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Myosin II Adjusts Motility Properties and Regulates Force Production Based on Motor Environment

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Abstract

Introduction

Myosin II has been investigated with optical trapping, but single motor-filament assay arrangements are not reflective of the complex cellular environment. To understand how myosin interactions propagate up in scale to accomplish system force generation, we devised a novel actomyosin ensemble optical trapping assay that reflects the hierarchy and compliancy of a physiological environment and is modular for interrogating force effectors.

Methods

Hierarchical actomyosin bundles were formed in vitro. Fluorescent template and cargo actin filaments (AF) were assembled in a flow cell and bundled by myosin. Beads were added in the presence of ATP to bind the cargo AF and activate myosin force generation to be measured by optical tweezers.

Results

Three force profiles resulted across a range of myosin concentrations: high force with a ramp-plateau, moderate force with sawtooth movement, and baseline. The three force profiles, as well as high force output, were recovered even at low solution concentration, suggesting that myosins self-optimize within AFs. Individual myosin steps were detected in the ensemble traces, indicating motors are taking one step at a time while others remain engaged in order to sustain productive force generation.

Conclusions

Motor communication and system compliancy are significant contributors to force output. Environmental conditions, motors taking individual steps to sustain force, the ability to backslip, and non-linear concentration dependence of force indicate that the actomyosin system contains a force-feedback mechanism that senses the local cytoskeletal environment and communicates to the individual motors whether to be in a high or low duty ratio mode.

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Abbreviations

AF:

Actin filament

OT:

Optical tweezers, optical trapping

SM:

Single molecule

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Acknowledgments

This work is supported in part by the University of Mississippi Graduate Student Council Research Fellowship (OA), University of Mississippi Sally McDonnell-Barksdale Honors College (JCW, JER), the Mississippi Space Grant Consortium under Grant Number NNX15AH78H (JCW, DNR), and the American Heart Association under Grant Number 848586 (DNR).

Author Contributions

OA was involved in all aspects of the work, including assay development, performing experiments, data analysis, and manuscript preparation. OA, JCW, JED, JER, and DNR aided in assay development, data acquisition, and analysis. OA and DNR designed the experiments, analyzed data, and prepared the manuscript.

Conflict of interest

Omayma M. Al Azzam, Janie C. Watts, Justin E. Reynolds, Juliana E. Davis, and Dana N. Reinemann declare that they have no conflict of interest.

Research Involving Human Rights

No human studies were carried out by the authors for this article.

Research Involving Animal Rights

No animal studies were carried out by the authors for this article.

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Authors and Affiliations

  1. Department of Chemical Engineering, University of Mississippi, University, MS, 38677, USA

    Omayma Y. Al Azzam, Janie C. Watts & Dana N. Reinemann

  2. Department of Biomedical Engineering, University of Mississippi, University, MS, 38677, USA

    Justin E. Reynolds, Juliana E. Davis & Dana N. Reinemann

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  1. Omayma Y. Al Azzam
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Al Azzam, O.Y., Watts, J.C., Reynolds, J.E. et al. Myosin II Adjusts Motility Properties and Regulates Force Production Based on Motor Environment. Cel. Mol. Bioeng. 15, 451–465 (2022). https://doi.org/10.1007/s12195-022-00731-1

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