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Direct Measurements of Local Coupling between Myosin Molecules Are Consistent with a Model of Muscle Activation.

Walcott S, Kad NM - PLoS Comput. Biol. (2015)

Bottom Line: This result contrasts with a theoretical model, which reproduces several indirect measurements of coupling between myosin, that assumes a single myosin head can activate the thin filament.The conclusion that two myosin molecules are required to activate the thin filament arises from an assumption, made during data analysis, that the intensity of the fluorescent tags attached to myosin varies depending on experimental condition.We provide an alternative explanation that reconciles theory and experiment without assuming that the intensity of the fluorescent tags varies.

View Article: PubMed Central - PubMed

Affiliation: Mathematics, University of California at Davis, Davis, California, United States of America.

ABSTRACT
Muscle contracts due to ATP-dependent interactions of myosin motors with thin filaments composed of the proteins actin, troponin, and tropomyosin. Contraction is initiated when calcium binds to troponin, which changes conformation and displaces tropomyosin, a filamentous protein that wraps around the actin filament, thereby exposing myosin binding sites on actin. Myosin motors interact with each other indirectly via tropomyosin, since myosin binding to actin locally displaces tropomyosin and thereby facilitates binding of nearby myosin. Defining and modeling this local coupling between myosin motors is an open problem in muscle modeling and, more broadly, a requirement to understanding the connection between muscle contraction at the molecular and macro scale. It is challenging to directly observe this coupling, and such measurements have only recently been made. Analysis of these data suggests that two myosin heads are required to activate the thin filament. This result contrasts with a theoretical model, which reproduces several indirect measurements of coupling between myosin, that assumes a single myosin head can activate the thin filament. To understand this apparent discrepancy, we incorporated the model into stochastic simulations of the experiments, which generated simulated data that were then analyzed identically to the experimental measurements. By varying a single parameter, good agreement between simulation and experiment was established. The conclusion that two myosin molecules are required to activate the thin filament arises from an assumption, made during data analysis, that the intensity of the fluorescent tags attached to myosin varies depending on experimental condition. We provide an alternative explanation that reconciles theory and experiment without assuming that the intensity of the fluorescent tags varies.

No MeSH data available.


Related in: MedlinePlus

Histograms, collected under three different experimental conditions [52], collapse upon rescaling fluorescence.Scaled intensity, defined in the text, gives a single fluorescently-tagged myosin (GFP-S1) an intensity of 1. The observation that each histogram has a peak near 1 suggests that 1) under these three conditions, mostly single GFP-S1s bind; and 2) the emission of an excited GFP is constant and measured fluorescence is inversely proportional to frame rate. This latter is a central assumption of our analysis.
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pcbi.1004599.g006: Histograms, collected under three different experimental conditions [52], collapse upon rescaling fluorescence.Scaled intensity, defined in the text, gives a single fluorescently-tagged myosin (GFP-S1) an intensity of 1. The observation that each histogram has a peak near 1 suggests that 1) under these three conditions, mostly single GFP-S1s bind; and 2) the emission of an excited GFP is constant and measured fluorescence is inversely proportional to frame rate. This latter is a central assumption of our analysis.

Mentions: To support the assumption of constant GFP-S1 fluorescence intensity, at e = 450 fluorescent units/s, we reanalyzed the data from Desai et al [52]. If GFP-S1 fluorescence varied significantly, then the fluorescent spots from single GFP-S1s between conditions could not be rationally rescaled. Using we rescaled the histograms of fluorescent intensity observed by Desai et al. [52] under three conditions where isolated single GFP-S1s are expected to dominate binding. These conditions are: high ATP (pCa 6, ATP = 1μM, [Myo] = 15nM, f = 3.8Hz), low myosin (pCa 6, ATP = 0.1μM, [Myo] = 1nM, f = 10Hz), and low calcium (pCa 7, ATP = 0.5μM, [Myo] = 15nM, f = 3.8Hz). The resulting histograms are similar, supporting the assumption of equal intensity (Fig 6).


