Claudin-10a Deficiency Shifts Proximal Tubular Cl- Permeability to Cation Selectivity via Claudin-2 Redistribution

Abstract

Background The tight junction proteins claudin-2 and claudin-10a form paracellular cation and anion channels, respectively, and are expressed in the proximal tubule. However, the physiological role of claudin-10a in the kidney has been unclear. Methods To investigate the physiologic role of claudin-10a, we generated claudin-10a-deficient mice; confirmed successful knockout by Southern blot, Western blot, and immunofluorescence staining; and analyzed urine and serum of knockout and wild-type animals. We also used electrophysiologic studies to investigate the functionality of isolated proximal tubules, and studied compensatory regulation by pharmacologic intervention, RNA sequencing analysis, Western blot, immunofluorescence staining, and respirometry. Results Mice deficient in claudin-10a were fertile and without overt phenotypes. Upon knockout, claudin-10a was replaced by claudin-2 in all proximal tubule segments. Electrophysiology showed conversion from paracellular anion preference to cation preference and a loss of paracellular Cl- over HCO3- preference. As a consequence, there was tubular retention of calcium and magnesium, higher urine pH, and mild hypermagnesemia. A comparison of other urine and serum parameters under control conditions and sequential pharmacologic transport inhibition, as well as unchanged fractional lithium excretion, suggested compensative measures in proximal and distal tubular segments. Changes in proximal tubular oxygen handling and differential expression of genes regulating fatty acid metabolism indicated proximal tubular adaptation. Western blot and immunofluorescence revealed alterations in distal tubular transport. Conclusions Claudin-10a is the major paracellular anion channel in the proximal tubule and its deletion causes calcium and magnesium hyperreabsorption by claudin-2 redistribution. Transcellular transport in proximal and distal segments and proximal tubular metabolic adaptation compensate for loss of paracellular anion permeability.

Molecular mechanisms underlying paracellular calcium and magnesium reabsorption in the proximal tubule and thick ascending limb

“…Supporting the capacity for CLDN2 to reabsorb magnesium from the proximal tubule are observations from the Cldn10a knockout mouse. These animals display hypermagnesemia, hypomagnesuria, and have CLDN10a‐deficient proximal tubule junctions repopulated by increased CLDN2 expression, in line with CLDN2 enabling the hyperabsorption of magnesium from this segment in the absence of claudin‐10a 29 …”

“…The appearance of a lumen‐positive potential occurs as a consequence of the preferential reabsorption of bicarbonate along the early proximal tubule by processes involving the sodium hydrogen exchanger isoform 3 (NHE3), the apical and cytosolic carbonic anhydrases, and sodium bicarbonate cotransporter 1 (NBCe1), which leads to the concentration of chloride in the tubular lumen, since HCO 3 − is the primary counterion moving with sodium in the early portions of the proximal tubule 7 . Chloride is then reabsorbed via a paracellular pathway 29 . Contributing to this change from slightly lumen negative to slightly lumen positive is a change in relative ion selectivity from cation selective in the early proximal tubule to slightly anion selective in the late proximal tubule 29 .…”

“…Claudins (CLDN)‐2, ‐12, and ‐10a are expressed in the proximal tubule of adult animals 30,33,34 . CLDN2 and CLDN12 confer cation permeability to this segment, 30,35–37 while CLDN‐10a confers anion permeability 29 …”

“…However, there are very limited experimental data supporting this. A single epithelial cell culture model examining magnesium permeability in response to overexpression of claudin‐2 found increased magnesium permeability relative to an empty vector 29 . Furthermore, Cldn2 single knockout mice do not have hypermagnesiuria, nor reduced plasma magnesium levels when fed normal rodent chow 30,35,36 .…”

Imputation-powered whole-exome analysis identifies rare coding variants and genes associated with kidney function and disease in the UK Biobank

“…Isoform claudin-10a is expressed in the kidney proximal tubule and the uterus, claudin-10b in the thick ascending limb of Henle's loop and various exocrine glands. [27][28][29] Mutations in CLDN10 can cause the autosomal recessively inherited HELIX syndrome, 30 which features hypohidrosis, electrolyte imbalance, lacrimal gland dysfunction, ichthyosis, and xerostomia. Abnormal eGFR was found for some mutations, [31][32][33][34] but not for others.…”