Multistep expression and assembly of neuronal nicotinic receptors is both host-cell- and receptor-subtype-dependent

We tested the hypothesis that the folding, assembly and insertion of neuron al nicotinic receptors are critically dependent on the host cell line. We u sed recombinant adenoviruses encoding either the rat alpha 7, alpha 4 or be ta 2 subunits in which expression of the subunit is controlled by a tetracy cline-dependent promoter to screen five cell lines (GH4C1, SH-EP1, CVI, SN- 56, and CHO-CAR). All five lines do not express detectable nicotinic recept or but do express receptor for human adenovirus, and all expressed mRNA for alpha 7, alpha 4 and beta 2 subunits when infected with viruses. Each cell Line expressed varying levels of alpha 4 beta 2 receptors that bound [H-3] cytisine, but only the GH4C1 and SH-EP1 cell lines expressed either surface or internal alpha 7 receptors that bound [I-125]alpha bungarotoxin ([I-125 ]alpha-BGT). All five cell lines expressed a 60 kDa protein immunoblotted b y anti-alpha 7 antibodies when infected with the alpha 7 virus, presumably representing unassembled alpha 7 subunits, In addition, GH4C1 cells express ed over 10-fold more surface alpha 7 receptor than SH-EP1 cells, even thoug h the total alpha 7 receptor in the two cell lines was similar. Sedimentati on experiments indicate that SH-EP1 cells only partially assemble alpha 7 r eceptors compared with GH4C1 cells and control alpha 7 from rat brain. Thes e data suggest that not only is surface alpha 7 receptor expression a multi step process, but that each step may involve cell-specific assembly factors . (C) 2000 Elsevier Science B.V. All rights reserved.