2002 Global Researcher Conference Proceeding
April 26 - 28, 2002
| Conference: | 2002 Global Researcher Conference |
|---|---|
| Title: | Functional rescue of truncated V2-Vasopressin-receptors by aminoglycoside-induced misreading of nonsense-mutations |
| Authors: | Sangkuhl, Katrin; Schulz, Angela; Schultz, Prof. Dr. Med. Gunter; Schoneberg, Torsten |
| Institutions: | University of Leipzig, Freie University of Berlin |
About 15% out of all mutations found in patients with X chromosome-linked nephrogenic diabetes insipidus (NDI) result in a truncation and complete loss of V2 vasopressin receptor (V2R) function due to nonsense mutations. It has been demonstrated in vitro that aminoglycoside antibiotics can increase the frequency of erroneous misreading of premature nonsense codons thereby permitting the translation of alleles carrying nonsense mutations to continue reading to the end of the gene. In our study we demonstrate that pretreatment of transiently transfected COS-7 cells with several aminoglycosides partially restored function of clinically relevant nonsense codon-truncated V2R. Additionally, immunological studies verified the existence of full-length V2R molecules after aminoglycoside pretreatment. The property of functional reconstitution was obviously independent from cytotoxicity of the chosen aminoglycosides. Interestingly, an exposition period with the aminoglycoside geneticin of less than 3 hours resulted in a significant gain of receptor function. It was noted that the efficiency of this potential therapeutic strategy depends on the precise position of the stop mutation within the V2R gene.
To test the therapeutic usefulness of this approach, primary cultures of collecting tube cells were established from kidneys of wild-type and heterozygous female mice which carry a nonsense mutation in the V2R gene known to cause X chromosome-linked NDI. We demonstrated that aminoglycoside pretreatment is effective to rescue function of nonsense mutation-truncated V2R due to premature stop codon misreading in primary cultured cells from these knock-in mice. This approach may lead to additional therapeutic options for some NDI patients.
There is a mutation called a nonsense mutation. It produces a protein that is not fully developed because one of the sections of the gene that signals when the protein is to end occurs earlier in the genetic message than it should. This causes a premature termination of the transcription process and results in the release of an incomplete, generally nonfunctional protein.
About 15% of X-linked NDI mutations are nonsense mutations. Working with laboratory cell cultures, researchers found that aminoglycoside antibiotics applied to genes with nonsense mutations can increase a gene's likelihood of misreading its genetic message such that the protein synthesis is allowed to complete. Sangkuhl, et al., tested to see if using aminoglycoside could have a therapeutic value for patients with NDI caused by nonsense mutations. Working with the kidney cells of mice with this type of mutation, they determined that pretreatment with aminoglcoside effectively restored the function of vasopressin 2 receptor protein. They suggest that this approach may provide a treatment option for those NDI patients with certain NDI nonsense mutations.