NG108-15 cells express neuregulin that induces AChR α-subunit synthesis in cultured myotubes

Abstract

A cholinergic neuroblastoma×glioma hybrid cell line NG108-15 is able to form functional synapses, and contains both AChR-aggregating and AChR-inducing activities when co-cultured with myotubes. Several lines of evidence indicate that the AChR-inducing activity of NG108-15 cells is derived from neuregulin. The conditioned medium of cultured NG108-15 cells induced the expression of AChR α-subunit as well as the tyrosine phosphorylation of erbB-3 receptor. NG108-15 cells expressed neuregulin with a protein of ∼100 kDa in size and transcripts of ∼6.8 kbp, ∼2.6 kbp and ∼1.8 kbp; mRNAs encoding β1 and α2 isoforms of neuregulin were revealed. NG108-15 cells were induced to differentiate by chemicals, and the chemical-induced differentiation of NG108-15 cells increased the level of neuregulin mRNA expression ∼3-fold while the expression of a housekeeping gene remained relatively unchanged. The activity of neuregulin in the conditioned medium of NG108-15 cells was reduced by treating the medium with heparin and anti-neuregulin antibody. In addition, NG108-15 cells were transfected with antisense neuregulin cDNA and its expression of neuregulin was reduced, while its neuregulin-induced tyrosine phosphorylation activity was markedly decreased. This is the first direct demonstration that the NG108-15 cell-induced AChR up-regulation on cultured myotubes is mediated by neuron-derived neuregulin.