Characterization of D150E and G196D aquaporin-2 mutations responsible for nephrogenic diabetes insipidus: importance of a mild phenotype.
Aquaporin-2 (AQP2) is a water channel responsible for the final water reabsorption in renal collecting ducts. Alterations in AQP2 function induce nephrogenic diabetes insipidus (NDI), a condition characterized by severe polyuria and polydipsia. Three patients affected with severe NDI, who were compound heterozygous for the AQP2 mutations D150E and G196D, are presented here along with a mildly affected D150E homozygous patient from another family. Using Xenopus oocytes as an expression system, these two mutations (G196D and D150E) were compared with the wild-type protein (AQP2-wt) for functional activity (water flux analysis), protein maturation, and plasma membrane targeting. AQP2-wt induces a major increase in water permeability (P(f) = 47.4 +/- 12.2 x 10(-4) cm/s) whereas D150E displays intermediate P(f) values (P(f) = 12.5 +/- 3.0 x 10(-4) cm/s) and G196D presents no specific water flux, similar to controls (P(f) = 2.1 +/- 0.8 x 10(-4) cm/s and 2.2 +/- 0.7 x 10(-4) cm/s, respectively). Western blot and immunocytochemical evaluations show protein targeting that parallels activity levels with AQP2-wt adequately targeted to the plasma membrane, partial targeting for D150E, and complete sequestration of G196D within intracellular compartments. When coinjecting AQP2-wt with mutants, no (AQP2-wt + D150E) or partial (AQP2-wt + G196D) reduction of water flux were observed compared with AQP2-wt alone, whereas complete loss of function was found when both mutants were coinjected. These results essentially recapitulate the clinical profiles of the family members, showing a typical dominant negative effect when G196D is coinjected with either AQP2-wt or D150E but not between AQP2-wt and D150E mutant.
Guyon
Cécile
C
Groupe d'Etude des Protéines Membranaires, Département de Physiologie, Université de Montréal, Montréal, Québec, Canada.
Lussier
Yoann
Y
Bissonnette
Pierre
P
Leduc-Nadeau
Alexandre
A
Lonergan
Michèle
M
Arthus
Marie-Françoise
MF
Perez
Rafael Bedoya
RB
Tiulpakov
Anatoly
A
Lapointe
Jean-Yves
JY
Bichet
Daniel G
DG
D000595:Amino Acid Sequence; D000818:Animals; D051399:Aquaporin 2 / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D002463:Cell Membrane Permeability; D048429:Cell Size; D018500:Diabetes Insipidus, Nephrogenic / Q000235:genetics* / Q000378:metabolism; D005260:Female; D020022:Genetic Predisposition to Disease; D006579:Heterozygote; D006720:Homozygote; D006801:Humans; D008297:Male; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D009865:Oocytes; D010375:Pedigree; D010641:Phenotype; D011487:Protein Conformation; D021381:Protein Transport; D012720:Severity of Illness Index; D013329:Structure-Activity Relationship; D014162:Transfection; D014867:Water / Q000378:metabolism; D014982:Xenopus laevis
C496023:AQP2 protein, human; D051399:Aquaporin 2; D014867:Water