title: Impact of disease mutations on the desmin filament assembly process.
abstract: It has been documented that mutations in the human desmin gene lead to a severe type of myofibrillar myopathy, termed more specifically desminopathy, which affects cardiac and skeletal as well as smooth muscle. We showed recently that 14 recombinant versions of these disease-causing desmin variants, all involving single amino acid substitutions in the alpha-helical rod domain, interfere with in vitro filament formation at distinct stages of the assembly process. We now provide mechanistic details of how these mutations affect the filament assembly process by employing analytical ultracentrifugation, time-lapse electron microscopy of negatively stained and glycerol-sprayed/low-angle rotary metal-shadowed samples, quantitative scanning transmission electron microscopy, and viscometric studies. In particular, the soluble assembly intermediates of two of the mutated proteins exhibit unusually high s-values, compatible with octamers and other higher-order complexes. Moreover, several of the six filament-forming mutant variants deviated considerably from wild-type desmin with respect to their filament diameters and mass-per-length values. In the heteropolymeric situation with wild-type desmin, four of the mutant variants caused a pronounced "hyper-assembly", when assayed by viscometry. This indicates that the various mutations may cause abortion of filament formation by the mutant protein at distinct stages, and that some of them interfere severely with the assembly of wild-type desmin. Taken together, our findings provide novel insights into the basic intermediate filament assembly mechanisms and offer clues as to how amino acid changes within the desmin rod domain may interfere with the normal structural organization of the muscle cytoskeleton, eventually leading to desminopathy.
authors.affiliation: Department of Cardiology, University of Heidelberg, D-69120 Heidelberg, Germany.
authors.forename: Harald
authors.initials: H
authors.lastname: Bär
authors.forename: Norbert
authors.initials: N
authors.lastname: Mücke
authors.forename: Philippe
authors.initials: P
authors.lastname: Ringler
authors.forename: Shirley A
authors.initials: SA
authors.lastname: Müller
authors.forename: Laurent
authors.initials: L
authors.lastname: Kreplak
authors.forename: Hugo A
authors.initials: HA
authors.lastname: Katus
authors.forename: Ueli
authors.initials: U
authors.lastname: Aebi
authors.forename: Harald
authors.initials: H
authors.lastname: Herrmann
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D003893:Desmin / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure; D006801:Humans; D007382:Intermediate Filaments / Q000378:metabolism* / Q000648:ultrastructure; D051379:Mice; D017348:Microscopy, Electron, Scanning Transmission; D009135:Muscular Diseases / Q000235:genetics*; D009154:Mutation; D009210:Myofibrils / Q000378:metabolism / Q000648:ultrastructure; D011485:Protein Binding; D014461:Ultracentrifugation; D014783:Viscosity
chemical_list: D003893:Desmin