Functional acetylcholine receptors expressed in Xenopus oocytes after injection of Torpedo beta, gamma, and delta subunit RNAs are a consequence of endogenous oocyte gene expression.
The nicotinic acetylcholine receptor (AChR) is an oligomeric transmembrane glycoprotein consisting of four homologous subunits in a stoichiometry alpha 2 beta gamma delta. Xenopus oocytes were used to study the effects of selectively deleting the alpha subunit of the Torpedo californica AChR on functional receptor expression. Oocytes microinjected with only Torpedo beta, gamma, and delta subunit RNAs showed small acetylcholine-elicited currents. These "alpha-less" AChRs were pharmacologically similar to the wild-type (i.e., Torpedo alpha 2 beta gamma delta) receptor. Actinomycin D, which blocks endogenous RNA transcription, completely inhibited the expression of alpha-less but not wild-type receptor. Coinjection of antisense RNA to the alpha subunit of the Xenopus muscle AChR with Torpedo beta, gamma, and delta subunit RNAs significantly reduced expression of the alpha-less AChRs without altering expression of wild-type receptors. These results indicate that Xenopus oocytes express low levels of AChR alpha subunit mRNA that, when translated, can lead to the formation of functional Xenopus-Torpedo AChR hybrids. These results strongly suggest that, unless the potential contribution of endogenous subunits can be determined, caution must be exercised when analyzing the effects of subunit deletions on multisubunit protein expression in Xenopus oocytes.
Department of Pharmacology, University of Pennsylvania School of Medicine, Philadelphia 19104-6084.
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Buller
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White
D000109:Acetylcholine / Q000494:pharmacology; D000818:Animals; D002038:Bungarotoxins / Q000494:pharmacology; D003609:Dactinomycin / Q000494:pharmacology; D015870:Gene Expression / Q000187:drug effects; D046911:Macromolecular Substances; D008845:Microinjections; D009865:Oocytes / Q000502:physiology*; D012313:RNA; D016372:RNA, Antisense; D012333:RNA, Messenger / Q000235:genetics; D011978:Receptors, Nicotinic / Q000235:genetics* / Q000502:physiology; D014101:Torpedo; D014403:Tubocurarine / Q000494:pharmacology; D014982:Xenopus laevis
D002038:Bungarotoxins; D046911:Macromolecular Substances; D016372:RNA, Antisense; D012333:RNA, Messenger; D011978:Receptors, Nicotinic; D003609:Dactinomycin; D012313:RNA; D000109:Acetylcholine; D014403:Tubocurarine