Abstract
Little is known of the roles played by ion channels in cancer. Here we describe a pair of closely related calcium-activated chloride channels whose differential regulation in normal, apoptotic, and transformed mouse cells suggests that channel function is proapoptotic and antineoplastic. While mCLCA1 predominates over mCLCA2 under normal physiological conditions, this relationship is reversed by apoptotic stress both in developing mammary gland and in cultured HC11 mammary epithelial cells. Consistent with an apoptosis-promoting role, splicing of mCLCA2 is disrupted in apoptosis-resistant tumor cell lines and in HC11 cells selected for resistance to detachment-induced apoptosis (anoikis). Unexpectedly, mCLCA1 message is also down-regulated in these cells by at least 30-fold. These results suggest that both genes antagonize survival of mammary tumor cells by sensitizing them to anoikis. When MCF7 or HEK293 tumor cells were transfected with plasmids encoding either mCLCA1 or mCLCA2, colony formation was greatly reduced relative to a vector-transfected control, demonstrating that calcium-sensitive chloride channel (CLCA) expression is deleterious to tumor cell survival. Furthermore, mammary epithelial cells overexpressing mCLCA2 had twice the rate of apoptosis of normal cells when subjected to serum starvation and formed multinuclear giants at a high frequency in normal culture, suggesting that mCLCA2 can promote either apoptosis or senescence.
Highlights
The CLCA1 family of calcium-activated chloride channels comprises about a dozen members far expressed in a variety of organ and tissue types in mammals [1,2,3,4,5,6,7,8]
While mCLCA1 predominates over mCLCA2 under normal physiological conditions, this relationship is reversed by apoptotic stress both in developing mammary gland and in cultured HC11 mammary epithelial cells
When MCF7 or HEK293 tumor cells were transfected with plasmids encoding either mCLCA1 or mCLCA2, colony formation was greatly reduced relative to a vector-transfected control, demonstrating that calcium-sensitive chloride channel (CLCA) expression is deleterious to tumor cell survival
Summary
The CLCA1 family of calcium-activated chloride channels comprises about a dozen members far expressed in a variety of organ and tissue types in mammals [1,2,3,4,5,6,7,8]. The CLCA precursor is about 900 amino acids long with one proteolytic cleavage site following the amino-terminal signal sequence and another around residue 600 The latter results in a 90-kDa product with perhaps four transmembrane segments and a 30 – 40-kDa product that may be found in close association with the first [1, 3, 4, 6]. We reasoned that if CLCA channels played a similar role in mammary epithelial cells, the mCLCA2 gene should be down-regulated in cells that have become apoptosis-resistant, e.g. tumor cell lines. We asked whether re-expression of CLCA genes would be tolerated by tumor cell lines
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