Friday, May 9, 2025
5/9/2025
The myosin super-relaxed state and human dilated cardiomyopathy - Project Summary/Abstract Dilated cardiomyopathies are a frequent cause of heart failure worldwide, characterized by reduced systolic function and left ventricular dilation. Dilated cardiomyopathy has been linked to various causes, including viral infections, drug abuse, endocrine dysfunction, and genetic mutations. Reduced effective myocardial contractility presents as a symptom early in disease progression of dilated cardiomyopathy. Studying the molecular basis of decreased contractility by investigating the myosin motors that power contractions can provide important insight and allow testing of potential therapeutic modulation. Current therapeutics for dilated cardiomyopathy (e.g., ACE inhibitors, β-blockers, etc.) target symptoms but do not attenuate the underlying source of contractile dysfunction. At the molecular level, force generation in cardiac muscle is determined by the number of myosin motors that bind to the actin thin-filament during the systolic rise of calcium. Once myosin motors bind, myosins hydrolyze ATP and undergo conformational steps to generate force. Recent attention has focused on the myosin biochemical super-relaxed state, an energy-conserving, auto-inhibited state of cardiac myosin that determines the proportion of myosin motors available. Thus, the amount of myosin motors in the super-relaxed state modulates the magnitude of myosin motors available to generate sarcomere power upon activation. The stability of the super-relaxed state can be modulated by phosphorylation of regulatory light chain and myosin binding protein C. Additionally, as force increases, stress on the thick filament rises resulting in the release of myosin heads, allowing greater force production. A decrease in available myosin motors as a result of a shift towards the super-relaxed state is a potential mechanism of systolic dysfunction in human dilated cardiomyopathy and has yet to be fully investigated. Therefore, our central hypothesis is that depressed contractile force in dilated cardiomyopathy results from increased myosins in the super-relaxed state (Aim 1) mediated by depressed regulatory protein phosphorylation (Aim 2), which may be able to be reversed with the use of myosin activators (Aim 3). To test our hypothesis, we will utilize the fluorescent ATP pulse-chase technique to quantify the proportion of myosins in the super-relaxed state and test the effects of myosin activators at short and long sarcomere lengths. Also, using Phos-TagTM western blotting, we will measure the phosphorylation status of regulatory proteins. This proposal will provide novel insight into the effects of myosin activators on the super-relaxed state, demonstrate fundamental aspects of myosin biochemical states, and their role in the molecular basis of human dilated cardiomyopathy.
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