Revealing the intricate mechanisms of excitation-contraction coupling in skeletal muscle weakness associated with heart failure

Revealing the intricate mechanisms of excitation-contraction coupling in skeletal muscle weakness associated with heart failure. Abstract Heart failure (HF) affects 2-3% of the population and is the leading cause of hospital admissions for individuals over 65, with symptomatic patients facing a 45% mortality rate within one year. HF is characterized by severe fatigue and reduced exercise capacity, limiting daily activities and significantly diminishing quality of life. However, the specific mechanisms causing muscle weakness and fatigue in HF remain largely unknown, and effective targeted treatments are lacking. HF triggers pathological calcium (Ca2+)-dependent signaling that disrupts muscle cell function. Excitation-contraction coupling (ECC)—the physiological process connecting neural excitation to muscle contraction—is notably impaired in HF, with ryanodine receptor type 1 (RyR1) Ca2+ channels playing a key role in these abnormalities. Despite substantial advances in understanding ECC in both healthy and diseased muscles, many details about the molecular processes driving muscle contraction remain unclear. Current research mainly addresses whole-muscle cell dysfunction rather than focusing on defective protein microenvironments or altered amino acids. This proposal seeks to identify oxidized cysteine residues within RyR1 channels that contribute to abnormal Ca2+ handling in HF-affected skeletal muscle, using mass spectrometry, the channel's high-resolution structure, and CRISPR mutagenesis. Furthermore, we will study the role of NADPH oxidases and explore changes in RyR1-associated proteins and interactions under both normal and pathological conditions. This will involve a mouse model engineered with plant peroxidase (TurboID) attached to the RyR1 cytosolic shell, allowing for the labeling of nearby proteins within a 10-20 nm range by adding biotin-phenol. By comparing protein abundances near the channel in HF and control mice using multiplexed quantitative mass spectrometry, we aim to map changes in the RyR1 microenvironment during HF. These studies will provide new insights into the specific mechanisms of skeletal muscle dysfunction, enhancing our understanding of ECC defects, particularly in the context of HF.