Characterization of the altered oligosaccharide composition of the insulin receptor on neural-derived cells.

ABSTRACT

Typical insulin receptors are present on neuroblastoma cell lines. High affinity binding for insulin was present in membrane preparations from NG108 (a hybrid mouse neuroblastoma-rat glioma) as well as in membranes from SK-N-MC and SK-N-SH, two human neuroblastoma cell lines. Specific [125I]insulin binding was 24.4% for NG108, 16.9% for SK-N-MC and 5.2% for SK-N-SH at membrane protein concentrations of 0.4 mg/ml. IC50 for [125I]insulin binding was 3.4 nM in NG108 membrane preparations and 0.9 nM for SK-N-SH and 1.8 nM in SK-N-MC membranes. Apparent mol. wt. for the alpha subunits (identified by specific immunoprecipitation using the anti-insulin receptor antiserum B10) on SDS PAGE was 134 kDa for NG108; 124 kDa for SK-N-MC and 120 kDa for SK-N-SH. Neuraminidase digestion increased the mobility of the alpha subunit from both NG108 and SK-N-MC receptors to 120 kDa, whereas that from SK-N-SH were unaffected. Endoglycosidase H and endoglycosidase F digestions increased the mobility of the alpha subunits of all 3 cell lines to varying degrees, suggesting the presence of N-linked glycosylation. Insulin induced autophosphorylation of the insulin receptor beta subunit in WGA-purified membranes from all 3 cell lines. In addition, phosphorylation of a protein with an apparent mol. wt. 105 kDa was stimulated by insulin in WGA purified membranes from NG108. Tyrosine-specific kinase activity was present in the membranes from each cell line and was stimulated by insulin in a dose-dependent manner from 10(-9) to 10(-6) M. Proinsulin was about 100 times less potent in stimulating phosphorylation of the artificial substrate poly (Glu, Tyr)41 when compared to insulin in accordance with its lower binding affinity to the insulin receptor. Hexose transport was stimulated by insulin in all 3 cell lines. These results indicate that neuroblastoma cells contain specific insulin receptors and that they may be useful as models for studying the role of insulin in nervous tissue.