Acute effects of desmin mutations on cytoskeletal and cellular integrity in cardiac myocytes

Abstract

Mutations in desmin have been associated with a subset of human myopathies. Symptoms typically appear in the second to third decades of life, but in the most severe cases can manifest themselves earlier. How desmin mutations lead to aberrant muscle function, however, remains poorly defined. We created a series of four mutations in rat desmin and tested their in vitro filament assembly properties. RDM-G, a chimera between desmin and green fluorescent protein, formed protofilament-like structures in vitro. RDM-1 and RDM-2 blocked in vitro assembly at the unit-length filament stage, while RDM-3 had more subtle effects on assembly. When expressed in cultured rat neonatal cardiac myocytes via adenovirus infection, these mutant proteins disrupted the endogenous desmin filament to an extent that correlated with their defects in in vitro assembly properties. Disruption of the desmin network by RDM-1 was also associated with disruption of plectin, myosin, and alpha-actinin organization in a significant percentage of infected cells. In contrast, expression of RDM-2, which is similar to previously characterized human mutant desmins, took longer to disrupt desmin and plectin organization and had no significant effect on myosin or alpha-actinin organization over the 5-day time course of our studies. RDM-3 had the mildest effect on in vitro assembly and no discernable effect on either desmin, plectin, myosin, or alpha-actinin organization in vivo. These results indicate that mutations in desmin have both direct and indirect effects on the cytoarchitecture of cardiac myocytes.

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

“…Nevertheless, all desmin mutants revealing defective assembly properties were dominant because, in 1:1 mixtures with WT desmin, they led to aberrant assembly as demonstrated by viscometry and cDNA-transfection into vimentin-containing 3T3 fibroblasts (data not shown). Consistent with these data, the disruption of the desmin cytoskeleton in cultured rat neonatal cardiac myocytes by overexpression of an engineered desmin mutant with an arginine to cysteine change in coil 1B has demonstrated that also in an authentic desmin environment such mutants exhibit a dominant effect (34).…”

Designing Heart Performance by Gene Transfer

“…Desmin mutations have been linked to both dilated (482,584) and restrictive cardiomyopathies (20,712,930). Gene transfer of mutated desmin into neonatal cardiac myocytes disrupted the sarcomeric banding pattern (341). These studies demonstrate that alterations in desmin filament assembly have a dominant effect on cellular sarcomeric assembly, an observation consistent with the dominant mode of inheritance that characterizes desminrelated cardiomyopathies.…”

Splice site, frameshift, and chimericGFAPmutations in Alexander disease

“…The de novo nature of the mutation in our patient is strong evidence that it is disease-causing, and we have also found that the mutant GFAP fails to polymerize normally in transiently transfected cells, and is dominant to the wild type. These polymerization properties of the extended, mutant GFAP add further caution to the use of C-terminal tags of intermediate filament proteins (Haubold et al, 2003). …”