Incorporation of the beta3 subunit has a dominant-negative effect on the function of recombinant central-type neuronal nicotinic receptors.
The beta3 neuronal nicotinic subunit is localized in dopaminergic areas of the central nervous system, in which many other neuronal nicotinic subunits are expressed. So far, beta3 has only been shown to form functional receptors when expressed together with the alpha3 and beta4 subunits. We have systematically tested in Xenopus laevis oocytes the effects of coexpressing human beta3 with every pairwise functional combination of neuronal nicotinic subunits likely to be relevant to the central nervous system. Expression of alpha7 homomers or alpha/beta pairs (alpha2, alpha3, alpha4, or alpha6 together with beta2 or beta4) produced robust nicotinic currents for all combinations, save alpha6beta2 and alpha6beta4. Coexpression of wild-type beta3 led to a nearly complete loss of function (measured as maximum current response to acetylcholine) for alpha7 and for all functional alpha/beta pairs except for alpha3beta4. This effect was also seen in hippocampal neurons in culture, which lost their robust alpha7-like responses when transfected with beta3. The level of surface expression of nicotinic binding sites (alpha3beta4, alpha4beta2, and alpha7) in tsA201 cells was only marginally affected by beta3 expression. Furthermore, the dominant-negative effect of beta3 was abolished by a valine-serine mutation in the 9' position of the second transmembrane domain of beta3, a mutation believed to facilitate channel gating. Our results show that incorporation of beta3 into neuronal nicotinic receptors other than alpha3beta4 has a powerful dominant-negative effect, probably due to impairment in gating. This raises the possibility of a novel regulatory role for the beta3 subunit on neuronal nicotinic signaling in the central nervous system.
Broadbent
Steven
S
Department of Pharmacology, University College London, Gower St., London, WC1E 6BT, UK.
Groot-Kormelink
Paul J
PJ
Krashia
Paraskevi A
PA
Harkness
Patricia C
PC
Millar
Neil S
NS
Beato
Marco
M
Sivilotti
Lucia G
LG
D000818:Animals; D002460:Cell Line; D002478:Cells, Cultured; D005260:Female; D006624:Hippocampus / Q000166:cytology / Q000378:metabolism; D006801:Humans; D008564:Membrane Potentials; D009474:Neurons / Q000378:metabolism*; D009865:Oocytes / Q000502:physiology*; D018408:Patch-Clamp Techniques; D011978:Receptors, Nicotinic / Q000235:genetics* / Q000502:physiology*; D011994:Recombinant Proteins / Q000378:metabolism; D014982:Xenopus laevis
D011978:Receptors, Nicotinic; D011994:Recombinant Proteins