The Effect of Eight V2 Vasopressin Receptor Mutations on Stimulation of Adenylyl Cyclase and Binding to Vasopressin
| Title: | The Effect of Eight V2 Vasopressin Receptor Mutations on Stimulation of Adenylyl Cyclase and Binding to Vasopressin |
|---|---|
| Authors: | Pan, Yang; Wilson, Paul; Gitschier, Ph.D., Jane |
| Publisher: | Journal of Biological Chemistry |
| Date Published: | December 16, 1994 |
| Reference Number: | 83 |
We previously identified six V2 vasopressin receptor mutations in five unrelated nephrogenic diabetes insipidus(NDI) families. In order to elucidate the effect of these mutations on the function of the V2 vasopressin receptor, we introduced these six and two additional, naturally occurring mutations into the V2 vasopressin receptor gene byin vitro mutagenesis. Five of the mutants (two frameshift, one nonsense, and two missense) failed to stimulate adenylyl cyclase due to their inability to bind vasopressin under the experimental conditions. In contrast, ligandbinding and cAMP accumulation were normal for two other mutations, a A61V missense mutation and an in-frame deletion of four amino acids (Arg-247 to Gly-250), suggesting that they are not the cause of NDI in these families. The deletion mutation was found in a family in conjunction with a second mutation, R181C, which yielded a much reduced ligand-binding capacity. The KD of R181C was at least 26 times higher than that of the wild type. Furthercharacterization by an immunofluorescent assay showed that the R181C mutant receptor is expressed and distributed on the cell surface in a manner similar to that of the wild type. This finding indicates that the inability of this mutant to stimulate adenylyl cyclase is caused by the reduced capacity for vasopressin binding and that the R181C mutation is responsible for NDI in this family.
|
You may, however, read this article at the Journal of Biological Chemistry website. In order to view this document, you will need Acrobat Reader. If you do not already have Acrobat Reader or need to upgrade, click here. To return to this page, use your "back" key. |
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)
The most common form of inherited NDI is caused by mutations in the V2R gene. Yang Pan, et al., analyzed the V2Rs of eight mutated V2R genes in order to determine how each mutation affected the structure -- and therefore the ability to function -- of the V2Rs each synthesized. As indices of the V2Rs' functional ability, the researchers measured the V2Rs' ability to increase cAMP and to bind with AVP.
The authors found that six of eight of the V2R gene mutations produced V2Rs that could not increase cAMP. This inability was caused because, though these V2Rs could travel to the cell surface, which is the AVP/V2R binding site, they could not bind with AVP once they were there. Since they could not bind with AVP, the molecular sequence that leads to water reabsorption and urine concentrationwhen AVP and V2R bind could not take place.
However, one of the remaining two mutations had no effect on cAMP production, nor did it affect the V2Rs' ability to bind with AVP. This suggested that this particular V2R gene mutation was not the cause of NDI in the person who bore it. The researchers speculated that this patient's NDI may be due to a mutation in another gene.
One of the patients had two mutations in his V2R genes. One fell in the first group of six mutations as it resulted in non-functional V2Rs incapable of binding with AVP. The other mutation could generate V2Rs in laboratory cell cultures that could increase cAMP production.
Researchers are able to make such fine distinctions (e.g., being able to discern which of the two mutations in one NDI patient's V2R gene is responsible for his NDI) due to the growing body of knowledge concerning the relationship between the V2R's structure and its function. Researchers have deduced the structure of the V2R (You can look at a diagram of a V2R here) and they are beginning to discover roles its different sections play in V2R function. For example, a disulfide bond exists between the first and second extracellular loops of the V2R. A structural flaw in either of these loops could disrupt the bond, which may be essential for the V2R tofold itself into the right shape. Researchers hope that further research will lead to effective therapeutic approaches to NDI.