Calcium Inhibits Paracellular Sodium Conductance through Claudin-2 by Competitive Binding |
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Authors: | Alan S L Yu Mary H Cheng Rob D Coalson |
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Institution: | From the ‡Division of Nephrology, Department of Medicine, and ;the §Department of Physiology and Biophysics, University of Southern California Keck School of Medicine, Los Angeles, California 90089 and ;the ¶Department of Chemistry, University of Pittsburgh, Pittsburgh, Pennsylvania 15260 |
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Abstract: | Claudins form paracellular pores at the tight junction in epithelial cells. Profound depletion of extracellular calcium is well known to cause loosening of the tight junction with loss of transepithelial resistance. However, moderate variations in calcium concentrations within the physiological range can also regulate transepithelial permeability. To investigate the underlying molecular mechanisms, we studied the effects of calcium on the permeability of claudin-2, expressed in an inducible MDCK I cell line. We found that in the physiological range, calcium acts as a reversible inhibitor of the total conductance and Na+ permeability of claudin-2, without causing changes in tight junction structure. The effect of calcium is enhanced at low Na+ concentrations, consistent with a competitive effect. Furthermore, mutation of an intrapore negatively charged binding site, Asp-65, to asparagine partially abrogated the inhibitory effect of calcium. This suggests that calcium competes with Na+ for binding to Asp-65. Other polyvalent cations had similar effects, including La3+, which caused severe and irreversible inhibition of conductance. Brownian dynamics simulations demonstrated that such inhibition can be explained if Asp-65 has a relatively high charge density, thus favoring binding of Ca2+ over that of Na+, reducing Ca2+ permeation by inhibiting its dissociation from this site, and decreasing Na+ conductance through repulsive electrostatic interaction with Ca2+. These findings may explain why hypercalcemia inhibits Na+ reabsorption in the proximal tubule of the kidney. |
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Keywords: | Calcium Epithelium Ion Channels Junctions Sodium Transport Claudin Tight Junction |
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