Vasopressin V2 Receptor Mutants that Cause X-linked Nephrogenic Diabetes Insipidus: Analysis of Expression, Processing, and Function

Title: Vasopressin V2 Receptor Mutants that Cause X-linked Nephrogenic Diabetes Insipidus: Analysis of Expression, Processing, and Function
Authors: Oksche, Alexander; Schulein, Ralf; Rutz, Claudia; Liebenhoff, Ursula; Dickson, MD, John; Muller, Helmut; Birnbaumer, Mariel; Rosenthal, Walter
Publisher: Molecular Pharmacology
Date Published: October 01, 1996
Reference Number: 15
We investigated the biochemical and functional properties of five vasopressin V2 receptor mutants (L44F, L44P, W164S, S167L, and S167T) that were recently described in families with a history of X-linked nephrogenic diabetes insipidus. COS.M6 cells transfected with cDNA encoding these mutants acquired < 4% specific [3H] arginine vasopressin (AVP) binding sites on the cell surface in comparison with cells transfected with cDNA coding for the wild-type receptor. Membrane preparations from COS.M6 cells or human embryonic kidney 293 cells expressing these mutants did not respond with an increase in adenylyl cyclase activity in response to AVP, which is in contrast to membranes from cells expressing the wild-type. By analyzing fusion proteins of the V2 receptor and Escherichia coli alkaline phosphatase attached to the carboxyl terminus of the receptor moiety, we found that the mutants L44P, W164S, S167L, and S167T lacked complex glycosylation and were expressed at low levels. The data suggest that the mutants L44P, W164S, S176T, and S167L are misfolded and therefore retained within the endoplasmic reticulum and degraded. In contrast, the fusion proteins carrying the mutant L44F and the in vitro mutant S167A were expressed in their mature form at wild-type levels; however, only the mutant S167A was functionally active. Site-directed mutagenesis of S167 revealed that elimination of the endogenous hydroxyl group (S167A) yielded a protein with properties identical to those of the wild-type receptor, whereas both the introduction of a methyl group (S167T) and the replacement of the hydroxyl group by an isopropyl group (S167L) profoundly disturbed receptor processing. The data show that minute changes at codon 167 nearly abolish expression of a mature protein, thus defining structural requirements of this codon.

This translation by the NDI Foundation is to assist the lay reader. To provide a clear, accessible interpretation of the original article, we eliminated or simplified some technical detail and complicated scientific language. We concentrated our translation on those aspects of the article dealing directly with NDI. The NDI Foundation thanks the researchers for their work toward understanding and more effectively treating this disorder.
© Copyright NDI Foundation 2007 (JC)

In nephrogenic diabetes insipidus (NDI), the kidney fails to concentrate urine (i.e., the kidneys pass too much water into the urine) despite normal or elevated levels of the hormone arginine vasopressin (AVP). AVP tells certain kidney cells to retain water and direct it into the blood stream rather than passing it into the urine and expelling it as waste. To do this, AVP works with vasopressin V2 receptors which are found on the walls of certain kidney cells. It is now well established that most, if not all, cases of X-linked recessive NDI are caused by inactive V2 receptor gene mutations.

In this study, the properties of five V2 receptor mutants were analyzed: L44F, L44P, W164S, S167L, and S167T.

The L44F and L44P mutations were found in two unrelated families with X-linked NDI. The S167L mutation caused X-linked NDI in at least seven unrelated families. In addition, the authors found an affected family with an S167T mutation.

Oksche, et al., biochemically demonstrated that the five naturally occurring point mutations in the V2 receptors (L44F, L44P, W164S, S167L, and S167T), found in families with X-linked NDI, are responsible for the disease. (Point mutations result from a change in the DNA molecule.) These mutant receptors are not able to bind to certain cells and, therefore, cannot do the job they are supposed to do.

The authors showed that four of five X-linked NDI mutations which were analyzed severely disturbed protein processing. If cells do not get the protein they need, they cannot function as they should.