Originally published in ASBMB Today
Insights into treatment of Brody disease through inhibition of the ubiquitin-proteasome system
A recent study in the Journal of Biological Chemistry about a muscular disease in cattle may offer clues about how to treat a similar disease found in humans.
Both the Chianina cattle muscular disease pseudomyotonia and the human Brody disease are characterized by an inability of skeletal muscles to relax after strenuous physical exercise, leading to temporary muscle stiffness. The cause is a mutation in the ATP2A1 gene encoding a protein called SERCA1, which is crucial for pumping calcium from the cytosol back to the lumen of sarcoplasmic reticulum, thus enabling muscle relaxation. Because of such phenotypic and genetic overlap, Chianina pseudomyotonia is studied as a model for Brody disease.
Interestingly, the mutated SERCA1 protein retains its basic calcium-dependent ATPase activity like the normal protein, suggesting that the mutation does not affect its function. What is affected, however, is the amount of the mutant protein in skeletal muscles, which is much lower in comparison to normal protein levels. This is despite normal mRNA levels of the ATP2A1 gene. These key observations prompted Roberta Sacchetto and Dorianna Sandona at the University of Padova in Italy to join forces and investigate the potential roles of the ubiquitin-proteasome system in degrading the mutant proteins.
The team blocked the ubiquitin-proteasome system with different chemical inhibitors and measured the effect on the levels of mutant SERCA1 in a cellular model. Disrupting the pathway dramatically rescued the expression levels and membrane localization of SERCA1 as determined by Western blot analysis and immunofluorescence analyses.
To corroborate further the role of the ubiquitin-proteasome system in degradation of the mutant SERCA1, the team measured polyubiquitination of the mutant against that of the normal protein. Their rationale was that since the chemical inhibitor used to disrupt the ubiquitin-proteasome system pathway works downstream of the ubiquitination step, it would not affect the accumulation of polyubiquitaned forms of mutant SERCA1 protein. Results from immunoprecipitation assays confirmed the scientists’ hypothesis, and, indeed, there was an increase in the polyubiquitinated mutant SERCA1.
The researchers demonstrated that the pharmacologically rescued SERCA1 was able to restore cytoplasmic calcium homeostasis in a cellular model and was also fully active in muscle fibers isolated from a PMT-affected cow.
A Chianina cow and calf in a field in Tuscany. Images courtesty of Steven Walling, a wikimedia commons user
The significance of the study is that it demonstrates for the first time the role of the ubiquitin-proteasome system in degrading the mutant SERCA1 protein, explaining the symptoms associated with Chianina cattle pseudomyotonia and some forms of Brody disease. The findings also suggest specific inhibition of the ubiquitin-proteasome system could be a sound therapeutic strategy against the two diseases.