Assembly of the mammalian muscle acetylcholine receptor in transfected COS cells

Abstract

We have investigated the mechanisms of assembly and transport to the cell surface of the mouse muscle nicotinic acetylcholine receptor (AChR) in transiently transfected COS cells. In cells transfected with all four subunit cDNAs, AChR was expressed on the surface with properties resembling those seen in mouse muscle cells (Gu, Y., A. F. Franco, Jr., P. D. Gardner, J. B. Lansman, J. R. Forsayeth, and Z. W. Hall. 1990. Neuron. 5:147-157). When incomplete combinations of AChR subunits were expressed, surface binding of 125I-α-bungarotoxin was not detected except in the case of αβγ which expressed <15% of that seen with all four subunits. Immunoprecipitation and sucrose gradient sedimentation experiments showed that in cells expressing pairs of subunits, αδ and αγ heterodimers were formed, but αβ was not. When three subunits were expressed, αδβ and αγβ complexes were formed. Variation of the ratios of the four subunit cDNAs used in the transfection mixture showed that surface AChR expression was decreased by high concentrations of δ or γ cDNAs in a mutually competitive manner. High expression of δ or γ subunits also each inhibited formation of a heterodimer with α and the other subunit. These results are consistent with a defined pathway for AChR assembly in which αδ and αγ heterodimers are formed first, followed by association with the β subunit and with each other to form the complete AChR.