Caveolin-3 T78M and T78K missense mutations lead to different phenotypes in vivo and in vitro

Abstract

Caveolins are the principal protein components of caveolae, invaginations of the plasma membrane involved in cell signaling and trafficking. Caveolin-3 (Cav-3) is the muscle-specific isoform of the caveolin family and mutations in the CAV3 gene lead to a large group of neuromuscular disorders. In unrelated patients, we identified two distinct CAV3 mutations involving the same codon 78. Patient 1, affected by dilated cardiomyopathy and limb girdle muscular dystrophy (LGMD)-1C, shows an autosomal recessive mutation converting threonine to methionine (T78M). Patient 2, affected by isolated familiar hyperCKemia, shows an autosomal dominant mutation converting threonine to lysine (T78K). Cav-3 wild type (WT) and Cav-3 mutations were transiently transfected into Cos-7 cells. Cav-3 WT and Cav-3 T78M mutant localized at the plasma membrane, whereas Cav-3 T78K was retained in a perinuclear compartment. Cav-3 T78K expression was decreased by 87% when compared with Cav-3 WT, whereas Cav-3 T78M protein levels were unchanged. To evaluate whether Cav-3 T78K and Cav-3 T78M mutants behaved with a dominant negative pattern, Cos-7 cells were cotransfected with green fluorescent protein (GFP)-Cav-3 WT in combination with either mutant or WT Cav-3. When cotransfected with Cav-3 WT or Cav-3 T78M, GFP-Cav-3 WT was localized at the plasma membrane, as expected. However, when cotransfected with Cav-3 T78K, GFP-Cav-3 WT was retained in a perinuclear compartment, and its protein levels were reduced by 60%, suggesting a dominant negative action. Accordingly, Cav-3 protein levels in muscles from a biopsy of patient 2 (T78K mutation) were reduced by 80%. In conclusion, CAV3 T78M and T78K mutations lead to distinct disorders showing different clinical features and inheritance, and displaying distinct phenotypes in vitro.

Caveolinopathies: from the biology of caveolin-3 to human diseases

“…78 Only recently, Traverso et al reported a 58-year old patient affected by LGMD-1C and dilatative cardiomyopathy (p.T77M mutation). 47 It is possible that compensatory mechanisms to Cav-3 deficiency are different in cardiac and skeletal muscle tissues, but the molecular basis for this difference is not known.…”

“…In the patient Cav-3 protein levels are reduced by approximately 80%. Modified and reproduced by Traverso et al 47 Caveolin-3 and muscle diseasesH-CK and proven CAV3 mutations (p.G55S and p.T77M) with normal Cav-3 expression at the muscle biopsy. It is feasible that these mutants result into a stable but dysfunctional protein.…”

“…Finally, the same group identified one patient affected by dilated cardiomyopathy and LGMD-1C and carrying a Cav-3 p.T77M substitution. 47 In a screening of 663 patients affected by primary myopathies of unknown etiology, CAV3 mutations represented the one percent of unclassified LGMD and other phenotypes including isolated H-CK, RMD and proximal and distal myopathy. 48 In addition, four CAV3 mutants have been identified in patients affected by LQTS and SIDS, confirming a role for Cav-3 in the regulation of cardiac ion channels.…”

Delineating the role of alterations in lipid metabolism to the pathogenesis of inherited skeletal and cardiac muscle disorders

“…The missense T63S mutation in the CAV3 gene resulted in decreased cell surface expression of caveolin-3 ( 228 ). Recently, another missense T78M mutation in the CAV3 gene has been identifi ed in a patient suffering from both DCM and LGMD-1C ( 229 ). Interestingly, these patients do not display skeletal muscle anomalies that are common in LGMD with underlying caveolin-3 mutations ( 228,230 ).…”

“…Skeletal muscle contracts on demand, whereas the continuous contraction and relaxation of cardiac muscle requires more caveolin-3 to deliver a larger amount of fatty acid to mitochondria for sustained energy production ( 228 ). Moreover, caveolin-3 may be regulated by different mechanisms in skeletal and cardiac muscles ( 229 ). Because hypertrophy occurs as a secondary response to other conditions, such as valvular disease, hypertension, and ischemic heart disease ( 231, 232 ), delineation of inherited FMC and DCM changes from other noninherited conditions has become quite a challenge.…”