The relative importance of plasma membrane Ca 2؉ -ATPase (PMCA) 1 and PMCA4 was assessed in mice carrying null mutations in their genes (Atp2b1 and Atp2b4). Loss of both copies of the gene encoding PMCA1 caused embryolethality, whereas heterozygous mutants had no overt disease phenotype. Despite widespread and abundant expression of PMCA4, PMCA4 null (Pmca4 ؊/؊ ) mutants exhibited no embryolethality and appeared outwardly normal. Loss of PMCA4 impaired phasic contractions and caused apoptosis in portal vein smooth muscle in vitro; however, this phenotype was dependent on the mouse strain being employed. Pmca4 ؊/؊ mice on a Black Swiss background did not exhibit the phenotype unless they also carried a null mutation in one copy of the Pmca1 gene. Pmca4 ؊/؊ male mice were infertile but had normal spermatogenesis and mating behavior. Pmca4؊/؊ sperm that had not undergone capacitation exhibited normal motility but could not achieve hyperactivated motility needed to traverse the female genital tract. Ultrastructure of the motility apparatus in Pmca4 ؊/؊ sperm tails was normal, but an increased incidence of mitochondrial condensation indicated Ca 2؉ overload. Immunoblotting and immuno-histochemistry showed that PMCA4 is the most abundant isoform in testis and sperm and that it is localized to the principle piece of the sperm tail, which is also the location of the major Ca 2؉ channel (CatSper) required for sperm motility. These results are consistent with an essential housekeeping or developmental function for PMCA1, but not PMCA4, and show that PMCA4 expression in the principle piece of the sperm tail is essential for hyperactivated motility and male fertility.

embryos had an open rostral neural tube, but hematopoiesis and cardiovascular development were ostensibly normal. Golgi membranes of Spca1 ؊/؊ embryos were dilated, had fewer stacked leaflets, and were expanded in amount, consistent with increased Golgi biogenesis. The number of Golgi-associated vesicles was also increased, and rough endoplasmic reticulum had fewer ribosomes. Coated pits, junctional complexes, desmosomes, and basement membranes appeared normal in mutant embryos, indicating that processing and trafficking of proteins in the secretory pathway was not massively impaired. However, apoptosis was increased, possibly the result of secretory pathway stress, and a large increase in cytoplasmic lipid was observed in mutant embryos, consistent with impaired handling of lipid by the Golgi. Adult heterozygous mice appeared normal and exhibited no evidence of Hailey-Hailey disease; however, aged heterozygotes had an increased incidence of squamous cell tumors of keratinized epithelial cells of the skin and esophagus. These data show that loss of the Golgi Ca 2؉ pump causes Golgi stress, expansion of the Golgi, increased apoptosis, and embryonic lethality and demonstrates that SPCA1 haploinsufficiency causes a genetic predisposition to cancer. and Mn 2ϩ (10). Loss of PMR1 affects outer chain glycosylation, proteolytic processing, and trafficking of proteins in the secretory pathway (9). In mammals, SPCA1 is expressed in all tissues (1), whereas SPCA2 is expressed in only a limited set of tissues (3). Like PMR1, both SPCA1 and SPCA2 are localized to the Golgi and transport Ca 2ϩ and Mn 2ϩ (3, 11). There is evidence that the cell biological functions of SPCA1 are also similar to those of PMR1 (12).Loss of one copy of the human ATP2C1 gene, encoding SPCA1, causes Hailey-Hailey disease (HHD), an autosomal dominant skin disorder (13,14). SPCA1 protein levels in HHD keratinocytes are reduced to about half of normal levels, and Golgi Ca 2ϩ handling is impaired (15). HHD is similar to Darier disease, which is caused by null mutations in one copy of the human ATP2A2 gene, encoding SERCA2 (16). Both diseases are characterized by acantholysis (a disruption of cell-cell contacts) in the suprabasal layers of the skin. As the major ER Ca 2ϩ pump in most tissues, including keratinocytes, the function of SERCA2 is similar to that of SPCA1 in that it maintains luminal Ca 2ϩ concentrations in a major compartment of the secretory pathway. In mice, SERCA2 haploinsufficiency does not cause Darier disease but does lead to squamous cell tumors of keratinized epithelial cells (17, 18), the same cell type affected in Darier disease. In humans, a low incidence of squamous cell tumors has been reported in both Darier disease (19) and HHD (20, 21), but it is unclear whether this is a chance association or is caused by the reduction in Ca 2ϩ pump levels and activity. In the current study, we developed a gene-targeted mouse model for SPCA1 and analyzed the phenotype resulting from heterozygous and homozygous null mutations. The results sho...

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ClC-2 is localized to the apical membranes of secretory epithelia where it has been hypothesized to play a role in fluid secretion. Although ClC-2 is clearly the inwardly rectifying anion channel in several tissues, the molecular identity of the hyperpolarization-activated Cl ؊ current in other organs, including the salivary gland, is currently unknown. To determine the nature of the hyperpolarization-activated Cl ؊ current and to examine the role of ClC-2 in salivary gland function, a mouse line containing a targeted disruption of the Clcn2 gene was generated. The resulting homozygous Clcn2 ؊/؊ mice lacked detectable hyperpolarization-activated chloride currents in parotid acinar cells and, as described previously, displayed postnatal degeneration of the retina and testis. The magnitude and biophysical characteristics of the volume-and calcium-activated chloride currents in these cells were unaffected by the absence of ClC-2. Although ClC-2 appears to contribute to fluid secretion in some cell types, both the initial and sustained salivary flow rates were normal in Clcn2 ؊/؊ mice following in vivo stimulation with pilocarpine, a cholinergic agonist. In addition, the electrolytes and protein contents of the mature secretions were normal. Because ClC-2 has been postulated to contribute to cell volume control, we also examined regulatory volume decrease following cell swelling. However, parotid acinar cells from Clcn2 ؊/؊ mice recovered volume with similar efficiency to wild-type littermates. These data demonstrate that ClC-2 is the hyperpolarization-activated Cl ؊ channel in salivary acinar cells but is not essential for maximum chloride flux during stimulated secretion of saliva or acinar cell volume regulation.

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