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Am J Physiol Cell Physiol 274: C1545-C1551, 1998;
0363-6143/98 $5.00
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Vol. 274, Issue 6, C1545-C1551, June 1998

Effect of cell swelling on membrane and cytoplasmic distribution of pICln

Francesco Emma1, Sylvie Breton2, Rebecca Morrison3, Stephen Wright4, and Kevin Strange3

1 Renal Division, Children's Hospital, Boston 02115; 2 Renal Unit, Massachusetts General Hospital, Boston, Massachusetts 02129; 3 Laboratory of Cellular and Molecular Physiology, Departments of Anesthesiology and Pharmacology, Vanderbilt University Medical Center, Nashville, Tennessee 37232; and 4 Department of Physiology, University of Arizona, Tucson, Arizona 85724

pICln is found ubiquitously in mammalian cells and is postulated to play a critical role in cell volume regulation. Mutagenesis studies led to the proposal that pICln is a swelling-activated anion channel. However, recent studies in Madin-Darby canine kidney cells and endothelial cells have shown that the protein is localized primarily to the cytoplasm. It has therefore been postulated that activation involves reversible translocation of pICln from the cytoplasm and insertion into the plasma membrane. We tested this hypothesis using several different approaches. Fractionation of C6 glioma cells into plasma membrane- and cytoplasm-containing fractions demonstrated that ~90% of the recovered pICln was confined to the cytosol. Swelling had no effect on the relative amount of protein present in the plasma membrane fraction. Immunofluorescence microscopy revealed that pICln is localized primarily, if not exclusively, to the cytoplasm of swollen and nonswollen cells. Similarly, transfection of cells with a green fluorescent protein-labeled pICln construct failed to reveal any membrane localization of the protein. These findings do not support the hypothesis that pICln is a volume regulatory anion channel activated by swelling-induced membrane insertion.

anion channels; organic osmolytes; cell volume regulation; osmoregulation


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