycin (TM), has been shown to kill malignant cells. The aims of this thesiswere to clarify the role of MVA in regulation of N-linked glycosylation and cellmembrane expression of IGF-lR with special regard to growth control of malignantcells, and to elucidate the mechanisms involved in TM induced cell death. Inhibition of MVA synthesis led to depression of N-linked glycosylation and growtharrest in melanoma cells. This was correlated to a drastic decrease in translocationof IGF-1 R to the cell surface. By stimulating MVA-depleted arrested cells I foundthat IGF-lR expression occured before initiation of DNA synthesis. Using alphaR-3,an antibody blocking the IGF-lR binding domain, it was confirmed that the IGF-lRexpression was necessary for initiation of DNA synthesis. I could also provide evidencethat dolichyl phosphate, by its participation in N-linked glycosylation of IGF-lR,comprises the MVA product involved in cell growth control. Furthermore, my resultssuggest that the low expression of membrane-bound IGF 1 R in estrogen-negative breastcancer cells is due to a low rate of de novo synthesis of dolichyl phosphate andN-linked glycosylation of IGF-lR. Since these cells normally produce functional (ligand-binding)IGF-lR and sythesize IGF-1 my data raise the possibilty that dolichyl phosphate maybe important for whether breast cancer cells use an extracellular or intracellularautocrine IGF-1 pathway. Inhibition of N-linked glycosylation with tunicamycin induced apoptosis in melanomacells and virus-transformed fibroblasts. In melanoma cells this effect was due toa down regulation of IGF-lR at the cell surface. In SV40-transformed cells the apoptoticeffect, which was mediated through very rapid mechanisms, was related to an increasein cytoplasmic Ca2+ ISBN 91-628-2751-0