Direct Measurements of Local Coupling between Myosin Molecules Are Consistent with a Model of Muscle Activation.

Walcott S, Kad NM - PLoS Comput. Biol. (2015)

Histograms, collected under three different experimental conditions [52], collapse upon rescaling fluorescence.Scaled intensity, defined in the text, gives a single fluorescently-tagged myosin (GFP-S1) an intensity of 1. The observation that each histogram has a peak near 1 suggests that 1) under these three conditions, mostly single GFP-S1s bind; and 2) the emission of an excited GFP is constant and measured fluorescence is inversely proportional to frame rate. This latter is a central assumption of our analysis.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4633106&req=5

pcbi.1004599.g006: Histograms, collected under three different experimental conditions [52], collapse upon rescaling fluorescence.Scaled intensity, defined in the text, gives a single fluorescently-tagged myosin (GFP-S1) an intensity of 1. The observation that each histogram has a peak near 1 suggests that 1) under these three conditions, mostly single GFP-S1s bind; and 2) the emission of an excited GFP is constant and measured fluorescence is inversely proportional to frame rate. This latter is a central assumption of our analysis.
Mentions: To support the assumption of constant GFP-S1 fluorescence intensity, at e = 450 fluorescent units/s, we reanalyzed the data from Desai et al [52]. If GFP-S1 fluorescence varied significantly, then the fluorescent spots from single GFP-S1s between conditions could not be rationally rescaled. Using we rescaled the histograms of fluorescent intensity observed by Desai et al. [52] under three conditions where isolated single GFP-S1s are expected to dominate binding. These conditions are: high ATP (pCa 6, ATP = 1μM, [Myo] = 15nM, f = 3.8Hz), low myosin (pCa 6, ATP = 0.1μM, [Myo] = 1nM, f = 10Hz), and low calcium (pCa 7, ATP = 0.5μM, [Myo] = 15nM, f = 3.8Hz). The resulting histograms are similar, supporting the assumption of equal intensity (Fig 6).

Bottom Line: This result contrasts with a theoretical model, which reproduces several indirect measurements of coupling between myosin, that assumes a single myosin head can activate the thin filament.The conclusion that two myosin molecules are required to activate the thin filament arises from an assumption, made during data analysis, that the intensity of the fluorescent tags attached to myosin varies depending on experimental condition.We provide an alternative explanation that reconciles theory and experiment without assuming that the intensity of the fluorescent tags varies.

View Article: PubMed Central - PubMed

Affiliation: Mathematics, University of California at Davis, Davis, California, United States of America.

ABSTRACT
Muscle contracts due to ATP-dependent interactions of myosin motors with thin filaments composed of the proteins actin, troponin, and tropomyosin. Contraction is initiated when calcium binds to troponin, which changes conformation and displaces tropomyosin, a filamentous protein that wraps around the actin filament, thereby exposing myosin binding sites on actin. Myosin motors interact with each other indirectly via tropomyosin, since myosin binding to actin locally displaces tropomyosin and thereby facilitates binding of nearby myosin. Defining and modeling this local coupling between myosin motors is an open problem in muscle modeling and, more broadly, a requirement to understanding the connection between muscle contraction at the molecular and macro scale. It is challenging to directly observe this coupling, and such measurements have only recently been made. Analysis of these data suggests that two myosin heads are required to activate the thin filament. This result contrasts with a theoretical model, which reproduces several indirect measurements of coupling between myosin, that assumes a single myosin head can activate the thin filament. To understand this apparent discrepancy, we incorporated the model into stochastic simulations of the experiments, which generated simulated data that were then analyzed identically to the experimental measurements. By varying a single parameter, good agreement between simulation and experiment was established. The conclusion that two myosin molecules are required to activate the thin filament arises from an assumption, made during data analysis, that the intensity of the fluorescent tags attached to myosin varies depending on experimental condition. We provide an alternative explanation that reconciles theory and experiment without assuming that the intensity of the fluorescent tags varies.

No MeSH data available.


Related in: MedlinePlus