Severe muscle disease-causing desmin mutations interfere with in vitro filament assembly at distinct stages

Abstract

Desmin is the major intermediate filament (IF) protein of muscle.Recently, mutations of the desmin gene have been reported to cause familial or sporadic forms of human skeletal, as well as cardiac, myopathy, termed desmin-related myopathy (DRM). The impact of any of these mutations on filament assembly and integration into the cytoskeletal network of myocytes is currently not understood, despite the fact that all cause the same histopathological defect, i.e., desmin aggregation. To gain more insight into the molecular basis of this process, we investigated how mutations within the ␣-helical rod domain of desmin affect both the assembly of the recombinant protein in vitro as well as the filament-forming capacity in cDNA-transfected cells. Whereas 6 of 14 mutants assemble into seemingly normal IFs in the test tube, the other mutants interfere with the assembly process at distinct stages, i.e., tetramer formation, unit-length filament (ULF) formation, filament elongation, and IF maturation. Correspondingly, the mutants with in vitro assembly defects yield dot-like aggregates in transfected cells, whereas the mutants that form IFs constitute a seemingly normal IF cytoskeleton in the cellular context. At present, it is entirely unclear why the latter mutant proteins also lead to aggregate formation in myocytes. Hence, these findings may be a starting point to dissect the contribution of the individual subdomains for desmin pathology and, eventually, the development of therapeutic interventions.desmin-related myopathy ͉ intermediate filaments

Intermediate Filament Diseases: Desminopathy

“…Although p.Ala213Val desmin created a filamentous network in SW13 cells and preserved the existing network in the C2C12 cells, 40 filament assembly experiments showed it aggregated in the viscometer; in the BMGE +H cells, the p.Ala213Val filaments were bundle-like, suggesting that the pathomechanisms of this mutation probably involve subtle but critical interactions with non-IF components in muscle cells. 61 A heterozygous single-nucleotide (adenine) insertion mutation occurring at the third position of codon 241 causes a frameshift leading to serial amino acid replacements: Val242Glu, His243Ser, Glu244Ala and eventually a premature termination signal at codon 245 (numbering according to the updated sequence, GenBank submission # AF167579). This mutation is predicted to create a truncated desmin molecule with molecular weight of 27 kDa.…”

“…40,61 However, when the p.Ala360Pro and p.Asn393Ile mutations were cotransfected, this caused devastating effects in each cell line used in the experiment. 40 Indeed, in a family segregating both p.Ala360Pro and p.Asn393Ile mutations, a highly aggressive early onset cardioskeletal myopathy affected only those having both mutations in a compound heterozygous fashion but spared the carriers of either mutation.…”

“…61 Analysis of filament assembly behavior demonstrates that some desmin mutants do not prevent filament formation (p.Ala213Val, p.Glu245Asp, p.Ala360Pro, p.Glu389Pro, p.Asn393Ile, p.Asp399Tyr). Some others interfere with the assembly process at distinct stages and these are classified into three groups: (1) mutations compromising longitudinal annealing properties (p.Leu385Pro, p.Arg406Trp); (2) mutants with enhanced adhesiveness leading to filament aggregation (p.Ala337Pro, p.Asn342Pro, p.Ala357Pro); and (3) mutants showing rapid disintegration of assembly precursors (p.Leu345Pro, p.Arg350Pro, p.Leu370Pro).…”

Monitoring intermediate filament assembly by small-angle x-ray scattering reveals the molecular architecture of assembly intermediates

“…The next major challenge is to establish the molecular mechanism of these diseases and, ultimately, to develop efficient treatment approaches. Here, the in vitro-derived structural information on the IF architecture and assembly is of utmost importance, as seen, e.g., in the recent studies on desminopathies (35)(36)(37).…”

The desmin coil 1B mutation K190A impairs nebulin Z-disc assembly and destabilizes actin thin filaments

“…Although these aggregates are mainly composed of desmin, many sarcomere proteins such as actin, titin, myosin and nebulin have also been detected (Schroder et al, 2007). Interestingly, although mutations have been identified in all domains of desmin, no obvious correlation between the localization of an individual mutation within desmin and its effect on protein assembly has been recognized (Bar et al, 2005;Hong et al, 2011;Taylor et al, 2007;Vernengo et al, 2010).…”