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Intracellular localization of ClC chloride channels and their ability to form hetero‐oligomers
Author(s) -
Suzuki Tatsunori,
Rai Tatemitsu,
Hayama Atsushi,
Sohara Eisei,
Suda Shin,
Itoh Tomohiro,
Sasaki Sei,
Uchida Shinichi
Publication year - 2006
Publication title -
journal of cellular physiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.529
H-Index - 174
eISSN - 1097-4652
pISSN - 0021-9541
DOI - 10.1002/jcp.20516
Subject(s) - intracellular , chloride channel , immunoprecipitation , epitope , immunofluorescence , chemistry , microbiology and biotechnology , transfection , hek 293 cells , cytoplasm , membrane , organelle , biophysics , oligomer , biochemistry , biology , gene , antibody , genetics , organic chemistry
ClC chloride channels (ClCs) can be classified into two groups in terms of their cellular localizations: ClCs present in the plasma membranes and those residing in intracellular organelles. Members of the latter group, including ClC‐3, ClC‐4, ClC‐5, ClC‐6, and ClC‐7, are often co‐expressed in a variety of cell types in many organs. Although the localization of individual channels within cells has been investigated, the degree of overlap between the locations of different ClCs in the same cell has not been clarified. To address this question, different combinations of ClCs, engineered to encode specific epitope tags (FLAG or HA), were either transiently or stably transfected into HEK293 cells, and we then compared the intracellular localization of the expressed channel proteins by immunofluorescence microscopy. Immunofluorescence images of the alternatively labeled channels clearly showed significant co‐localization between all pair‐wise combinations of ClCs. In particular, ClC‐3, ClC‐4, and ClC‐5 showed a high degree of co‐localization. As a significant degree of co‐localization between ClCs was observed, we used co‐immunoprecipitation to evaluate oligomer formation, and found that each ClC tested could form homo‐oligomers, and that any pair‐wise combination of ClC‐3, ClC‐4, and ClC‐5 could also form hetero‐oligomers. Neither ClC‐6 nor ClC‐7 was co‐precipitated with any other channel protein. These results suggest that within cells ClC‐3, ClC‐4, and ClC‐5 may have combinatorial functions, whereas ClC‐6 and ClC‐7 are more likely to function as homo‐oligomers. © 2005 Wiley‐Liss, Inc.