Beta-synemin expression in cardiotoxin-injected rat skeletal muscle

Background: β-synemin was originally identified in humans as an α-dystrobrevin-binding protein through a yeast two-hybrid screen using an amino acid sequence derived from exons 1 through 16 of α-dystrobrevin, a region common to both α-dystrobrevin-1 and -2. α-Dystrobrevin-1 and -2 are both expressed in muscle and co-localization experiments have determined which isoform preferentially functions with β-synemin in vivo. The aim of our study is to show whether each α-dystrobrevin isoform has the same affinity for β-synemin or whether one of the isoforms preferentially functions with β-synemin in muscle. Methods: The two α-dystrobrevin isoforms (-1 and -2) and β-synemin were localized in regenerating rat tibialis anterior muscle using immunoprecipitation, immunohistochemical and immunoblot analyses. Immunoprecipitation and co-localization studies for α-dystrobrevin and β-synemin were performed in regenerating muscle following cardiotoxin injection. Protein expression was then compared to that of developing rat muscle using immunoblot analysis.Results: With an anti-α-dystrobrevin antibody, β-synemin co-immunoprecipitated with α-dystrobrevin whereas with an anti-β-synemin antibody, α-dystrobrevin-1 (rather than the -2 isoform) preferentially co-immunoprecipitated with β-synemin. Immunohistochemical experiments show that β-synemin and α-dystrobrevin co-localize in rat skeletal muscle. In regenerating muscle, β-synemin is first expressed at the sarcolemma and in the cytoplasm at day 5 following cardiotoxin injection. Similarly, β-synemin and α-dystrobrevin-1 are detected by immunoblot analysis as weak bands by day 7. In contrast, immunoblot analysis shows that α-dystrobrevin-2 is expressed as early as 1 day post-injection in regenerating muscle. These results are similar to that of developing muscle. For example, in embryonic rats, immunoblot analysis shows that β-synemin and α-dystrobevin-1 are weakly expressed in developing lower limb muscle at 5 days post-birth, while α-dystrobrevin-2 is detectable before birth in 20-day post-fertilization embryos. Conclusion: Our results clearly show that β-synemin expression correlates with that of α-dystrobrevin-1, suggesting that β-synemin preferentially functions with α-dystrobrevin-1 in vivo and that these proteins are likely to function coordinately to play a vital role in developing and regenerating muscle

Destabilization of the Dystrophin-Glycoprotein Complex without Functional Deficits in α-Dystrobrevin Null Muscle

α-Dystrobrevin is a component of the dystrophin-glycoprotein complex (DGC) and is thought to have both structural and signaling roles in skeletal muscle. Mice deficient for α-dystrobrevin (adbn−/−) exhibit extensive myofiber degeneration and neuromuscular junction abnormalities. However, the biochemical stability of the DGC and the functional performance of adbn−/− muscle have not been characterized. Here we show that the biochemical association between dystrophin and β-dystroglycan is compromised in adbn−/− skeletal muscle, suggesting that α-dystrobrevin plays a structural role in stabilizing the DGC. However, despite muscle cell death and DGC destabilization, costamere organization and physiological performance is normal in adbn−/− skeletal muscle. Our results demonstrate that myofiber degeneration alone does not cause functional deficits and suggests that more complex pathological factors contribute to the development of muscle weakness in muscular dystrophy

An Alpha-Catulin Homologue Controls Neuromuscular Function through Localization of the Dystrophin Complex and BK Channels in Caenorhabditis elegans

The large conductance, voltage- and calcium-dependent potassium (BK) channel serves as a major negative feedback regulator of calcium-mediated physiological processes and has been implicated in muscle dysfunction and neurological disorders. In addition to membrane depolarization, activation of the BK channel requires a rise in cytosolic calcium. Localization of the BK channel near calcium channels is therefore critical for its function. In a genetic screen designed to isolate novel regulators of the Caenorhabditis elegans BK channel, SLO-1, we identified ctn-1, which encodes an α-catulin homologue with homology to the cytoskeletal proteins α-catenin and vinculin. ctn-1 mutants resemble slo-1 loss-of-function mutants, as well as mutants with a compromised dystrophin complex. We determined that CTN-1 uses two distinct mechanisms to localize SLO-1 in muscles and neurons. In muscles, CTN-1 utilizes the dystrophin complex to localize SLO-1 channels near L-type calcium channels. In neurons, CTN-1 is involved in localizing SLO-1 to a specific domain independent of the dystrophin complex. Our results demonstrate that CTN-1 ensures the localization of SLO-1 within calcium nanodomains, thereby playing a crucial role in muscles and neurons