INHIBITION OF PROTEASOME ACTIVITY PROMOTES THE CORRECT LOCALIZATION OF DESEASE-CAUSING ALPHA-SARCOGLYCAN MUTANTS IN A HETEROLOGOUS CELL SYSTEM EXPRESSING BETA-, GAMMA-, AND DELTA-SARCOGLYCAN

Sarcoglycanopathies are progressive muscle wasting disorders caused by genetic defects of four proteins, alpha-, beta-, gamma-, and delta-sarcoglycan, elements of a key transmembrane complex of striated muscle. The proper assembly of the sarcoglycan complex represents a critical issue of sarcoglycanopathies, as several mutations severely perturb tetramer formation. Misfolded proteins are generally discarded through the cell’s quality-control system that, however, could lead to the removal of functional polypeptides. To explore whether it is possible to rescue sarcoglycan mutants by preventing their degradation, we generated a heterologous cell system constitutively expressing three (beta, gamma, and delta) of the four sarcoglycans. In these cells (bgd-HEK), the lack of alpha-sarcoglycan prevented complex formation and cell surface localization, while the presence of alpha-sarcoglycan allowed maturation and targeting of the tetramer. On the contrary, transfection of bgd-HEK cells with disease-causing alpha-sarcoglycan mutants led to a dramatic reduction of mutated proteins, compared to wild type, and the absence of the complex from cell surface. This result closely reproduces the observations made in muscle patients. Proteasomal inhibition reduced the degradation of mutants and facilitated the assembly and targeting of the sarcoglycan complex to the plasma membrane. However, as expected for a critical process necessary for cell survival, the prolonged inhibition of proteasome severely compromised cell viability. Importantly, when the treatment included the inhibition of caspase activity, the cytotoxic effect of proteasome inhibitor was almost completely abolished. Finally, in a trial application of this approach to a skeletal muscle explant isolated from an LGMD-2D patient, incubation with the FDA-approved proteasome inhibitor Velcade (bortezomib) rescued the expression of mutant alpha-sarcoglycan to the cell membrane. The present data provide important insights for the development of pharmacological therapies for sarcoglycanopathies. Funded by AFM (grant # 12988) and University of Padova.