Smooth muscle and skeletal muscle myosins produce similar unitary forces and displacements in the laser trap

Guilford, William H.; Dupuis, D. E.; Kennedy, Guy G.; Wu, Jinjin; Patlak, Joseph B.; Warshaw, David M.

doi:10.1016/s0006-3495(97)78753-8

Cited by 247 publications

(278 citation statements)

References 37 publications

Supporting

Mentioning

253

Contrasting

Order By: Relevance

“…Although the F764L mutant exhibited a 20% reduction in V actin (Table 3), the single-molecule measurements could not detect any changes in either d or t on . If, as for the S532P mutant myosin, both mechanical and kinetic components contribute equally to the 20% reduction in V actin , then 10% changes in both d and t on are below our detection limits (18). Regardless of the molecular mechanism, both the S532P and the F764L mutations share a common phenotype by having reduced actin filament velocities and actomyosin ATPase activities ( Fig.…”

Section: Resultsmentioning

confidence: 83%

See 1 more Smart Citation

Cardiac myosin missense mutations cause dilated cardiomyopathy in mouse models and depress molecular motor function

Schmitt

Debold²,

Ahmad³

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

108

View full text Add to dashboard Cite

Dilated cardiomyopathy (DCM) leads to heart failure, a leading cause of death in industrialized nations. Approximately 30% of DCM cases are genetic in origin, with some resulting from point mutations in cardiac myosin, the molecular motor of the heart. The effects of these mutations on myosin's molecular mechanics have not been determined. We have engineered two murine models characterizing the physiological, cellular, and molecular effects of DCM-causing missense mutations (S532P and F764L) in the ␣-cardiac myosin heavy chain and compared them with WT mice. Mutant mice developed morphological and functional characteristics of DCM consistent with the human phenotypes. Contractile function of isolated myocytes was depressed and preceded left ventricular dilation and reduced fractional shortening. In an in vitro motility assay, both mutant cardiac myosins exhibited a reduced ability to translocate actin (V actin) but had similar force-generating capacities. Actin-activated ATPase activities were also reduced. Single-molecule laser trap experiments revealed that the lower V actin in the S532P mutant was due to a reduced ability of the motor to generate a step displacement and an alteration of the kinetics of its chemomechanical cycle. These results suggest that the depressed molecular function in cardiac myosin may initiate the events that cause the heart to remodel and become pathologically dilated.animal models ͉ biophysics ͉ genetics ͉ single molecule ͉ laser trap D ilated cardiomyopathy (DCM) is an early step in the pathway leading to heart failure, the leading cause of human morbidity and mortality (1, 2). Although etiological factors such as ischemia and diabetes can result in DCM, at least 30% of cases are genetic in origin, with mutations found in sarcomeric proteins associated with the cytoskeletal and contractile systems (3). Clinically, DCM is characterized by myocardial hypertrophy, thin-walled ventricles, and myocyte hypoplasia (2, 4). Dilated hearts are hypocontractile, with systolic dysfunction leading to a reduced ejection fraction: a hallmark of a failing heart (2, 3). Only recently have autosomal dominant missense mutations to the ␤-cardiac myosin heavy chain (MHC) been identified that result in DCM (5, 6). Because the primary defect resides within the heart's molecular motor, we hypothesized that altered cardiac contractile function in DCM hearts might reflect changes in the mutant myosin's ability to generate force and motion as it cyclically interacts with actin during its hydrolysis of ATP.To investigate the impact of two distinct DCM-causing MHC mutations (S532P and F764L) on cardiac myosin's molecular performance, we have genetically engineered these missense mutations separately into the murine ␣-cardiac MHC gene, the human ␤-MHC homolog, to generate two separate DCM knockin mouse models. In addition to cardiac dilation and dysfunction, defects in isolated myocyte contractility were observed that are consistent with DCM. Interestingly, defects at the whole-heart and cellular levels were cor...

show abstract

Section: Resultsmentioning

confidence: 83%

“…Nineteen to 39 independent traces (Ϸ2 min in duration) were obtained for each myosin type. Estimates of the step size and duration of strong binding were derived by using the mean-variance technique (18) and are reported in Table 3.…”

Section: (2) Nsmentioning

confidence: 99%

Cardiac myosin missense mutations cause dilated cardiomyopathy in mouse models and depress molecular motor function

Schmitt

Debold²,

Ahmad³

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

108

View full text Add to dashboard Cite

show abstract

“…Strain energy is a particularly useful measure of the contractile strength. For instance, a single actin-myosin interaction can generate a maximum energy of about 10 Ϫ8 pJ (9,21). The strain energy we measured in a single cell stimulated with 10 M histamine is of the order of 1 pJ.…”

