Research Article: Auxotonic to isometric contraction transitioning in a beating heart causes myosin step-size to down shift

Date Published: April 19, 2017

Publisher: Public Library of Science

Author(s): Thomas P. Burghardt, Xiaojing Sun, Yihua Wang, Katalin Ajtai, Miklos S. Kellermayer.

http://doi.org/10.1371/journal.pone.0174690

Abstract

Myosin motors in cardiac ventriculum convert ATP free energy to the work of moving blood volume under pressure. The actin bound motor cyclically rotates its lever-arm/light-chain complex linking motor generated torque to the myosin filament backbone and translating actin against resisting force. Previous research showed that the unloaded in vitro motor is described with high precision by single molecule mechanical characteristics including unitary step-sizes of approximately 3, 5, and 8 nm and their relative step-frequencies of approximately 13, 50, and 37%. The 3 and 8 nm unitary step-sizes are dependent on myosin essential light chain (ELC) N-terminus actin binding. Step-size and step-frequency quantitation specifies in vitro motor function including duty-ratio, power, and strain sensitivity metrics. In vivo, motors integrated into the muscle sarcomere form the more complex and hierarchically functioning muscle machine. The goal of the research reported here is to measure single myosin step-size and step-frequency in vivo to assess how tissue integration impacts motor function.

Partial Text

The myosin motor protein powers the beating heart with transduction of ATP to mechanical work. Rationalizing “bottom-up” single myosin mechanics with “top-down” whole animal muscle physiology is indispensable to solving myosin’s structure/function paradigm for creating an ensemble capable nanomotor and to providing the insight into muscle disease mechanisms demanded by translational science. A time-resolved in vivo imaging approach characterizes single myosin mechanics in contracting striated muscle of live zebrafish embryos. It provides the means for linking bottom-up myosin characteristics to top-down muscle physiology or phenotype in the zebrafish embryo model for human muscle.

Myosin in striated muscle transduces ATP free energy into the mechanical work of moving actin. It does so using cyclical rotary movement of the lever-arm/light-chain complex linking motor generated torque to the myosin filament backbone. The linear actin displacement due to lever-arm rotation defines the unitary myosin step-size. The essential in vitro myosin structure/function paradigm is captured by its single molecule mechanical characteristics measured using the Qdot assay characterizing motor step-size and step-frequency [32]. Muscle myosin performance beyond the essential structure/function paradigm is influenced by self-assembly and integration with other motors and proteins in the muscle sarcomere. The native integrated myosin, with potential for hierarchical coordinated functionality and regulation because of its structured environment, was investigated here with cardiac myosin in live zebrafish. We developed tools to measure and interpret in vivo single cardiac myosin lever-arm rotation in a beating heart and estimate the cardiac myosin step-size and step-frequency. These metrics provide unprecedented insight into native cardiac myosin structure/function.

Cardiac ventriculum myosin transitioning from low to high force causes motor down-shifting to a 3 nm step-size accounting for >80% of all steps in the near-isometric contraction phase. We propose that strain delays ATP dissociation of actomyosin at isometric force using a mechanism involving the ratcheting effect of the actin bound ELC N-terminus. The significance of ELC N-terminus actin binding follows directly from earlier in vitro single myosin measurements [5, 17] showing close correlation between in vitro and in vivo systems. Enhanced occupation of the 0 length step hints at a mechanism for restrictive cardiomyopathy (RCM) caused by mutation in ELC [45]. Strongly bound myosins fail to detach quickly due to a fault in the ELC N-terminus strain sensing inhibiting detachment under low load conditions. It produces a drag force during stretch when the muscle should be fully relaxed.

 

Source:

http://doi.org/10.1371/journal.pone.0174690

 

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