Résumé :
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Communication n° 280 Cardiac myosin binding protein C (cMyBP-C) gene mutations are a frequent cause of familial hypertrophic cardiomyopathy (FHC), and most of them result in C-terminal truncated cMyBP-Cs. However, truncated cMyBP-Cs were undetectable in myocardial tissue of FHC patients. In the present study, we investigated whether truncated cMyBP-Cs are subject to accelerated degradation by the lysosome or the ubiquitin-proteasome system (UPS). Neonatal rat cardiomyocytes were infected with recombinant adenovirus encoding wild type (wt) or truncated cMyBP-Cs (M6t 3% truncation, M7t 80% truncation, both mutations have been identified in FHC patients). Despite similar mRNA levels, protein expression of M6t and M7t mutants were markedly reduced to 70Æ4% and 11Æ5% of wt, respectively (n=10, p<0.05). Treatment with the lysosomal inhibitors bafilomycin A1 (50 nM, 6 h) or wortmannin (500 nM, 6 h) slightly raised the protein level of the M7t mutant, but did not affect protein levels of wt. Treatment with the UPS inhibitors MG132 (50 µM, 2.5 h) or lactacystin (25 µM, 24 h) was more effective and raised protein levels of M6t and M7t to wt level. Immunostaining showed correct incorporation of wt into the A-band, low A-band incorporation of M6t, and mis-incorporation of M7t into the Z-disc. In addition, M7t, but not wt or M6t, showed intracellular ubiquitin-positive aggregates. Using an adenovirus encoding a fluorescent substrate of UPS (UbG76V-DsRed), we demonstrate that M7t mutant cMyBP-C impair the proteolytic capacity of the UPS. In conclusion, these data provide evidence that truncated cMyBP-C are rapidly and quantitatively degraded by the UPS, most likely explaining why these mutants are not detected in patients tissue before. Importantly, the rapid degradation of the truncated mutant was associated with the formation of ubiquitin-positive aggregates and inhibition of the proteolytic capacity of the UPS. Since UPS plays an important role in a variety of fundamental cellular process, we propose impairment of this system by mutant cMyBP-C as a contributing factor to the pathogenesis of FHC.
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