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J. Biol. Chem., Vol. 265, Issue 29, 17560-17567, 10, 1990
S Couturier, L Erkman, S Valera, D Rungger, S Bertrand, J Boulter, M Ballivet and D Bertrand
In vertebrates, neuronal nicotinic acetylcholine receptors (nAChRs)
assemble in an unknown stoichiometry from two homologous subunits, an alpha
and a non-alpha. How large is the repertoire of these subunits and how many
subtypes of functionally different nAChRs can they constitute? We found in
the avian genome a cluster of three closely linked genes spanning 28
kilobase pairs and encoding three proteins, n alpha 3, alpha 3, and alpha
5, that have the features expected of neuronal nAChR subunits. Gene n alpha
3 lies 5' of alpha 3 (whose role in cholinoception has already been
established) and is transcribed from the same DNA strand, whereas alpha 5
lies 3' of alpha 3 and is transcribed from the opposite DNA strand. The
structure of the n alpha 3 and alpha 5 genes consists of six exons with
precisely conserved splice sites and is identical to the structure of the
previously characterized avian neuronal receptor subunit genes alpha 2,
alpha 3, alpha 4, and n alpha 1. alpha 3, n alpha 3, and alpha 5
transcripts are rare in the central nervous system, but alpha 3 and n alpha
3 are readily detectable in embryonic superior cervical and ciliary
ganglia. In order to assay function, the gene encoding n alpha 3 and the
cDNAs encoding alpha 3, alpha 4, alpha 5, and n alpha 1 were subcloned into
an expression vector, and the constructs were injected into Xenopus oocyte
nuclei, either singly or in pairwise combinations of one alpha and one
non-alpha. One to five days later, ACh sensitivity of the injected oocytes
was examined in voltage clamp. The n alpha 3 gene and n alpha 1 cDNA
elicited assembly of nAChRs when coinjected with alpha 3 or alpha 4 cDNA
and the electrophysiological properties of the four pairwise combinations
were significantly different. alpha 5, however, did not direct the assembly
of functional nAChRs when injected alone or in combination with n alpha 1
or n alpha 3.
Alpha 5, alpha 3, and non-alpha 3. Three clustered avian genes encoding neuronal nicotinic acetylcholine receptor-related subunits
Department of Biochemistry, University of Geneva, Switzerland.
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