Efficient expression of functional (?6?2)2?3 AChRs in Xenopus oocytes from free subunits using slightly modified ?6 subunits.

ABSTRACT: Human (?6?2)(?4?2)?3 nicotinic acetylcholine receptors (AChRs) are essential for addiction to nicotine and a target for drug development for smoking cessation. Expressing this complex AChR is difficult, but has been achieved using subunit concatamers. In order to determine what limits expression of ?6* AChRs and to efficiently express ?6* AChRs using free subunits, we investigated expression of the simpler (?6?2)2?3 AChR. The concatameric form of this AChR assembles well, but is transported to the cell surface inefficiently. Various chimeras of ?6 with the closely related ?3 subunit increased expression efficiency with free subunits and produced pharmacologically equivalent functional AChRs. A chimera in which the large cytoplasmic domain of ?6 was replaced with that of ?3 increased assembly with ?2 subunits and transport of AChRs to the oocyte surface. Another chimera replacing the unique methionine 211 of ?6 with leucine found at this position in transmembrane domain 1 of ?3 and other ? subunits increased assembly of mature subunits containing ?3 subunits within oocytes. Combining both ?3 sequences in an ?6 chimera increased expression of functional (?6?2)2?3 AChRs to 12-fold more than with concatamers. This is pragmatically useful, and provides insights on features of ?6 subunit structure that limit its expression in transfected cells.

SUBMITTER: Ley CK 

PROVIDER: S-EPMC4113361 | biostudies-literature | 2014

REPOSITORIES: biostudies-literature