Evolutionarily conserved surface residues constitute actin binding sites of tropomyosin

Barua, Bipasha; Pamula, Melissa C.; Hitchcock‐DeGregori, Sarah E.

doi:10.1073/pnas.1101221108

Cited by 68 publications

(127 citation statements)

References 42 publications

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“…1.9 ± 0.1 48.3 0.6 ± 0.5**, n = 4 (29), 202.7 ± 60 1.1 ± 0.1 6.8 ± 0.6 0.3 ± 0.1 † The values for the binding constant of Tm to actin, K app , shown with SE, and T M , the temperature at which the ellipticity at 222 nm, normalized to a scale of 0-1, is equal to 0.5, from Barua et al (23). ‡ Filament speeds ± SD of actin and actin-Tm from in vitro motility assays in the absence of NEM-S1 and Tn.…”

Section: Resultsmentioning

confidence: 99%

“…1) are close to actin residues D25, K326, K328, and P333 (23,33,40). The weak but specific electrostatic interactions with actin place Tm in the closed position in the absence of myosin, partially occupying the myosin binding site on actin.…”

Section: Discussionmentioning

confidence: 99%

“…The Tm mutants used in this study have been described previously (23). Details about actin, troponin, myosin, and myosin S1 are provided in SI Materials and Methods.…”

Section: Methodsmentioning

confidence: 99%

“…We previously reported an evolutionary analysis of Tms from 26 species, ranging from cnidarians to vertebrates, and determined the conserved residues of Tm (23). Mutations at conserved surface positions b, c, and f of the heptad repeat of the Tm coiled-coil revealed that residues in the first halves of Tm's periodic repeats or periods (P) 2-6 of 7 repeats are important for actin affinity.…”

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confidence: 99%

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Regulation of actin-myosin interaction by conserved periodic sites of tropomyosin

Barua

Winkelmann

White

et al. 2012

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

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Cooperative activation of actin-myosin interaction by tropomyosin (Tm) is central to regulation of contraction in muscle cells and cellular and intracellular movements in nonmuscle cells. The steric blocking model of muscle regulation proposed 40 y ago has been substantiated at both the kinetic and structural levels. Even with atomic resolution structures of the major players, how Tm binds and is designed for regulatory function has remained a mystery. Here we show that a set of periodically distributed evolutionarily conserved surface residues of Tm is required for cooperative regulation of actomyosin. Based on our results, we propose a model of Tm on a structure of actin-Tm-myosin in the "open" (on) state showing potential electrostatic interactions of the residues with both actin and myosin. The sites alternate with a second set of conserved surface residues that are important for actin binding in the inhibitory state in the absence of myosin. The transition from the closed to open states requires the sites identified here, even when troponin + Ca 2+ is present. The evolutionarily conserved residues are important for actomyosin regulation, a universal function of Tm that has a common structural basis and mechanism.actin filament structure | cell motility | cytoskeleton | muscle contraction | motility assay A ctin and myosin are found in almost all eukaryotic cells and are required for diverse cellular functions, including muscle contraction, cell motility, cell adhesion, cytokinesis, and organelle transport (1). The actin-myosin interaction produces two types of movements: force generation between actin filaments leading to contractions, such as in muscle contraction, cell motility, and cytokinesis; and transport of subcellular organelles and macromolecular complexes by myosin motors along actin filaments. The actomyosin contractile apparatus is best characterized in striated muscle contraction, which is regulated in a Ca 2+ -dependent manner by the proteins, tropomyosin (Tm) and troponin (Tn).Tm is a universal regulator of the actin cytoskeleton. It is a twochained, α-helical coiled-coil actin binding protein that associates end-to-end to form a continuous strand along both sides of actin filaments and regulates the functions and stability of actin filaments (2, 3). Tm regulates the interaction of actin filaments with actin binding proteins, including myosin, Tn, ADF-cofilin, Arp2/3, formin, and tropomodulin. Tm can activate or inhibit actomyosin, depending on the myosin and Tm isoforms (4-7). The Tm-dependent regulation is widespread, having been reported in fission yeast and mammalian muscle and nonmuscle systems. The regulation of actomyosin by Tm may be viewed as universal, although there is little understanding of the mechanism or structural basis, the focus of the present work.In striated muscle, regulation of myosin binding to the thin filament (actin-Tm-Tn) is described by a three-state model, where the three kinetic states are thought to correspond to three positions of Tm on the actin filament (8-1...

