Title

Regulation By Cycloheximide And Lowered Temperature Of Cell-Surface α7-Nicotinic Acetylcholine Receptor Expression On Transfected Sh-Ep1 Cells

Department

neurobiology

Document Type

Article

Abstract

Heterologous expression of functional, nicotinic acetylcholine receptors (nAChR) in mammalian cells has been difficult to achieve or optimize, even for nAChR containing only one kind of subunit. In this study, we determined effects of lowered temperature or of exposure to the protein synthesis inhibitor cycloheximide (CHX) on cell surface expression of homomeric α7-nAChR in transfected SH-EP1 human epithelial cells. We found that incubation of cells for 2 days at 25°C or in the presence of 0.5-2 μg/mL of CHX caused ∼four- or ∼eight-fold increases, respectively, in surface binding sites for 125I-labeled α-bungarotoxin (I-Bgt). These increases were accompanied by increases in peak whole-cell current responses to nicotinic agonists. Either treatment lowered protein synthesis and cell proliferation, but experiments using puromycin indicated that a reduction in protein synthesis or cell proliferation per se was not sufficient to increase surface binding. I-Bgt binding to whole-cell membrane pools increased in response to either treatment, suggesting that the increase in surface binding was due, at least in part, to an increase in intracellular receptor levels. The cyclophilin inhibitor cyclosporin A reduced surface expression in untreated as well as CHX- or 25°C-treated cells. The results suggest practical means for increasing cell surface and functional expression of α7-nAChR. Although these effects are not simply due to protein synthesis inhibition or reduced cell proliferation, they do involve an increase in intracellular receptor pool size.

Publication Date

1-1-2003

Publication Title

Journal of Neurochemistry

ISSN

00223042

Volume

85

Issue

3

First Page

581

Last Page

591

Digital Object Identifier (DOI)

10.1046/j.1471-4159.2003.01658.x

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