Tropomyosin and Troponin Regulation of Wild Type and E93K Mutant Actin Filaments from Drosophila Flight Muscle

Bing, Wu; Razzaq, Azam; Sparrow, John C.; Marston, Steven B.

doi:10.1074/jbc.273.24.15016

Cited by 49 publications

(37 citation statements)

References 46 publications

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“…Actin mutants provide an alternative approach to assess specific interactions between tropomyosin and actin. In one such study, tropomyosin suppressed the in vitro motility of E93K actin filaments, leading to the suggestion that tropomyosin binding to the outer domain of actin is affected by charged surface residues on subdomain 2 (16). The present D56A/E57A binding data support the conclusion that acidic residues on subdomain 2 normally act to weaken tropomyosin binding.…”

Section: Discussionsupporting

confidence: 77%

“…In the case of the Ca 2ϩ G-actin, polymerization by Mg 2ϩ addition causes a heterogeneous population of Ca 2ϩ or Mg 2ϩ F-actin, here termed conventional F-actin. Similar to other actins in which the amino acid residues of subdomain 2 have been modified (17,19) or mutated (16), polymerization of D56A/E57A actin was altered under some conditions. Polymerization was normal in the presence of 3 mM MgCl 2 and low ionic strength, but it was necessary to add phalloidin to prevent depolymerization of the mutant filaments under the high salt conditions of the co-sedimentation binding assays (data not shown).…”

Section: Methodsmentioning

confidence: 76%

“…For example, a Drosophila melanogaster actin subdomain 2 mutant, E93K, which inhibits tropomyosin-actin sliding in isolated protein preparations, has been utilized to investigate tropomyosin-based regulation (16). These and other data (17)(18)(19)(20) suggested that actin subdomain 2 may be important for tropomyosin and troponin function.…”

mentioning

confidence: 99%

“…Helical reconstruction was carried out using standard methods (38 -40) as described previously (8,36). (16,17,19) suggest that modification or mutation of actin subdomain 2 can alter interactions of actin with tropomyosin (16). To explore this quantitatively, we examined the binding of tropomyosin to yeast actin mutant D56A/E57A (Fig.…”

mentioning

confidence: 99%

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An Actin Subdomain 2 Mutation That Impairs Thin Filament Regulation by Troponin and Tropomyosin

Korman¹,

Hatch²,

Dixon³

et al. 2000

Journal of Biological Chemistry

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Striated muscle thin filaments adopt different quaternary structures, depending upon calcium binding to troponin and myosin binding to actin. Modification of actin subdomain 2 alters troponin-tropomyosin-mediated regulation, suggesting that this region of actin may contain important protein-protein interaction sites. We used yeast actin mutant D56A/E57A to examine this issue. The mutation increased the affinity of tropomyosin for actin 3-fold. The addition of Ca 2؉ to mutant actin filaments containing troponin-tropomyosin produced little increase in the thin filament-myosin S1 MgATPase rate. Despite this, three-dimensional reconstruction of electron microscope images of filaments in the presence of troponin and Ca 2؉ showed tropomyosin to be in a position similar to that found for muscle actin filaments, where most of the myosin binding site is exposed. Troponin-tropomyosin bound with comparable affinity to mutant and wild type actin in the absence and presence of calcium, and in the presence of myosin S1, tropomyosin bound very tightly to both types of actin. The mutation decreased actin-myosin S1 affinity 13-fold in the presence of troponin-tropomyosin and 2.6-fold in the absence of the regulatory proteins. The results suggest the importance of negatively charged actin subdomain 2 residues 56 and 57 for myosin binding to actin, for tropomyosinactin interactions, and for regulatory conformational changes in the actin-troponin-tropomyosin complex.Cardiac and skeletal muscle contraction is controlled by a complex allosteric system in which myosin-actin interactions are regulated by tropomyosin and troponin. Tropomyosin is an extended, dimeric coiled-coil and spans seven actin monomers on the thin filament. Troponin is composed of three subunits: troponin T connects the other two subunits and tropomyosin to the actin filament, troponin I inhibits myosin binding to actin, and troponin C serves as a Ca 2ϩ sensor for the system (for reviews see Refs. 1 and 2). Structural studies have shown that tropomyosin can bind to three different regions of the actin filament (3-7). In the absence of Ca 2ϩ , tropomyosin appears to bind to subdomain 1 of actin and to bridge over subdomain 2 to the next actin monomer (4). In the presence of Ca 2ϩ , tropomyosin moves toward subdomains 3 and 4 (3) and moves even further in the presence of myosin (8). Whereas these and other results support a three state model of muscle regulation involving tropomyosin movement (9 -12), information defining the position and interactions of troponin on the filament and actin residues that specifically interact with the regulatory proteins is limited.One strategy to examine muscle protein-protein interactions has been to exploit functionally altered mutants (13-15). For example, a Drosophila melanogaster actin subdomain 2 mutant, E93K, which inhibits tropomyosin-actin sliding in isolated protein preparations, has been utilized to investigate tropomyosin-based regulation (16). These and other data (17)(18)(19)(20) suggested that actin subdomain 2 may be ...

