Caldesmon: binding to actin and myosin and effects on elementary steps in the ATPase cycle

Chalovich, Joseph M.; Şen,; Resetar,; Leinweber,; Fredricksen,; J, Lu; Chen, Guihai

doi:10.1046/j.1365-201x.1998.00449.x

Cited by 45 publications

(28 citation statements)

References 55 publications

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“…The role of caldesmon in the stability of stress fibres and focal adhesions Although the detailed mechanisms by which caldesmon contributes to the regulation of cell motility and cell invasion has not been clearly defined, there is evidence to suggest that caldesmon: (1) regulates contractility by modulating the actinactivated myosin ATPase activity Dabrowska et al, 1996;Chalovich et al, 1998), and (2) stabilises actin stress fibres by inhibiting actin-severing proteins (Ishikawa et al, 1989a;Ishikawa et al, 1989b;Gusev et al, 1994;Dabrowska et al, 1996). Contractility and stress fibre stability are intimately interdependent of each other, such that any mechanism or reagent that inhibits contractility would also perturb stress fibre stability and vice versa (ChrzanowskaWodnicka and Burridge, 1996;Choquet et al, 1997;Hirose et al, 1998;Rottner et al, 1999;Riveline et al, 2001;Balaban et al, 2001;Tsubouchi et al, 2002;Burgstaller and Gimona, 2004).…”

Section: Discussionmentioning

confidence: 99%

“…There is strong in vitro evidence implicating caldesmon in the regulation of smooth muscle contraction and cell motility. Caldesmon inhibits the actin-activated myosin ATPase (Dabrowska et al, 1985;Smith et al, 1987;Horiuchi and Chacko, 1989;Chalovich et al, 1998) by blocking the interaction of actin and myosin (Chalovich et al, 1998;Sen et al, 2001) and/or inhibiting a kinetic step of the actomyosin ATPase cycle Marston et al, 1998). Inhibition of contraction by caldesmon can be released by binding of caldesmon to Ca 2+ /calmodulin and/or posttranslational phosphorylation.…”

Section: Introductionmentioning

confidence: 99%

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Caldesmon is an integral component of podosomes in smooth muscle cells

Eves

Webb

Zhou

et al. 2006

Journal of Cell Science

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Podosomes are highly dynamic actin-based structures commonly found in motile and invasive cells such as macrophages, osteoclasts and vascular smooth muscle cells. Here, we have investigated the role of caldesmon, an actinbinding protein, in the formation of podosomes in aortic smooth muscle A7r5 cells induced by the phorbol ester PDBu. We found that endogenous low molecular weight caldesmon (l-caldesmon), which was normally localised to actin-stress fibres and membrane ruffles, was recruited to the actin cores of PDBu-induced podosomes. Overexpression of l-caldesmon in A7r5 cells caused dissociation of actin-stress fibres and disruption of focal adhesion complexes, and significantly reduced the ability of PDBu to induce podosome formation. By contrast, siRNA interference of caldesmon expression enhanced PDBuinduced formation of podosomes. The N-terminal fragment of l-caldesmon, CaD40, which contains the myosin-binding site, did not label stress fibres and was not translocated to PDBu-induced podosomes. Cad39, the C-terminal fragment housing the binding sites for actin, tropomyosin and calmodulin, was localised to stress fibres and was translocated to podosomes induced by PDBu. The caldesmon mutant, CadCamAB, which does not interact with Ca 2+ /calmodulin, was not recruited to PDBu-induced podosomes. These results show that (1) l-caldesmon is an integral part of the actin-rich core of the podosome; (2) overexpression of l-caldesmon suppresses podosome formation, whereas siRNA knock-down of l-caldesmon facilitates its formation; and (3) the actin-binding and calmodulin-binding sites on l-caldesmon are essential for the translocation of l-caldesmon to the podosomes. In summary, this data suggests that caldesmon may play a role in the regulation of the dynamics of podosome assembly and that Ca 2+ /calmodulin may be part of a regulatory mechanism in podosome formation.

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Caldesmon is an integral component of podosomes in smooth muscle cells

Eves

Webb

Zhou

et al. 2006

Journal of Cell Science

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“…We propose in this work that this latter mechanism involves interference with the myosin-induced tropomyosin movement. This duality includes the myosin ATP-blocking model of Chalovich and colleagues (23,28,68,69) and the allosteric inhibitory mechanism of Marston and colleagues (14,27,70).…”

Section: Discussionmentioning

confidence: 99%

Effect of Caldesmon on the Position and Myosin-induced Movement of Smooth Muscle Tropomyosin Bound to Actin

