Assembly of an adult type acetylcholine receptor in a mouse cell line transfected with rat muscle epsilon-subunit DNA.
The mouse muscle cell line BC3H-1 expresses an acetylcholine receptor (AChR) composed of alpha-, beta-, gamma- and delta-subunits. The functional characteristics of this AChR are comparable to the non-synaptic AChR subtype in mouse muscle. To investigate the role of the epsilon-subunit, which is believed to replace the gamma-subunit in forming the adult AChR subtype, BC3H-1 cells were stably transfected with cDNA encoding the rat muscle AChR epsilon-subunit. Expression of this cDNA was under the control of a heat shock promoter, and the plasmid carried the neomycin resistance gene for selection. Several clones were isolated that had integrated the plasmid DNA in a stable form and produced epsilon-subunit specific RNA after heat induction. Single-channel current recording from cells which contained abundant epsilon-subunit mRNA identified a novel AChR channel having a larger conductance than the native AChR in these cells. These results suggest that the rat muscle epsilon-subunit may assemble with mouse muscle alpha-, beta- and delta-subunits to form a mouse-rat hybrid AChR with properties similar to that of end-plate channels in the mature mammalian neuromuscular synapse. The novel AChR channel appears in the surface membrane within a few hours following the rise in epsilon-subunit mRNA. Thus, the notion that replacement of the gamma-subunit by the epsilon-subunit during development is the result of the postnatal rise in the level of epsilon-subunit specific mRNA is further supported.
Max-Planck-Institut für Medizinische Forschung, Heidelberg, FRG.
M
M
Criado
M
M
Koenen
B
B
Sakmann
D000818:Animals; D015152:Blotting, Northern; D015139:Blotting, Southern; D002460:Cell Line; D006358:Hot Temperature; D007473:Ion Channels / Q000502:physiology; D051379:Mice; D009132:Muscles / Q000502:physiology; D010446:Peptide Fragments / Q000502:physiology; D051381:Rats; D011950:Receptors, Cholinergic / Q000096:biosynthesis / Q000502:physiology*; D011994:Recombinant Proteins / Q000096:biosynthesis; D014158:Transcription, Genetic; D014162:Transfection
D007473:Ion Channels; D010446:Peptide Fragments; D011950:Receptors, Cholinergic; D011994:Recombinant Proteins