Effects of Cardiomyopathy-Related Troponin T Mutations on the Ubiquitin-Proteasome System

Abstract

Familial hypertrophic cardiomyopathy (FHC) is a disease of the myocardium that can be caused by a mutation in a sarcomeric gene. TNNT2 encodes cardiac troponin T (cTnT), a sarcomeric protein important for cardiac muscle contraction. Mutations in TNNT2 are frequently linked to sudden cardiac death (SCD); however their role in heart failure and cardiomyopathy is unclear. We hypothesize that changes in the ubiquitin-proteasome system (UPS) are related to increased risk of cardiac death, and that changes in the UPS may be related to calcium sensitization caused by TNNT2 mutations. We examined two mutations in TNNT2, R278C, which is associated with mild effects and late-onset heart disease, and the calcium-sensitizing mutation I79N, which is early-onset and has severe cardiac effects, including SCD. In 3-month-old transgenic mice expressing the I79N mutant form of troponin T, expression of the proteasome subunit PSMA6 was decreased by approximately 35% in I79N mice when compared to hearts from wild-type transgenic mice. Protein levels of the 19S regulator subunit Rpt1 were increased in the hearts of 3-month-old I79N mice and decreased in the hearts of 1-year-old R278C mice. Expression of PKC alpha, an important regulator of cardiac contractility, was decreased in R278C mice at 1 year. Activity of the lysosomal protease cathepsin L was increased in 1-year-old I79N mice, indicating disruption of another major proteolytic system in these hearts. Quantitative real-time PCR of the genes involved in the ubiquitination pathway (E1, E2, and E3 enzymes) showed that transcript levels were increased or decreased for 25 genes in I79N mice and three genes in R278C mice. These data demonstrate that the UPS is affected at the gene expression and protein level in R278C and I79N mice, indicating that changes in the UPS may be involved in FHC.