title: TMEM16A is a Ca(2+) -activated Cl(-) channel expressed in the renal collecting duct.
abstract: OBJECTIVE
  In the renal collecting ducts, ATP stimulates a Ca(2+) -activated chloride current. The identity of the channel responsible for the current under physiological conditions is not known and it was hypothesized that TMEM16a is a relevant candidate in the renal collecting duct.
  
  
  METHODS
  The cortical collecting duct cell line M-1 was used as a model of the collecting duct. The ATP induced Ca(2+) signalling was imaged in cells loaded with Ca(2+) -sensitive fluorescent probes using confocal laser-scanning fluorescence microscopy. Chloride current was determined by mounting M-1 cell layers in Ussing chamber. The expression of TMEM16a in human kidney was tested by immunohistochemistry.
  
  
  RESULTS
  M-1 cells displayed a transient increase in intracellular Ca(2+) concentration in response to 100 nm ATP. This response was completely blocked by addition of 100 μm suramin, indicating that ATP signals through purinergic P2 receptors. Apical addition of 100 nm ATP induced a Cl(-) current, which was blocked by suramin, DPC and the cysteine-modifying compound MTSET. M-1 cells were found to express TMEM16a at the mRNA and protein level. Functionally, it was found that knock-down of TMEM16a expression in M-1 cells inhibited the ATP induced Cl(-) -current. In human and mouse kidney sections, TMEM16a protein expression was localized to the collecting duct, and TMEM16a was found to be excreted in human urinary exosomes.
  
  
  CONCLUSIONS
  TMEM16a is a Ca(2+) -activated Cl(-) channel expressed in the collecting ducts.
authors.affiliation: Department of Cardiovascular and Renal Research, Institute of Molecular Medicine, University of Southern Denmark, Odense, Denmark.
authors.forename: P
authors.initials: P
authors.lastname: Svenningsen
authors.forename: M R
authors.initials: MR
authors.lastname: Nielsen
authors.forename: N
authors.initials: N
authors.lastname: Marcussen
authors.forename: S
authors.initials: S
authors.lastname: Walter
authors.forename: B L
authors.initials: BL
authors.lastname: Jensen
mesh_terms: D000818:Animals; D000075369:Anoctamin-1; D015153:Blotting, Western; D002460:Cell Line; D018118:Chloride Channels / Q000378:metabolism*; D055785:Gene Knockdown Techniques; D006801:Humans; D007150:Immunohistochemistry; D007685:Kidney Tubules, Collecting / Q000378:metabolism*; D051379:Mice; D018613:Microscopy, Confocal; D009363:Neoplasm Proteins / Q000378:metabolism*; D034741:RNA, Small Interfering; D020133:Reverse Transcriptase Polymerase Chain Reaction
chemical_list: C476213:ANO1 protein, human; C531181:ANO1 protein, mouse; D000075369:Anoctamin-1; D018118:Chloride Channels; D009363:Neoplasm Proteins; D034741:RNA, Small Interfering
keywords: ATP; calcium-activated chloride channels; collecting duct; exosome; kidney; purinergic signalling