title: Activation of the bumetanide-sensitive Na+,K+,2Cl- cotransporter (NKCC2) is facilitated by Tamm-Horsfall protein in a chloride-sensitive manner.
abstract: Active transport of NaCl across thick ascending limb (TAL) epithelium is accomplished by Na(+),K(+),2Cl(-) cotransporter (NKCC2). The activity of NKCC2 is determined by vasopressin (AVP) or intracellular chloride concentration and includes its amino-terminal phosphorylation. Co-expressed Tamm-Horsfall protein (THP) has been proposed to interact with NKCC2. We hypothesized that THP modulates NKCC2 activity in TAL. THP-deficient mice (THP(-/-)) showed an increased abundance of intracellular NKCC2 located in subapical vesicles (+47% compared with wild type (WT) mice), whereas base-line phosphorylation of NKCC2 was significantly decreased (-49% compared with WT mice), suggesting reduced activity of the transporter in the absence of THP. Cultured TAL cells with low endogenous THP levels and low base-line phosphorylation of NKCC2 displayed sharp increases in NKCC2 phosphorylation (+38%) along with a significant change of intracellular chloride concentration upon transfection with THP. In NKCC2-expressing frog oocytes, co-injection with THP cRNA significantly enhanced the activation of NKCC2 under low chloride hypotonic stress (+112% versus +235%). Short term (30 min) stimulation of the vasopressin V2 receptor pathway by V2 receptor agonist (deamino-cis-D-Arg vasopressin) resulted in enhanced NKCC2 phosphorylation in WT mice and cultured TAL cells transfected with THP, whereas in the absence of THP, NKCC2 phosphorylation upon deamino-cis-D-Arg vasopressin was blunted in both systems. Attenuated effects of furosemide along with functional and structural adaptation of the distal convoluted tubule in THP(-/-) mice supported the notion that NaCl reabsorption was impaired in TAL lacking THP. In summary, these results are compatible with a permissive role for THP in the modulation of NKCC2-dependent TAL salt reabsorptive function.
authors: Department of Anatomy, Charité-Universitätsmedizin Berlin, Berlin, Germany.
authors: Kerim
authors: K
authors: Mutig
authors: Thomas
authors: T
authors: Kahl
authors: Turgay
authors: T
authors: Saritas
authors: Michael
authors: M
authors: Godes
authors: Pontus
authors: P
authors: Persson
authors: James
authors: J
authors: Bates
authors: Hajamohideen
authors: H
authors: Raffi
authors: Luca
authors: L
authors: Rampoldi
authors: Shinichi
authors: S
authors: Uchida
authors: Carsten
authors: C
authors: Hille
authors: Carsten
authors: C
authors: Dosche
authors: Satish
authors: S
authors: Kumar
authors: Maria
authors: M
authors: Castañeda-Bueno
authors: Gerardo
authors: G
authors: Gamba
authors: Sebastian
authors: S
authors: Bachmann
mesh_terms: D000818:Animals; D050034:Antidiuretic Agents / Q000494:pharmacology; D002461:Cell Line, Transformed; D002712:Chlorides / Q000378:metabolism*; D003894:Deamino Arginine Vasopressin / Q000494:pharmacology; D007686:Kidney Tubules, Distal / Q000378:metabolism*; D051379:Mice; D018345:Mice, Knockout; D009997:Osmotic Pressure; D010766:Phosphorylation / Q000187:drug effects / Q000502:physiology; D011817:Rabbits; D017483:Receptors, Vasopressin / Q000235:genetics / Q000378:metabolism; D028021:Sodium-Potassium-Chloride Symporters / Q000235:genetics / Q000378:metabolism*; D064466:Solute Carrier Family 12, Member 1; D058949:Uromodulin / Q000235:genetics / Q000378:metabolism*; D014982:Xenopus laevis
chemical_list: D050034:Antidiuretic Agents; D002712:Chlorides; D017483:Receptors, Vasopressin; C578715:Slc12a1 protein, mouse; D028021:Sodium-Potassium-Chloride Symporters; D064466:Solute Carrier Family 12, Member 1; C548613:Umod protein, mouse; D058949:Uromodulin; D003894:Deamino Arginine Vasopressin