title: Functional characterization and cell immunolocalization of AQP-CD water channel in kidney collecting duct.
abstract: Vasopressin-regulated water permeability of the kidney collecting duct is a key component of the urine concentration machinery. Recently, a cDNA for AQP-CD, the vasopressin-regulated water channel, initially reported as WCH-CD, has been isolated (K. Fushimi, S. Uchida, Y. Hara, Y. Hirata, F. Marumo, and S. Sasaki. Nature Lond. 361: 549-552, 1993). AQP-CD was expressed in oocyte membrane using a Xenopus expression vector, and functional characteristics of AQP-CD were examined. Osmotic water permeability (Pf) of oocytes expressing AQP-CD was 138 +/- 19 microns/s (mean +/- SE), 12 times greater than the control (11 +/- 3 microns/s), 90% inhibited by 0.3 mM HgCl2, and weakly temperature dependent (energy of activation for Pf was 4.0 kcal/mol). Urea influx measured from 15-min [14C]urea uptake by oocytes injected with AQP-CD/expression vector 1 cRNA was 86 +/- 17% of the control. Two-electrode voltage-clamp experiments revealed insignificant ion conductance of AQP-CD. Immunoblots of membranes from rat kidney medulla and oocytes expressing AQP-CD using anti-AQP-CD COOH-terminal antibody showed a 29-kDa protein and 35- to 50-kDa high-molecular-mass forms. Immunohistochemistry showed apical and subapical localization of AQP-CD in the collecting duct principal cells. Our results indicated that AQP-CD is a 29-kDa protein, a selective water channel, distinct from a urea channel, and localized to the membranes of vasopressin-sensitive components in kidney collecting duct principal cells.
authors.affiliation: Second Department of Internal Medicine, Tokyo Medical and Dental University, Japan.
authors.forename: K
authors.initials: K
authors.lastname: Fushimi
authors.forename: S
authors.initials: S
authors.lastname: Sasaki
authors.forename: T
authors.initials: T
authors.lastname: Yamamoto
authors.forename: M
authors.initials: M
authors.lastname: Hayashi
authors.forename: T
authors.initials: T
authors.lastname: Furukawa
authors.forename: S
authors.initials: S
authors.lastname: Uchida
authors.forename: M
authors.initials: M
authors.lastname: Kuwahara
authors.forename: K
authors.initials: K
authors.lastname: Ishibashi
authors.forename: M
authors.initials: M
authors.lastname: Kawasaki
authors.forename: I
authors.initials: I
authors.lastname: Kihara
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D051398:Aquaporin 1; D020346:Aquaporins; D002463:Cell Membrane Permeability; D004553:Electric Conductivity; D005260:Female; D005455:Fluorescent Antibody Technique; D007150:Immunohistochemistry; D007473:Ion Channels / Q000032:analysis / Q000096:biosynthesis / Q000502:physiology*; D007672:Kidney Cortex / Q000502:physiology*; D007679:Kidney Medulla / Q000502:physiology*; D007685:Kidney Tubules, Collecting / Q000166:cytology / Q000502:physiology* / Q000648:ultrastructure; D008564:Membrane Potentials; D016253:Microscopy, Immunoelectron; D008961:Models, Structural; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009865:Oocytes / Q000187:drug effects / Q000502:physiology*; D017433:Protein Structure, Secondary; D006180:Proton-Translocating ATPases / Q000032:analysis; D018075:RNA, Complementary; D051381:Rats; D017207:Rats, Sprague-Dawley; D014982:Xenopus laevis
chemical_list: C496022:Aqp1 protein, rat; D020346:Aquaporins; D007473:Ion Channels; D018075:RNA, Complementary; D051398:Aquaporin 1; D006180:Proton-Translocating ATPases