Abstract
We previously reported a 120-kDa phosphoprotein that translocated from cytosol to the apical membrane of gastric parietal cells in association with stimulation of HCl secretion. To determine the molecular identity of the protein, we performed molecular cloning and expression of the protein. Immunoblot analysis showed that this protein was highly enriched in tissues that secrete water, such as parietal cell, choroid plexus, salivary duct, lacrimal gland, kidney, airway epithelia, and chorioretinal epithelia. We named this protein "parchorin" based on its highest enrichment in parietal cells and choroid plexus. We obtained cDNA for parchorin from rabbit choroid plexus coding a protein consisting of 637 amino acids with a predicted molecular mass of 65 kDa. The discrepancy in size on 6% SDS-polyacrylamide gel electrophoresis is considered to be due to its highly acidic nature (pI = 4.18), because COS-7 cells transfected with parchorin cDNA produced a protein with apparent molecular mass of 120 kDa on 6% SDS-polyacrylamide gel electrophoresis. Parchorin is a novel protein that has significant homology to the family of chloride intracellular channels (CLIC), especially the chloride channel from bovine kidney, p64, in the C-terminal 235 amino acids. When expressed as a fusion protein with green fluorescent protein (GFP) in the LLC-PK1 kidney cell line, GFP-parchorin, unlike other CLIC family members, existed mainly in the cytosol. Furthermore, when Cl(-) efflux from the cell was elicited, GFP-parchorin translocated to the plasma membrane. These results suggest that parchorin generally plays a critical role in water-secreting cells, possibly through the regulation of chloride ion transport.
Highlights
When gastric acid secretion is stimulated, a marked redistribution of Hϩ,Kϩ-ATPase, the gastric proton pump, occurs in the parietal cell
Tissue Distribution of Parchorin—We previously observed that parchorin was highly enriched in rabbit parietal cells [6]
Because brain consists of quite heterologous cells, we further investigated the detailed distribution of parchorin in the brain
Summary
When gastric acid secretion is stimulated, a marked redistribution of Hϩ,Kϩ-ATPase, the gastric proton pump, occurs in the parietal cell. Tubulovesicles containing Hϩ,KϩATPase are incorporated into the apical membrane, the latter acquiring Kϩ and ClϪ permeability, and Hϩ,Kϩ-ATPase can exchange Kϩ for protons, creating HCl together with water secretion [1]. An interesting phosphoprotein was identified which migrated with apparent molecular mass of 120 kDa by 6 –7.5% SDS-polyacrylamide gel electrophoresis (SDS-PAGE).1 This so-called pp120 protein existed mainly in the cytosol and translocated to the apical membrane-rich fraction when acid secretion was activated. Using antibodies to examine the tissue distribution, we found that pp120 was highly enriched in tissues associated with water transport, including parietal cells and choroid plexus We named this protein “parchorin” based on the enrichment in the parietal and choroid cellular locations. We suggest that the expression of cellular parchorin is specified for water movement, possibly through the regulation of chloride ion transport
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