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Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

“…The Tm mutants used in this study have been described previously (23). Details about actin, troponin, myosin, and myosin S1 are provided in SI Materials and Methods.…”

Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 99%

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Regulation of actin-myosin interaction by conserved periodic sites of tropomyosin

Barua

Winkelmann

White

et al. 2012

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

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“…On the basis of sequence and structural analyses and considerations of charged-residue clusters (31,32), Glu-181 was identified as an evolutionarily conserved residue that should play an important role because of close interactions with actin (32). The residue is located in the C-terminal half of Tm period 5 and is likely to function to stabilize the blocked and closed states via electrostatic interactions with actin.…”

mentioning

confidence: 99%

Mechanistic Heterogeneity in Contractile Properties of α-Tropomyosin (TPM1) Mutants Associated with Inherited Cardiomyopathies

Gupte

Haque

Gangadharan

et al. 2015

Journal of Biological Chemistry

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Background: Single residue substitutions in sarcomeric proteins cause most inherited cardiomyopathies. Results: Mutant ␣-tropomyosins cause multiple functional alterations in actin affinity and Ca 2ϩ sensitivity. Conclusion: Mutants follow distinct mechanisms to change Ca 2ϩ sensitivity. Significance: Fluorescence assays to measure changes in troponin C conformation may provide a simple platform for preliminary high throughput screening of modulatory small molecules to treat inherited cardiomyopathies.

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Distinct sites in tropomyosin specify shared and isoform‐specific regulation of myosins II and V

et al. 2018

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Muscle contraction, cytokinesis, cellular movement, and intracellular transport depend on regulated actin-myosin interaction. Most actin filaments bind one or more isoform of tropomyosin, a coiled-coil protein that stabilizes the filaments and regulates interactions with other actin-binding proteins, including myosin. Isoform-specific allosteric regulation of muscle myosin II by actin-tropomyosin is well-established while that of processive myosins, such as myosin V, which transport organelles and macromolecules in the cell periphery, is less certain. Is the regulation by tropomyosin a universal mechanism, the consequence of the conserved periodic structures of tropomyosin, or is it the result of specialized interactions between particular isoforms of myosin and tropomyosin? Here, we show that striated muscle tropomyosin, Tpm1.1, inhibits fast skeletal muscle myosin II but not myosin Va. The non-muscle tropomyosin, Tpm3.1, in contrast, activates both myosins. To decipher the molecular basis of these opposing regulatory effects, we introduced mutations at conserved surface residues within the six periodic repeats (periods) of Tpm3.1, in positions homologous or analogous to those important for regulation of skeletal muscle myosin by Tpm1.1. We identified conserved residues in the internal periods of both tropomyosin isoforms that are important for the function of myosin Va and striated myosin II. Conserved residues in the internal and C-terminal periods that correspond to Tpm3.1-specific exons inhibit myosin Va but not myosin II function. These results suggest that tropomyosins may directly impact myosin function through both general and isoform-specific mechanisms that identify actin tracks for the recruitment and function of particular myosins.

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Evolutionarily conserved surface residues constitute actin binding sites of tropomyosin

Cited by 68 publications

References 42 publications

Regulation of actin-myosin interaction by conserved periodic sites of tropomyosin

Regulation of actin-myosin interaction by conserved periodic sites of tropomyosin

Mechanistic Heterogeneity in Contractile Properties of α-Tropomyosin (TPM1) Mutants Associated with Inherited Cardiomyopathies

Distinct sites in tropomyosin specify shared and isoform‐specific regulation of myosins II and V

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