Abstract 15252: Desmin A191T Mutation Aggravates δ-Sarcoglycan-Deficient Cardiomyopathy in TO-2 Hamster
Introduction: Cardiomyopathic hamsters have distinct sublines, among which BIO14.6 exhibits hypertrophic cardiomyopathy (CM) whereas TO-2 manifests severe dilated CM. An additional mutation aggravating CM should then exist in TO-2, since both hamsters share genetic loss of δ-sarcoglycan, a member of dystrophin-associated proteins (DAPs). Here we searched for such mutation in TO-2.
Methods: Male TO-2, BIO14.6 and normal hamsters were used (6, 12, 28, 45 week old, n=6 in each group). Ventricular ultrastructure was examined with transmission electron microscope and Z-band protein was biochemically measured. We isolated cDNAs for desmin, an intermediate filament specific to Z-band, and examined phosphorylation of TO-2 desmin with antibodies raised against the site-specific phosphopeptide. Filament formation of TO-2 desmin was explored by expressing its cDNA in SW13 cells. Pulldown assay determined the binding domain of desmin for desmuslin, another member of DAPs. Expression of DAPs including desmuslin was immunohistochemically examined along CM development.
Results: The Z-band appeared dim in TO-2 compared with that in BIO14.6 at the same age. In TO-2, desmin was progressively lost whereas α-actinin remained intact. While all the hamster desmin cDNAs harbored open reading frames of 1,407 nucleotides, guanine at the 571st nucleotide was replaced by adenine only in TO-2, resulting in substitution of alanine at the 191st amino acid in the coil-1 domain to threonine. This A191T substitution created a potential phosphorylation site for protein kinase C, but such phosphorylation was not detected in vivo. In SW13 cells, the transfected TO-2 desmin formed visible filaments with fragile structure and sparse distribution. The N-terminal head domain of desmin bound to desmuslin in vitro. Concordantly in TO-2 ventricles, desmuslin and other DAPs were reduced along with the gradual decrease in desmin.
Conclusions: We found TO-2 has A191T mutation in desmin, one of the known causative genes for CM. This natural mutation was considered to weaken the proper formation of desmin filament, causing the desmin/DAP complex more susceptible to mechanical stress in vivo. In conclusion, our data refined the molecular pathogenesis of TO-2, a widely used model for severe CM.
- © 2011 by American Heart Association, Inc.