Switching Muscles On and Off in Steps: The McKillop-Geeves Three-State Model of Muscle Regulation

Abstract

BJ Classic highlighting the article "Regulation of the interaction between actin and myosin subfragment 1: evidence for three states of the thin filament."

Figure 1

Dual effect of Tm and Tn-Tm on actomyosin subfragment 1 ATPase. Lehrer and Morris (29) showed linear plots of the activation of acto-S1 ATPase by F-actin when measured as a function of added S1, yet they are sigmoidal when either Tn-free Tm or Tn-Tm is additionally present (plateau values at high S1 levels not shown). Although acto-Tn-Tm- stimulated myosin ATPase is Ca²⁺ dependent at all S1 concentrations tested, Tm, both in the presence and absence of Tn and Ca²⁺, inhibits actomyosin ATPase at low S1/F-actin ratios, and stimulates ATPase at high S1/F-actin ratios. Inhibition and activation by Tm are greatest when Tn is present. The observations of Lehrer and Morris (29, 30), like those of Weber and colleagues (25, 26, 27), demonstrated that seemingly complex cooperative interactions depend on the presence of Tm on actin. The studies suggested that an isomerization of myosin takes place in a step after the initial Ca²⁺ activation of the thin filament, laying the foundation of the three-state MG model. This figure, originally published in The Journal of Biological Chemistry, was adapted, with permission, from (29).

Figure 2

Critical evidence provided by McKillop and Geeves (23) for a three-state model of Tn-Tm regulation. Simultaneous titration of light scattering (open circles) to measure actin-myosin binding and pyrene fluorescence (solid circles) to measure the A-R transition of myosin on pyrene-labeled F-actin complexed with Tm and Tn in the presence (A) and absence of Ca²⁺ (B). Butanedione monoxime (BDM) was included to reduce K₂ (the A-R equilibrium) when making these measurements. Solid lines (light scattering) and dotted lines (fluorescence) represent the best fits for a three-state model. In (B), the dashed line represents a fitting that would have satisfied a two-state model but did not fit the data. Note that the sigmoidal data plots in (B) indicate that a simple 1:1 first-order reaction cannot explain both accessibility of actin to myosin and the A-R transition, i.e., they do not agree with a binary on-off thin-filament switching mechanism. This figure was reprinted from (23) with permission.

Figure 3

Schematic representation of the McKillop-Geeves three-state model of thin-filament regulation. (A) “Regulatory units” of the thin filament are illustrated by single lines depicting one Tm molecule lying over seven circles representing actin; myosin is drawn as triangles. The blocked, closed, and open states are indicated. The dynamic equilibrium between states is depicted as a two-step translation of Tm across actin (K_B and K_T) and an isomerization of myosin into a strongly attached configuration (K₂). K_B and K_T are Ca²⁺-sensitive steps. (B) The same transitions represented in alternative cartoon format. The figure in (A) was adapted, with permission, from (48). The figure in (B) was adapted, with permission, from (23). Note that in (A), Tm is drawn as a single molecular unit spanning seven actin subunits; however, in muscle, neighboring Tm molecules polymerize end to end on actin, creating a cable. Once formed, the four-helix bundles between successive Tm molecules are relatively stiff (58). Thus, in situ, Tm appears to form a continuous semi-flexible cable (59, 60) with a persistence length typical of coiled coils (61). Also note that although the cartoon depicts distinct states, the MG model predicted dynamic equilibria between states, both in the absence and presence of myosin and calcium, a critical point for the interpretation of high-resolution structures. In this regard, myosin, Tn, and calcium can be viewed as allosteric effectors that bias the dynamic distribution of Tm states (30).

Figure 4

Three structural states of the thin filament revealed by fiber diffraction of intact muscle and EM reconstruction of isolated and reconstituted thin filaments. (A–C) Fitting crystal structures of actin and Tm to fiber diffraction patterns of intact muscle; reproduced from Holmes (51) with permission. Three actin subunits (white) are shown in each image, the outer two as C_α-chains and the middle one as a surface representation. Tm is shown as a black coiled coil. B-, C-, and M-states (A–C, respectively) are shown from fitting to low-Ca²⁺ (A) and high-Ca²⁺ (B) data, and (C) to data deduced from reconstructions of S1-decorated actin (51, 52, 53). Note the three positions of Tm. The arrow points to proline 333 protruding from the actin surface. Pro333, at the border between the blocked and closed states, is a good landmark for comparing Tm positions. (D) Fitting crystal structures to EM reconstructions of thin filaments highlighting Tm in three distinct positions on actin. Actin and Tm are shown as ribbon structures (actin, white, blue, and cyan; Tm, red, yellow, and green). Red shows the Tm in the low-Ca²⁺, B-/blocked-position covering most of the known myosin binding site on actin. Yellow shows Tm in the myosin-free, high-Ca²⁺, C-/closed-position, exposing most of the myosin site, but not all of it. Green shows the location of Tm when myosin is bound to actin in the open or M-state. This figure was reprinted from Poole et al. (42), with permission.