http://repub.eur.nl/res/aut/34431/ List of Publications en http://repub.eur.nl/static-eur/img/logo.png http://repub.eur.nl http://repub.eur.nl/res/pub/25286/ 2009-03-24T00:00:00Z Mutations in the MYBPC3 gene, encoding cardiac myosin-binding protein C (cMyBP-C), are a frequent cause of familial hypertrophic cardiomyopathy. In the present study, we investigated whether protein composition and function of the sarcomere are altered in a homogeneous familial hypertrophic cardiomyopathy patient group with frameshift mutations in MYBPC3 (MYBPC3mut).Methods and Results- Comparisons were made between cardiac samples from MYBPC3 mutant carriers (c.2373dupG, n=7; c.2864-2865delCT, n=4) and nonfailing donors (n= 13). Western blots with the use of antibodies directed against cMyBP-C did not reveal truncated cMyBP-C in MYBPC3mut. Protein expression of cMyBP-C was significantly reduced in MYBPC3mutby 33 ±5%. Cardiac MyBP-C phosphorylation in MYBPC3mutsamples was similar to the values in donor samples, whereas the phosphorylation status of cardiac troponin I was reduced by 84 ±5%, indicating divergent phosphorylation of the 2 main contractile target proteins of the β-adrenergic pathway. Force measurements in mechanically isolated Triton-permeabilized cardiomyocytes demonstrated a decrease in maximal force per cross- sectional area of the myocytes in MYBPC3mut(20.2±2.7 kN/m2) compared with donor (34.5± 1.1 kN/m2). Moreover, Ca2+sensitivity was higher in MYBPC3mut(pCa50=5.62±0.04) than in donor (pCa50=5.54±0.02), consistent with reduced cardiac troponin I phosphorylation. Treatment with exogenous protein kinase A, to mimic β-adrenergic stimulation, did not correct reduced maximal force but abolished the initial difference in Ca sensitivity between MYBPC3mut(pCa50=5.46±0.03) and donor (pCa50=5.48±0. 02).Conclusions- Frameshift MYBPC3 mutations cause haploinsufficiency, deranged phosphorylation of contractile proteins, and reduced maximal force-generating capacity of cardiomyocytes. The enhanced Ca2+sensitivity in MYBPC3mutis due to hypophosphorylation of troponin I secondary to mutation-induced dysfunction. http://repub.eur.nl/res/pub/29146/ 2008-03-01T00:00:00Z Sarcomeric dysfunction plays a central role in reduced cardiac pump function in heart failure. This review focuses on the alterations in sarcomeric proteins in diseased myocardium that range from altered isoform expression to post-translational protein changes such as proteolysis and phosphorylation. Recent studies in animal models of heart failure and human failing myocardium converge and indicate that sarcomeric dysfunction, including altered maximum force development, Ca2+sensitivity, and increased passive stiffness, largely originates from altered protein phosphorylation, caused by neurohumoral-induced alterations in the kinase-phosphatase balance inside the cardiomyocytes. Novel therapies, which specifically target phosphorylation sites within sarcomeric proteins or the kinases and phosphatases involved, might improve cardiac function in heart failure.