Journal of Neuroscience, Vol 10, 2771-2779, Copyright © 1990 by Society for Neuroscience
Expression of functional mouse muscle acetylcholine receptors in Chinese hamster ovary cells
JR Forsayeth, A Franco Jr, AB Rossi, JB Lansman and ZW Hall
Department of Physiology, School of Medicine, University of California, San Francisco 94143-0444.
CHO cells were transfected with cDNAs for all 4 subunits of the mouse
muscle acetylcholine receptor (AChR) to obtain a stable cell line (CHO- AR)
expressing the AChR on its surface. Immunoprecipitation experiments
established that the AChR contained alpha- and beta-subunits assembled with
gamma- and/or delta-subunits. In addition, one of the toxin- binding sites
of the AChR was blocked by a myasthenic serum that specifically recognizes
AChR containing the gamma-subunit. AChR from the CHO-AR cells had the same
sedimentation rate, association rate constant for the binding of
alpha-bungarotoxin (alpha BTX), and the same metabolic half-life as the
AChR in myotubes of the mouse muscle cell line C2C12. Electrophysiological
assay of CHO-AR cells by single- channel recording showed the presence of
ACh-responsive ion channels with the characteristics of the embryonic AChR
(gamma = 40 pS, tau = 5.6 msec). In some patches a smaller conductance
channel was also seen that may represent partially assembled receptor.
Fluorescence microscopy of fixed, permeabilized cells stained with
rhodamine-alpha- BTX demonstrated both perinuclear and diffuse surface
staining. The expression of fully assembled, functional mammalian muscle
AChR in nonmuscle cells will allow detailed investigation of its properties
and interactions with other cellular components.