Details

Autor(en) / Beteiligte

• Brumm, Harald
• Mühlhaus, Jessica
• Bolze, Florian
• Scherag, Susann
• Hinney, Anke
• Hebebrand, Johannes
• Wiegand, Susanna
• Klingenspor, Martin
• Grüters, Annette
• Krude, Heiko
• Biebermann, Heike

Titel

Rescue of Melanocortin 4 Receptor (MC4R) Nonsense Mutations by Aminoglycoside-Mediated Read-Through

Ist Teil von

• Obesity (Silver Spring, Md.), 2012-05, Vol.20 (5), p.1074-1081

Ort / Verlag

Oxford, UK: Blackwell Publishing Ltd

Erscheinungsjahr

2012

Link zum Volltext

Quelle

Wiley Online Library - AutoHoldings Journals

Beschreibungen/Notizen

• Aminoglycoside‐mediated read‐through of stop codons was recently demonstrated for a variety of diseases in vitro and in vivo. About 30 percent of human genetic diseases are the consequence of nonsense mutations. Nonsense mutations in obesity‐associated genes like the melanocortin 4 receptor (MC4R), expressed in the hypothalamus, show the impact of premature stop codons on energy homeostasis. Therefore, the MC4R could be a potential pharmaceutical target for obesity treatment and targeting MC4R stop mutations could serve as proof of principle for nonsense mutations in genes expressed in the brain. We investigated four naturally occurring nonsense mutations in the MC4R (W16X, Y35X, E61X, Q307X) located at different positions in the receptor for aminoglycoside‐mediated functional rescue in vitro. We determined localization and amount of full‐length protein before and after aminoglycoside treatment by fluorescence microscopy, cell surface and total enzyme linked immunosorbent assay (ELISA). Signal transduction properties were analyzed by cyclic adenosine monophosphate (cAMP) assays after transient transfection of MC4R wild type and mutant receptors into COS‐7 cells. Functional rescue of stop mutations in the MC4R is dependent on: (i) triplet sequence of the stop codon, (ii) surrounding sequence, (iii) location within the receptor, (iv) applied aminoglycoside and ligand. Functional rescue was possible for W16X, Y35X (N‐terminus), less successful for Q307X (C‐terminus) and barely feasible for E61X (first transmembrane domain). Restoration of full‐length proteins by PTC124 could not be confirmed. Future pharmaceutical applications must consider the potency of aminoglycosides to restore receptor function as well as the ability to pass the blood‐brain barrier.