Biomechanical analysis of sarcomeric mutations that cause early-onset cardiomyopa

描述（由申请人提供）：肥厚性心肌病（HCM）是一种涉及心脏心室壁增厚的疾病。在婴儿中，其表现特别严重，是儿科人群心源性猝死的主要原因。不幸的是，目前的治疗仅限于轻度病例的症状缓解和心脏移植

DESCRIPTION (provided by applicant): Hypertrophic cardiomyopathy (HCM) is a disease involving the thickening of the ventricular walls of the heart. In infants, its presentation is particularly severe, and it is the leading cause of sudden cardiac death in pediatric populations. Unfortunately, current therapy is limited to symptomatic relief for mild cases and heart transplant for severe cases, and the functional changes caused by such mutations are not well understood. To develop targeted therapies, we must first better understand HCM's causes, which are believed to be genetic in nature for ~75% of pediatric HCM cases. It is believed that alteration of β-cardiac myosin's power generation by HCM mutations leads to a series of cellular changes that gradually cause the HCM disease phenotype. To date, few mutations have been rigorously investigated, and previous studies were performed using non-human myosins. These studies have produced conflicting results, underscoring the need for the expression of human β-cardiac myosin. A new murine myoblast-based expression technique has been developed, and the Spudich Lab has just published the first biochemical characterization study of wild type and mutant human β-cardiac myosin in an entirely human system. Using this new expression system will assess the effects of 5 pediatric-specific HCM mutations on the biomechanical function of human β-cardiac myosin using multiple assays: the F-actin activated ATPase assay to measure total cycle time, the in vitro motility assay to measure the unloaded maximum contractile velocity, and the dual beam optical trap assay to measure intrinsic force generation. I will also utilize a newly-developed oscillation technique to apply varying forces and measure velocity of contraction as a function of force. By performing the first biomechanical analysis of pediatric-specific HCM mutations and the first forcevelocity analysis of any cardiomyopathy-causing mutation, I will determine the extent to which these mutations alter myosin's ability to move and produce force. By highlighting the mechanisms by which mutations can alter molecular motor function, these pediatric-specific mutations may offer excellent models for testing potential small molecule therapeutics for treating HCM.