Movements of Individual BKCa Channels in Live Cell Membrane Monitored by Site-Specific Labeling Using Quantum Dots

作者:Won Sehoon; Kim Hae Deun; Kim Ji Yeon; Lee Byoung Cheol; Chang Sunghoe; Park Chul Seung*
来源:Biophysical Journal, 2010, 99(9): 2853-2862.
DOI:10.1016/j.bpj.2010.08.049

摘要

The movements of BKCa channels were investigated in live cells using quantum dots (QDs). The extracellular N-terminus was metabolically tagged with biotin, labeled with streptavidin-conjugated QDs and then monitored using real-time time-lapse imaging in COS-7 cells and cultured neurons. By tracking hundreds of channels, we were able to determine the characteristics of channel movements quantitatively. Channels in COS-7 cells exhibited a confined diffusion in an area of 1.915 mu m(2), with an initial diffusion coefficient of 0.033 mu m(2)/s. In neurons, the channel movements were more heterogeneous and highly dependent on subcellular location. While the channels in soma diffused slowly without clear confinement, axodendritic channels showed more rapid and pseudo-one-dimensional movements. Intriguingly, the channel movement in somata was drastically increased by the neuronal beta 4 subunit, in contrast to the channels in the axodendritic area where the mobility were significantly decreased. Thus, our results demonstrate that the membrane mobility of BKCa channels can be greatly influenced by the expression system used, subunit composition, and subcellular location. This OD-based, single-molecule tracking technique can be utilized to investigate the cellular mechanisms that determine the mobility as well as the localization of various membrane proteins in live cells.

  • 出版日期2010-11-3