title: Disease-proportional proteasomal degradation of missense dystrophins.
abstract: The 427-kDa protein dystrophin is expressed in striated muscle where it physically links the interior of muscle fibers to the extracellular matrix. A range of mutations in the DMD gene encoding dystrophin lead to a severe muscular dystrophy known as Duchenne (DMD) or a typically milder form known as Becker (BMD). Patients with nonsense mutations in dystrophin are specifically targeted by stop codon read-through drugs, whereas out-of-frame deletions and insertions are targeted by exon-skipping therapies. Both treatment strategies are currently in clinical trials. Dystrophin missense mutations, however, cause a wide range of phenotypic severity in patients. The molecular and cellular consequences of such mutations are not well understood, and there are no therapies specifically targeting this genotype. Here, we have modeled two representative missense mutations, L54R and L172H, causing DMD and BMD, respectively, in full-length dystrophin. In vitro, the mutation associated with the mild phenotype (L172H) caused a minor decrease in tertiary stability, whereas the L54R mutation associated with a severe phenotype had a more dramatic effect. When stably expressed in mammalian muscle cells, the mutations caused steady-state decreases in dystrophin protein levels inversely proportional to the tertiary stability and directly caused by proteasomal degradation. Both proteasome inhibitors and heat shock activators were able to increase mutant dystrophin to WT levels, establishing the new cell lines as a platform to screen for potential therapeutics personalized to patients with destabilized dystrophin.
authors.affiliation: Department of Biochemistry, Molecular Biology, and Biophysics, University of Minnesota-Twin Cities, Minneapolis, MN 55455.
authors.forename: Dana M
authors.initials: DM
authors.lastname: Talsness
authors.affiliation: Department of Biochemistry, Molecular Biology, and Biophysics, University of Minnesota-Twin Cities, Minneapolis, MN 55455.
authors.forename: Joseph J
authors.initials: JJ
authors.lastname: Belanto
authors.affiliation: Department of Biochemistry, Molecular Biology, and Biophysics, University of Minnesota-Twin Cities, Minneapolis, MN 55455 jervasti@umn.edu.
authors.forename: James M
authors.initials: JM
authors.lastname: Ervasti
mesh_terms: D000818:Animals; D015153:Blotting, Western; D002460:Cell Line; D016189:Dystrophin / Q000235:genetics / Q000378:metabolism*; D006360:Heat-Shock Proteins / Q000378:metabolism; D006801:Humans; D036701:Limonins / Q000494:pharmacology; D018485:Muscle Fibers, Skeletal / Q000187:drug effects / Q000378:metabolism; D020388:Muscular Dystrophy, Duchenne / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D020125:Mutation, Missense; D010641:Phenotype; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism*; D061988:Proteasome Inhibitors / Q000494:pharmacology; D055550:Protein Stability; D059748:Proteolysis; D020133:Reverse Transcriptase Polymerase Chain Reaction
chemical_list: D016189:Dystrophin; D006360:Heat-Shock Proteins; D036701:Limonins; D061988:Proteasome Inhibitors; C106014:gedunin; D046988:Proteasome Endopeptidase Complex
keywords: Duchenne muscular dystrophy; heat shock activator; missense mutation; proteasome inhibitor; protein folding