title: Characterization of the melanocortin-4-receptor nonsense mutation W16X in vitro and in vivo.
abstract: Several genetic diseases are triggered by nonsense mutations leading to the formation of truncated and defective proteins. Aminoglycosides have the capability to mediate a bypass of stop mutations during translation thus resulting in a rescue of protein expression. So far no attention has been directed to obesity-associated stop mutations as targets for nonsense suppression. Herein, we focus on the characterization of the melanocortin-4-receptor (MC4R) nonsense allele W16X identified in obese subjects. Cell culture assays revealed a loss-of-function of Mc4r(X16) characterized by impaired surface expression and defect signaling. The aminoglycoside G-418 restored Mc4r(X16) function in vitro demonstrating that Mc4r(X16) is susceptible to nonsense suppression. For the evaluation of nonsense suppression in vivo, we generated a Mc4r(X16) knock-in mouse line by gene targeting. Mc4r(X16) knock-in mice developed hyperphagia, impaired glucose tolerance, severe obesity and an increased body length demonstrating that this new mouse model resembles typical characteristics of Mc4r deficiency. In a first therapeutic trial, the aminoglycosides gentamicin and amikacin induced no amelioration of obesity. Further experiments with Mc4r(X16) knock-in mice will be instrumental to establish nonsense suppression for Mc4r as an obesity-associated target gene expressed in the central nervous system.
authors.affiliation: Technische Universität München, Molecular Nutritional Medicine, Else Kröner-Fresenius Center and ZIEL-Research Center for Nutrition and Food Science, Freising, Germany. bolze@wzw.tum.de
authors.forename: F
authors.initials: F
authors.lastname: Bolze
authors.forename: N
authors.initials: N
authors.lastname: Rink
authors.forename: H
authors.initials: H
authors.lastname: Brumm
authors.forename: R
authors.initials: R
authors.lastname: Kühn
authors.forename: S
authors.initials: S
authors.lastname: Mocek
authors.forename: A-E
authors.initials: AE
authors.lastname: Schwarz
authors.forename: C
authors.initials: C
authors.lastname: Kless
authors.forename: H
authors.initials: H
authors.lastname: Biebermann
authors.forename: W
authors.initials: W
authors.lastname: Wurst
authors.forename: J
authors.initials: J
authors.lastname: Rozman
authors.forename: M
authors.initials: M
authors.lastname: Klingenspor
mesh_terms: D000617:Aminoglycosides / Q000235:genetics / Q000378:metabolism; D000818:Animals; D001823:Body Composition / Q000235:genetics; D001831:Body Temperature / Q000235:genetics; D001835:Body Weight / Q000235:genetics; D019556:COS Cells; D002460:Cell Line; D002522:Chlorocebus aethiops; D018389:Codon, Nonsense; D002149:Energy Intake / Q000235:genetics; D015870:Gene Expression / Q000235:genetics; D057809:HEK293 Cells; D006801:Humans; D007031:Hypothalamus / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D009765:Obesity / Q000235:genetics / Q000378:metabolism; D044105:Receptor, Melanocortin, Type 4 / Q000235:genetics* / Q000378:metabolism*
chemical_list: D000617:Aminoglycosides; D018389:Codon, Nonsense; D044105:Receptor, Melanocortin, Type 4