The Caveolin‐3 G56S sequence variant of unknown significance: Muscle biopsy findings and functional cell biological analysis

1. PURPOSE: In the era of next‐generation sequencing, we are increasingly confronted with sequence variants of unknown significance. This phenomenon is also known for variations in Caveolin‐3 and can complicate the molecular diagnosis of the disease. Here, we aimed to study the ambiguous character of the G56S Caveolin‐3 variant. 2. EXPERIMENTAL DESIGN: A comprehensive approach combining genetic and morphological studies of muscle derived from carriers of the G56S Caveolin‐3 variant were carried out and linked to biochemical assays (including phosphoblot studies and proteome profiling) and morphological investigations of cultured myoblasts. 3. RESULTS: Muscles showed moderate chronic myopathic changes in all carriers of the variant. Myogenic RCMH cells expressing the G56S Caveolin‐3 protein presented irregular Caveolin‐3 deposits within the Golgi in addition to a regular localization of the protein to the plasma membrane. This result was associated with abnormal findings on the ultra‐structural level. Phosphoblot studies revealed that G56S affects EGFR‐signaling. Proteomic profiling demonstrated alterations in levels of physiologically relevant proteins which are indicative for antagonization of G56S Caveolin‐3 expression. Remarkably, some proteomic alterations were enhanced by osmotic/mechanical stress. 4. CONCLUSIONS AND CLINICAL RELEVANCE: Our studies suggest that G56S might influence the manifestation of myopathic changes upon the presence of additional cellular stress burden. Results of our studies moreover improve the current understanding of (genetic) causes of myopathic disorders classified as caveolinopathies.