title: cDNA and genomic cloning of mouse aquaporin-2: functional analysis of an orthologous mutant causing nephrogenic diabetes insipidus.
abstract: As the first step in generating a transgenic mouse model of nephrogenic diabetes insipidus (NDI), we have analyzed the mouse aquaporin-2 (Aqp2) cDNA and gene and generated a mutated Aqp2 orthologous to NDI-causing human AQP2-T126M. Aqp2 cDNA was isolated from mouse kidney and encoded a 271-amino-acid protein with 90.4% identity to human AQP2. Expression in Xenopus oocytes indicated that Aqp2 encoded a mercurial-sensitive, water-selective channel. Northern blot analysis showed a single 1.7-kb Aqp2 transcript expressed only in kidney (medulla > cortex); transcript expression was increased approximately 20-fold in 48-h water-deprived mice. Immunoblot analysis revealed a 29-kDa glycoprotein in mouse kidney. Sequence comparison of the Aqp2 cDNA with a 5.5-kb mouse genomic DNA indicated three introns (lengths 2.4, 0.9, and 0.6 kb) separating four exons with boundaries at amino acids 120, 175, and 202. Genomic Southern blot analysis revealed a single-copy Aqp2 gene. The mutant Aqp2-T126M was water permeable when expressed in Xenopus oocytes, but was retained at the endoplasmic reticulum (ER) in transfected mammalian cells. The chemical chaperone glycerol produced a redistribution of Aqp2-T126M from ER to plasma membrane/endosomes. These results establish a basis for an Aqp2-T126M transgenic knock-in model of NDI.
authors: Department of Physiology, Cardiovascular Research Institute, San Francisco, California, 94143-0521, USA.
authors: B
authors: B
authors: Yang
authors: T
authors: T
authors: Ma
authors: Z
authors: Z
authors: Xu
authors: A S
authors: AS
authors: Verkman
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D051399:Aquaporin 2; D051403:Aquaporin 6; D020346:Aquaporins / Q000235:genetics*; D001483:Base Sequence; D015152:Blotting, Northern; D015139:Blotting, Southern; D016466:CHO Cells; D003001:Cloning, Molecular; D006224:Cricetinae; D018500:Diabetes Insipidus, Nephrogenic / Q000235:genetics*; D005091:Exons; D015870:Gene Expression; D015151:Immunoblotting; D051379:Mice; D008957:Models, Genetic; D008969:Molecular Sequence Data; D009865:Oocytes / Q000166:cytology / Q000378:metabolism; D013997:Time Factors; D014018:Tissue Distribution; D014981:Xenopus / Q000378:metabolism
chemical_list: C496024:Aqp2 protein, mouse; D051399:Aquaporin 2; D051403:Aquaporin 6; D020346:Aquaporins