A common MLP (muscle LIM protein) variant is associated with cardiomyopathy.

RATIONALE: We previously discovered the human 10T-->C (Trp4Arg) missense mutation in exon 2 of the muscle LIM protein (MLP, CSRP3) gene. OBJECTIVE: We sought to study the effects of this single-nucleotide polymorphism in the in vivo situation. METHODS AND RESULTS: We now report the generation and detailed analysis of the corresponding Mlp(W4R/+) and Mlp(W4R/W4R) knock-in animals, which develop an age- and gene dosage-dependent hypertrophic cardiomyopathy and heart failure phenotype, characterized by almost complete loss of contractile reserve under catecholamine induced stress. In addition, evidence for skeletal muscle pathology, which might have implications for human mutation carriers, was observed. Importantly, we found significantly reduced MLP mRNA and MLP protein expression levels in hearts of heterozygous and homozygous W4R-MLP knock-in animals. We also detected a weaker in vitro interaction of telethonin with W4R-MLP than with wild-type MLP. These alterations may contribute to an increased nuclear localization of W4R-MLP, which was observed by immunohistochemistry. CONCLUSIONS: Given the well-known high frequency of this mutation in Caucasians of up to 1%, our data suggest that (W4R-MLP) might contribute significantly to human cardiovascular disease.

Insulin signaling regulates cardiac titin properties in heart development and diabetic cardiomyopathy.

Isoform-switching of the giant elastic protein titin is a main mechanism for adjusting passive myocardial stiffness in perinatal heart development and chronic heart disease. Previous evidence suggested that thyroid-hormone signaling converging onto the phosphoinositol-3-kinase (PI3K)/AKT pathway regulates titin-isoform composition in developing cardiomyocytes. Here we hypothesized that insulin, another activator of PI3K/AKT, alters titin-isoform composition and titin-based stiffness. We also checked for insulin-induced changes in titin phosphorylation. In embryonic rat cardiomyocytes cultured in the presence of insulin for 7 days, the mean proportion of the stiff N2B-titin isoform (M(w), 3000 kDa) significantly increased from 53% in controls to 65% in insulin-treated cells, the remainder being the more compliant N2BA-isoforms (M(w), 3200-3700 kDa). This insulin-dependent titin-isoform shift was blocked by PI3K-inhibitor, LY294002, suggesting that insulin controls the cardiac titin-isoform pattern by activating PI3K/AKT. Titin phosphorylation was substantially increased in insulin-treated compared to control cardiomyocytes. The impact of insulin-deficiency in vivo on titin-isoform expression, titin phosphorylation, and passive myocardial stiffness was studied in streptozotocin-treated (STZ) rats as a model of diabetes mellitus type-1. Within 5 months, STZ rats developed cardiac hypertrophy and mild left ventricular fibrosis, concomitant with elevated glucose levels. The mean proportion of N2B-titin was slightly but significantly decreased from 86% in controls to 79% in STZ hearts. Titin phosphorylation levels remained unchanged. Mechanical measurements on skinned cardiac fibers showed only minor passive stiffness modifications in STZ myocardium. We conclude that insulin signaling regulates titin-isoform composition and titin phosphorylation in embryonic cardiomyocytes and could also contribute to altered diastolic function in diabetic cardiomyopathy.