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Section: Discussionsupporting

confidence: 77%

Section: Methodsmentioning

confidence: 76%

mentioning

confidence: 99%

mentioning

confidence: 99%

See 2 more Smart Citations

An Actin Subdomain 2 Mutation That Impairs Thin Filament Regulation by Troponin and Tropomyosin

Korman¹,

Hatch²,

Dixon³

et al. 2000

Journal of Biological Chemistry

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“…The effect varies according to myosin type and isoform. Smooth muscle tropomyosin increases actin sliding velocity ϳ10-fold with phosphorylated Limulus striated muscle myosin (54) and 2-4-fold on smooth muscle myosin (55), but only slightly increases velocities on rabbit skeletal HMM (56). On this basis the Emb 18 isoform acts like a smooth muscle myosin, while the IFM myosin behaves more like rabbit skeletal myosin.…”

Section: Figmentioning

confidence: 97%

Alternative Exon-encoded Regions of Drosophila Myosin Heavy Chain Modulate ATPase Rates and Actin Sliding Velocity

Swank¹,

Bartoo

Knowles

et al. 2001

Journal of Biological Chemistry

137

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To investigate the molecular functions of the regions encoded by alternative exons from the single Drosophila myosin heavy chain gene, we made the first kinetic measurements of two muscle myosin isoforms that differ in all alternative regions. Myosin was purified from the indirect flight muscles of wild-type and transgenic flies expressing a major embryonic isoform. The in vitro actin sliding velocity on the flight muscle isoform (6.4 m⅐s ؊1 at 22°C) is among the fastest reported for a type II myosin and was 9-fold faster than with the embryonic isoform. With smooth muscle tropomyosin bound to actin, the actin sliding velocity on the embryonic isoform increased 6-fold, whereas that on the flight muscle myosin slightly decreased. No difference in the step sizes of Drosophila and rabbit skeletal myosins were found using optical tweezers, suggesting that the slower in vitro velocity with the embryonic isoform is due to altered kinetics. Basal ATPase rates for flight muscle myosin are higher than those of embryonic and rabbit myosin. These differences explain why the embryonic myosin cannot functionally substitute in vivo for the native flight muscle isoform, and demonstrate that one or more of the five myosin heavy chain alternative exons must influence Drosophila myosin kinetics.

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The pathogenesis of ACTA1‐related congenital fiber type disproportion

Clarke

Ilkovski

Cooper

et al. 2007

Annals of Neurology

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Tropomyosin and Troponin Regulation of Wild Type and E93K Mutant Actin Filaments from Drosophila Flight Muscle

Cited by 49 publications

References 46 publications

An Actin Subdomain 2 Mutation That Impairs Thin Filament Regulation by Troponin and Tropomyosin

An Actin Subdomain 2 Mutation That Impairs Thin Filament Regulation by Troponin and Tropomyosin

Alternative Exon-encoded Regions of Drosophila Myosin Heavy Chain Modulate ATPase Rates and Actin Sliding Velocity

The pathogenesis of ACTA1‐related congenital fiber type disproportion

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