Graceffa

Mazurkie

2005

Journal of Biological Chemistry

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It is known that the actin-binding protein caldesmon inhibits actomyosin ATPase activity and might in this way take part in the thin filament regulation of smooth muscle contraction. Although the molecular mechanism of this inhibition is unknown, it is clear that the presence of actin-bound tropomyosin is necessary for full inhibition. Recent evidence also suggests that the myosin-induced movement of tropomyosin plays a key role in regulation. In this work, fluorescence studies provide evidence to show that caldesmon interacts with and alters the position of tropomyosin in a reconstituted actin thin filament and thereby limits the ability of myosin heads to move tropomyosin. Caldesmon interacts with the Cys-190 region in the COOH-terminal half of tropomyosin, resulting in the movement of this part of tropomyosin to a new position on actin. Additionally, this constrains the myosin-induced movement of this region of tropomyosin. On the other hand, caldesmon does not appear to interact with the Cys-36 region in the NH 2 -terminal half of tropomyosin and neither alters the position of nor significantly constrains the myosin-induced movement of this part of tropomyosin. The ability of caldesmon to limit the myosin-induced movement of tropomyosin provides a possible molecular basis for the inhibitory function of caldesmon. The different movements of the two halves of tropomyosin indicate that actin-bound tropomyosin moves as a flexible molecule and not as a rigid rod. Interestingly, caldesmon, which inhibits tropomyosin's potentiation of actomyosin ATPase activity, moves tropomyosin in one direction, whereas myosin heads, which enhance potentiation, move tropomyosin in the opposite direction.Muscle contraction takes place when myosin heads, projecting from the thick filament, cyclically interact with actin protomers in the thin filament, causing the filaments to slide past each other with a resulting muscle shortening. Actin-activated myosin hydrolysis of ATP provides the energy for this process. Tropomyosin, a long coiled-coil dimer, which binds end-to-end on each side of the thin filament, is involved in switching the muscle on. In skeletal muscle, contraction is switched on by Ca 2ϩ binding to actin-bound troponin, which results in the relocation of tropomyosin and thus the exposure of myosinbinding sites on actin (2-5). The fact that each tropomyosin molecule covers seven actin protomers and binds end-to-end to itself endows the switching process with cooperativity (for a review, see Refs. 6 -9).Smooth muscle does not contain troponin, and the main regulatory switch is the phosphorylation of myosin light chains (for a review, see Refs. 10 and 11). However, myosin phosphorylation does not account for all aspects of smooth muscle regulation, i.e. there can be a decoupling between force and myosin phosphorylation levels, and thus it has been put forth that the actin thin filament and its associated proteins also play a role in regulation (for a review, see Refs. 12 and 13). The actin-binding protein caldesmon has bee...

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“…A competition model proposes that ATPase inhibition by caldesmon is caused by a reduction in the formation of the weakly bound actomyosin complex as a result of caldesmon binding to the same site on actin as S1⅐ADP⅐P i (13). This is supported by the inability of caldesmon to inhibit the crosslinked actin-S1 complex, which does not dissociate on ATP binding (14) and the reduction of the K m of the steady state ATPase (8).…”

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confidence: 99%

The Mechanism of Smooth Muscle Caldesmon-Tropomyosin Inhibition of the Elementary Steps of the Actomyosin ATPase

Alahyan

Webb

Marston

et al. 2006

Journal of Biological Chemistry

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Caldesmon is a component of smooth muscle thin filaments that inhibits the actomyosin ATPase via its interaction with actin-tropomyosin. We have performed a comprehensive transient kinetic characterization of the actomyosin ATPase in the presence of smooth muscle caldesmon and tropomyosin. At physiological ratios of caldesmon to actin (1 caldesmon/7 actin monomers) actomyosin ATPase is inhibited by about 75%. Inhibitory caldesmon concentrations had little effect upon the rate of S1 binding to actin, actin-S1 dissociation by ATP, and dissociation of ADP from actin-S1⅐ADP; however the rate of phosphate release from the actin-S1⅐ADP⅐P i complex was decreased by more than 80%. In addition the transient of phosphate release displayed a lag of up to 200 ms. The presence of a lag phase indicates that a step on the pathway prior to phosphate release has become rate-limiting. Premixing the actin-tropomyosin filaments with myosin heads resulted in the disappearance of the lag phase. We conclude that caldesmon inhibition of the rate of phosphate release is caused by the thin filament being switched by caldesmon to an inactive state. The active and inactive states correspond to the open and closed states observed in skeletal muscle thin filaments with no evidence for the existence of a third, blocked state. Taken together these data suggest that at physiological concentrations, caldesmon controls the isomerization of the weak binding complex to the strong binding complex, and this causes the inhibition of the rate of phosphate release. This inhibition is sufficient to account for the inhibition of the steady state actomyosin ATPase by caldesmon and tropomyosin.Smooth muscle contractility and actomyosin ATP hydrolysis are regulated by Ca 2ϩ at two levels. The first involves phosphorylation of the myosin light chain by myosin light chain kinase upon stimulation by Ca 2ϩ -calmodulin. The second occurs via the thin filament-associated protein caldesmon. Caldesmon (CaD) 2 binds to actin, calmodulin, and tropomyosin (1) and inhibits the actin activation of both smooth and skeletal muscle myosin ATPase in vitro (2) and force production when added to smooth and skeletal muscle fibers (3-5). Caldesmon also regulates the actin activation of myosin I ATPase in smooth muscle (6). The smooth muscle thin filament is Ca 2ϩ -regulated and is made up of actin, tropomyosin, caldesmon, and a Ca 2ϩ -binding protein in molar ratios ϳ14:2:1:1 (7). Previous studies of the molecular mechanism of caldesmon inhibition of the actomyosin ATPase have focused on its effect on the ATPase steady state parameters K m and V max (8) and on the equilibrium binding of myosin to actin filaments (9 -12). A detailed mechanism by which caldesmon regulates the actin activation of myosin ATPase and force production has not been demonstrated by such experiments but two potentially conflicting models have been developed.A competition model proposes that ATPase inhibition by caldesmon is caused by a reduction in the formation of the weakly bound actomyosin complex as a...

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Caldesmon: binding to actin and myosin and effects on elementary steps in the ATPase cycle

Cited by 45 publications

References 55 publications

Caldesmon is an integral component of podosomes in smooth muscle cells

Caldesmon is an integral component of podosomes in smooth muscle cells

Effect of Caldesmon on the Position and Myosin-induced Movement of Smooth Muscle Tropomyosin Bound to Actin

The Mechanism of Smooth Muscle Caldesmon-Tropomyosin Inhibition of the Elementary Steps of the Actomyosin ATPase

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