Temperature-sensitive expression of Drosophila neuronal nicotinic acetylcholine receptors.
Heterologous expression of cloned Drosophila nicotinic acetylcholine receptor (nAChR) subunits indicates that these proteins misfold when expressed in mammalian cell lines at 37 degrees C. This misfolding can, however, be overcome either by growing transfected mammalian cells at lower temperatures or by the expression of Drosophila nAChR subunits in a Drosophila cell line. Whereas the Drosophila nAChR beta subunit (SBD) cDNA, reported previously, lacked part of the SBD coding sequence, here we report the construction and expression of a full-length SBD cDNA. We have examined whether problems in expressing functional Drosophila nAChRs in either Xenopus oocytes or mammalian cell lines can be attributed to an inability of these expression systems to assemble correctly Drosophila nAChRs. Despite expression in what might be considered a more native cellular environment, we have been unable to detect functional nAChRs in a Drosophila cell line unless Drosophila nAChR subunit cDNAs are coexpressed with vertebrate nAChR subunits. Our results indicate that the folding of Drosophila nAChR subunits is temperature-sensitive and strongly suggest that the inability of these Drosophila nAChR subunits to generate functional channels in the absence of vertebrate subunits is due to a requirement for coassembly with as yet unidentified Drosophila nAChR subunits.
Lansdell
S J
SJ
Department of Pharmacology, University College London, U.K.
Schmitt
B
B
Betz
H
H
Sattelle
D B
DB
Millar
N S
NS
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D004330:Drosophila / Q000378:metabolism*; D005260:Female; D015870:Gene Expression; D006801:Humans; D007536:Isomerism; D008969:Molecular Sequence Data; D009474:Neurons / Q000378:metabolism*; D009865:Oocytes / Q000378:metabolism; D051381:Rats; D011978:Receptors, Nicotinic / Q000235:genetics / Q000378:metabolism*; D013696:Temperature; D014982:Xenopus laevis
D018076:DNA, Complementary; D011978:Receptors, Nicotinic