Equilibrium of the actin-tropomyosin interaction

Wegner, Albrecht

doi:10.1016/0022-2836(79)90204-3

Cited by 199 publications

(158 citation statements)

References 31 publications

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“…5. The form of the unbinding curves fits well with predictions from the linear lattice binding models previous proposed for Tm to actin (47,48). The variation in significance of the first inflection point between batches of actin matches the prediction of Vilfan that large filament sizes potentially produced in vitro would effectively increase the number of gaps present (48).…”

Section: Discussionsupporting

confidence: 82%

Ultra Short Yeast Tropomyosins Show Novel Myosin Regulation

Maytum

Hatch

Konrad

et al. 2008

Journal of Biological Chemistry

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Tropomyosin (Tm) is an ␣-helical coiled-coil actin-binding protein present in all eukaryotes from yeast to man. Its functional role has been best described in muscle regulation; however its much wider role in cytoskeletal actin regulation is still to be clarified. Isoforms vary in size from 284 or 248 amino acids in vertebrates, to 199 and 161 amino acids in yeast, spanning from 7 to 4 actin binding sites respectively. In Saccharomyces cerevisiae, the larger yTm1 protein is produced by an internal 38-amino acid duplication, corresponding to a single actin-binding site. We have produced an ultra-short Tm with only 125 amino acids by removing both of the 38 amino acid repeats from yTm1, with the addition of an Ala-Ser extension used to mimic the essential N-terminal acetylation. This short Tm, and an M1T mutant of it, bind to actin with a similar affinity to most Tms previously studied (K 50% ϳ 0.5 M). However, an equilibrium fluorescence binding assay shows a much greater inhibition of myosin binding to actin than any previously studied Tm. Actin cosedimentation assays show this is caused by direct competition for binding to actin. The M1T mutant shows a reduced inhibition, probably due to weaker end-to-end interactions making it easier for myosin to displace Tm. All previously characterized Tms, although able to sterically block the myosinbinding site, are able to bind to actin along with myosin. By showing that Tm can compete directly with myosin for the same binding site these new Tms provide direct evidence for the steric blocking model. Tropomyosin (Tm)2 appears to be as ubiquitous as actin in eukaryotes (1-3). It is a dimeric protein forming an elongated ␣-helical coiled-coil (4, 5). It is an actin-associated protein, the dimeric units interacting end-to-end along actin filaments to form a continuous strand that wraps around the surface of the actin filament (6, 7). In this way it acts to stabilize actin filaments, as shown by the effect of knockouts of TPM1 in yeast, which have a sickly phenotype due to loss of their actin stress fibers (8, 9). It has been shown to be an essential protein in yeast, with knockouts expressing no Tm being lethal (9). However Tm functions not just as a structural protein, but also a regulatory one. Its role is best understood in the regulation of muscle contraction, where it is directly involved in regulating the myosinactin interaction (reviewed in Ref. 10). Its wider role in regulation of non-muscle actin filaments is less well understood, although it is becoming clear that it plays an important role in functional regulation of the cytoskeleton (11).From its essential roles in cytoskeletal and muscle regulation it is clearly important to understand how Tm functions.Although structurally it appears like a simple rod-like molecule, higher vertebrates express a large diversity of Tm isoforms. Mammals express over 20 isoforms from four different genes (1). As highlighted in the skTm sequence shown in Fig. 1A, alternate splicing of the 9 exon mammalian tropomyosins is limited to ...

