NEURONAL NICOTINIC ACETYLCHOLINE-RECEPTORS ON BOVINE CHROMAFFIN CELLS- CLONING, EXPRESSION, AND GENOMIC ORGANIZATION OF RECEPTOR SUBUNITS

Neuronal nicotinic acetylcholine receptors from bovine adrenomedullary chromaffin cells play a primary role in triggering catecholamine secr etion, In the present study, their constituent subunits were character ized. In addition to the alpha 3 subunit, which we have previously clo ned, the presence of alpha 5 and beta 4 but not of beta 2 subunits was detected by reverse transcription-PCR analysis of mRNA from adrenal m edulla, In situ hybridization indicated that alpha 3, alpha 5, and bet a 4 subunits are coexpressed in all chromaffin cells. The primary stru cture of alpha 5 and beta 4 subunits was determined and functional rec eptors were obtained upon coinjection of subunit cRNAs into Xenopus oo cytes. In contrast to other beta 4-containing nicotinic receptors, the ones formed by the bovine beta 4 subunit are insensitive to the agoni st cytisine, Finally, we characterized the intergenic region of alpha 3 and alpha 5 subunits, which together with the beta 4 subunit, form a gene cluster in rats and chickens. RNase assays and the existence of overlapping cDNAs indicate that, in the bovine genome, the alpha 3 and alpha 5 genes overlap at their 3' ends. This fact is probably due to inefficient transcription termination, as a result of weak polyadenyla tion signals.