期刊名称：Proceedings of the National Academy of Sciences
印刷版ISSN：0027-8424
电子版ISSN：1091-6490
出版年度：2002
卷号：99
期号：6
页码：3950-3955
DOI：10.1073/pnas.052699299
语种：English
出版社：The National Academy of Sciences of the United States of America
摘要：The functional properties and cellular localization of the human neuronal 7 nicotinic acetylcholine (AcCho) receptor (7 AcChoR) and its L248T mutated (mut) form were investigated by expressing them alone or as gene fusions with the enhanced version of the green fluorescent protein (GFP). Xenopus oocytes injected with wild-type (wt), mut7, or the chimeric subunit cDNAs expressed receptors that gated membrane currents when exposed to AcCho. As already known, AcCho currents generated by wt7 receptors decay much faster than those elicited by the mut7 receptors. Unexpectedly, the fusion of GFP to the wt and mutated 7 receptors led to opposite results: the AcCho-current decay of the wt receptors became slower, whereas that of the mutated receptors was accelerated. Furthermore, repetitive applications of AcCho led to a considerable "run-down" of the AcCho currents generated by mut7-GFP receptors, whereas those of the wt7-GFP receptors remained stable or increased in amplitude. The AcCho-current run-down of mut7-GFP oocytes was accompanied by a marked decrease of -bungarotoxin binding activity. Fluorescence, caused by the chimeric receptors expressed, was seen over the whole oocyte surface but was more intense and abundant in the animal hemisphere, whereas it was much weaker in the vegetal hemisphere. We conclude that fusion of GFP to wt7 and mut7 receptors provides powerful tools to study the distribution and function of 7 receptors. We also conclude that fused genes do not necessarily recapitulate all of the properties of the original receptors. This fact must be borne close in mind whenever reporter genes are attached to proteins.
