Premium
ATP depletion inhibits the endocytosis of ClC‐2
Author(s) -
Dhani Sonja U.,
Kim Chiaw Patrick,
Huan LingJun,
Bear Christine E.
Publication year - 2008
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.21192
Subject(s) - dynamin , endocytosis , internalization , endocytic cycle , microbiology and biotechnology , endosome , biology , intracellular , caveolae , cell , chemistry , signal transduction , biochemistry
The chloride channel, ClC‐2 is expressed ubiquitously and participates in multiple physiological processes. In particular, ClC‐2 has been implicated in the regulation of neuronal chloride ion homeostasis and mutations in ClC‐2 are associated with idiopathic generalized epilepsy. Despite the physiological and pathophysiological significance of this channel, its regulation remains incompletely understood. The functional expression of ClC‐2 at the cell surface has been shown to be enhanced by depletion of cellular ATP, implicating its possible role in cellular energy sensing. In the present study, biochemical assays of cell surface expression suggest that this gain of function reflects, in part, an increase in channel number due to the reduction in ClC‐2 internalization by endocytosis. Cell surface expression of the disease‐causing mutant: G715E, thought to lack wild‐type nucleotide binding affinity, is similarly affected, suggesting that ATP‐depletion modifies the function of proteins in the endocytic pathway rather than ClC‐2 directly. Using a combination of immunofluorescence and biochemical studies, we confirmed that ClC‐2 is internalized via dynamin‐dependent endocytosis and that the change in surface expression evoked by ATP depletion is partially mimicked by inhibition of dynamin function using a dynamin dominant‐negative mutant (DynK44A). Furthermore, trafficking via the early endosomal compartment occurs in part through rab5‐associated vesicles and recycling of ClC‐2 to the cell surface occurs through a rab11 dependent pathway. In summary, we have determined that the internalization of ClC‐2 by endocytosis is inhibited by metabolic stress, highlighting the importance for understanding the molecular mechanisms mediating the endosomal trafficking of this channel. J. Cell. Physiol. 214:273–280, 2008. © 2007 Wiley‐Liss, Inc.