Section: Traction Maps Of Single Smooth Muscle Cellsmentioning

confidence: 80%

Spatial and temporal traction response in human airway smooth muscle cells

Tolic-Norrelykke

Butler

Chen

et al. 2002

American Journal of Physiology-Cell Physiology

126

104

View full text Add to dashboard Cite

. Spatial and temporal traction response in human airway smooth muscle cells. Am J Physiol Cell Physiol 283: C1254-C1266, 2002. First published June 26, 2002 10.1152/ajpcell.00169.2002Tractions that cells exert on their substrates are essential in cell spreading, migration, and contraction. These tractions can be determined by plating the cells on a flexible gel and measuring the deformation of the gel by using fluorescent beads embedded just below the surface of the gel. In this article we describe the image correlation method (ICM) optimized for determining the displacement field of the gel under a contracting cell. For the calculation of the traction field from the displacement field we use the recently developed method of Fourier transform traction cytometry (FTTC). The ICM and FTTC methods are applied to human airway smooth muscle cells during stimulation with the contractile agonist histamine or the relaxing agonist isoproterenol. The overall intensity of the cell contraction (the median traction magnitude, the energy transferred from the cell to the gel, and the net contractile moment) increased after activation with histamine, and decreased after treatment with isoproterenol. Cells exhibited regional differences in the time course of traction during the treatment. Both temporal evolution and magnitude of traction increase induced by histamine varied markedly among different cell protrusions, whereas the nuclear region showed the smallest response. These results suggest that intracellular mediators of cell adhesion and contraction respond to contractile stimuli with different rates and intensities in different regions of the cell.

show abstract

“…It should be noted that neither the specific cMyBP-C binding site on actin nor the physiologic relevance of such binding in muscle fibers (if present) has been defined. Thus, future studies characterizing the effect of cMyBP-C on the inherent mechanics and kinetics of the individual cross-bridge in the laser trap [54] using intact native thick filament [51] should offer insight into the molecular mechanisms by which cMyBP-C exerts its effect on contractility and its regulation by cMyBP-C phosphorylation. A) ProQ Diamond staining of untreated cMyBP-C (control) and cMyBP-C treated with PKA or PP2A.…”

Section: Discussionmentioning

confidence: 99%

Cardiac myosin binding protein-C modulates actomyosin binding and kinetics in the in vitro motility assay

Saber

Begin

Warshaw

et al. 2008

Journal of Molecular and Cellular Cardiology

View full text Add to dashboard Cite

The modulatory role of whole cardiac myosin binding protein-C (cMyBP-C) on myosin force and motion generation was assessed in an in vitro motility assay. The presence of cMyBP-C at an approximate molar ratio of cMyBP-C to whole myosin of 1:2, resulted in a 25% reduction in thin filament velocity (P<0.002) with no effect on relative isometric force under maximally activated conditions (pCa 5). Cardiac MyBP-C was capable of inhibiting actin filament velocity in a concentration-dependent manner using either whole myosin, HMM or S1, indicating that the cMyBP-C does not have to bind to myosin LMM or S2 subdomains to exert its effect. The reduction in velocity by cMyBP-C was independent of changes in ionic strength or excess inorganic phosphate. Cosedimentation experiments demonstrated S1 binding to actin is reduced as a function of cMyBP-C concentration in the presence of ATP. In contrast, S1 avidly bound to actin in the absence of ATP and limited cMyBP-C binding, indicating that cMyBP-C and S1 compete for actin binding in an ATP-dependent fashion. However, based on the relationship between thin filament velocity and filament length, the cMyBP-C induced reduction in velocity was independent of the number of crossbridges interacting with the thin filament. In conclusion, the effects of cMyBP-C on velocity and force at both maximal and submaximal activation demonstrate that cMyBP-C does not solely act as a tether between the myosin S2 and LMM subdomains but likely affects both the kinetics and recruitment of myosin cross-bridges through its direct interaction with actin and/or myosin head.

show abstract

Smooth muscle and skeletal muscle myosins produce similar unitary forces and displacements in the laser trap

Cited by 247 publications

References 37 publications

Cardiac myosin missense mutations cause dilated cardiomyopathy in mouse models and depress molecular motor function

Cardiac myosin missense mutations cause dilated cardiomyopathy in mouse models and depress molecular motor function

Spatial and temporal traction response in human airway smooth muscle cells

Cardiac myosin binding protein-C modulates actomyosin binding and kinetics in the in vitro motility assay

Contact Info

Product

Resources

About