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Section: Discussionsupporting

confidence: 82%

Ultra Short Yeast Tropomyosins Show Novel Myosin Regulation

Maytum

Hatch

Konrad

et al. 2008

Journal of Biological Chemistry

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“…The cosedimentation data also show that both TnT and TnT 1 have a relatively small effect on Tm affinity. Tm binding to actin has been described as a polymerization process controlled by two interactions, the single site affinity of Tm to actin which causes the initial binding and the Tm-Tm end-to-end interactions that drive polymerization (28,30). The former will tend to govern the initial concentration at which binding starts, whereas the latter controls the cooperativity of the binding process once nucleation has occurred.…”

Section: ϫ1 Mmentioning

confidence: 99%

A Modulatory Role for the Troponin T Tail Domain in Thin Filament Regulation

Maytum¹,

Geeves²,

Lehrer³

2002

Journal of Biological Chemistry

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In striated muscle the force generating acto-myosin interaction is sterically regulated by the thin filament proteins tropomyosin and troponin (Tn), with the position of tropomyosin modulated by calcium binding to troponin. Troponin itself consists of three subunits, TnI, TnC, and TnT, widely characterized as being responsible for separate aspects of the regulatory process. TnI, the inhibitory unit is released from actin upon calcium binding to TnC, while TnT performs a structural role forming a globular head region with the regulatory TnITnC complex with a tail anchoring it within the thin filament. We have examined the properties of TnT and the TnT 1 tail fragment (residues 1-158) upon reconstituted actin-tropomyosin filaments. Their regulatory effects have been characterized in both myosin S1 ATPase and S1 kinetic and equilibrium binding experiments. We show that both inhibit the actin-tropomyosin-activated S1 ATPase with TnT 1 producing a greater inhibitory effect. The S1 binding data show that this inhibition is not caused by the formation of the blocked B-state but by significant stabilization of the closed C-state with a 10-fold reduction in the C-to M-state equilibrium, K T , for TnT 1 . This suggests TnT has a modulatory as well as structural role, providing an explanation for its large number of alternative isoforms.

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“…First, Tpms are difficult to work with in protein chemistry experiments because, in solution, they are largely unstructured and have a tendency to oligomerize and/ or aggregate. Furthermore, Tpm dimers bind F-actin only with very low affinity (K d ∼10 −3 M) (Wegner, 1979) and their efficient incorporation into an actin filament requires the formation of a head-to-tail Tpm polymer, consisting of homopolymers of Tpm dimers that run along the actin filament (Tobacman, 2008). Finally, Tpms only interact with actin through ionic interactions and, in essence, 'float' above the surface of the actin polymer, making them unlike most other actin-binding proteins (von der Ecken et al, 2015).…”

Section: Introductionmentioning

confidence: 99%

Tropomyosin – master regulator of actin filament function in the cytoskeleton

Gunning

Hardeman

Lappalainen

et al. 2015

Journal of Cell Science

225

228

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Tropomyosin (Tpm) isoforms are the master regulators of the functions of individual actin filaments in fungi and metazoans. Tpms are coiled-coil parallel dimers that form a head-to-tail polymer along the length of actin filaments. Yeast only has two Tpm isoforms, whereas mammals have over 40. Each cytoskeletal actin filament contains a homopolymer of Tpm homodimers, resulting in a filament of uniform Tpm composition along its length. Evidence for this 'master regulator' role is based on four core sets of observation. First, spatially and functionally distinct actin filaments contain different Tpm isoforms, and recent data suggest that members of the formin family of actin filament nucleators can specify which Tpm isoform is added to the growing actin filament. Second, Tpms regulate wholeorganism physiology in terms of morphogenesis, cell proliferation, vesicle trafficking, biomechanics, glucose metabolism and organ size in an isoform-specific manner. Third, Tpms achieve these functional outputs by regulating the interaction of actin filaments with myosin motors and actin-binding proteins in an isoform-specific manner. Last, the assembly of complex structures, such as stress fibers and podosomes involves the collaboration of multiple types of actin filament specified by their Tpm composition. This allows the cell to specify actin filament function in time and space by simply specifying their Tpm isoform composition.

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Equilibrium of the actin-tropomyosin interaction

Cited by 199 publications

References 31 publications

Ultra Short Yeast Tropomyosins Show Novel Myosin Regulation

Ultra Short Yeast Tropomyosins Show Novel Myosin Regulation

A Modulatory Role for the Troponin T Tail Domain in Thin Filament Regulation

Tropomyosin – master regulator of actin filament function in the cytoskeleton

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