Structural Basis of the Differential Subcellular Localization of Rat Eag K+ Channels

• Main Authors: Huei-Min Lin, 林惠敏
• Other Authors: 湯志永
• Format: Others
• Language: zh-TW
• Published: 2006
• Online Access: http://ndltd.ncl.edu.tw/handle/96867859413428627716

碩士 === 國立臺灣大學 === 生理學研究所 === 94 === The EAG potassium channel is a member of the superfamility of voltage- gated potassium channels, and can be subdivided into three different subfamilies based on sequence homology：eag, erg and elk. In mammalians, the eag subfamily includes Eag1 and Eag2 K channels, which have been shown to express exclusively in central nervous system. Previous studies from our lab have demonstrated that rat Eag1 and rat Eag2 displayed differential subcellular localization patterns in hippocampus neurons. In the present study, we aim to determine the structural basis of differential localization of rEag1 and rEag2 in neurons. We intend to test the following hypotheses：(1) the differential localization pattern may be related to the different glycosylation patterns between rEag1 and rEag2. (2) the intracellular carboxyl terminal（C-terminal） region of rEag1 and rEag2 may contain the targeting signal that dictates the subcellular localization patterns of the two channels, and (3) rEag1 and rEag2 may form hetero-tetramers in neurons, suggesting the existence of a dominancy in the subcellular targeting within hetero-tetramer. We first studied the effect of abolishing either of the two known glycosylation sites in GFP-rEag1. Our results did not suppory a role of glycosylation in the subcellular localization of rEag1. We next engineered several chimeras that involved the C terminal regions of rEag1 and rEag2. By studing the confocal image of neuron transfected chimeric GFP constructs, we found evidence showing the C –terminal regions of rEag channels may confer the differential localization patterns of the two channels. Finally, we performed biochemical experiments to confirm that rEag1 and rEag2 are capable of forming hetero-tetramers in vivo. In conclusion, the C terminals of rEag channels play an important role in differential localization in neurons and rEag1 and rEag2 can assemble with each other in vivo.