Cooperative activation of myosin by light chain phosphorylation in permeabilized smooth muscle

Vyas, T. B.; Mooers, Susan U.; Narayan, S.; Witherell, J. C.; Siegman, Marion J.; Butler, Thomas M.

doi:10.1152/ajpcell.1992.263.1.c210

Cited by 113 publications

(51 citation statements)

References 25 publications

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“…Murphy and co-workers (25) proposed that force maintenance was because of the presence of actinattached dephosphorylated cross-bridges that have a significantly reduced rate of detachment and thus maintain force for long periods prior to relaxation. An alternate view proposed by Butler and co-workers (57) suggested that the cycling rate of a given myosin head, regardless of its phosphorylation state, depends on the fraction of phosphorylated heads in the ensemble and is thus modulated as the extent of phosphorylation changes during a contraction. Finally, the Somlyos and their co-workers (58) proposed that latch results from the high ADP affinity of minus-insert myosin leading to a longer attached lifetime, which activates the thin filament and allows cooperative binding of both phosphorylated and dephosphorylated myosin to actin.…”

Section: Discussionmentioning

confidence: 99%

The Unique Properties of Tonic Smooth Muscle Emerge from Intrinsic as Well as Intermolecular Behaviors of Myosin Molecules

Baker

Brosseau

Fagnant

et al. 2003

Journal of Biological Chemistry

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To better understand the molecular basis for some of the unique mechanical properties of tonic smooth muscle, we use a laser trap to assay the mechanochemistry of single smooth muscle heavy meromyosin molecules lacking a seven-amino acid insert in the nucleotide binding loop (minus insert). We measured a second-order ATP-induced actin dissociation rate, k T , of 2. , and an ADP affinity, K D , of 3.2 M, which is more than 100-fold greater than that measured for skeletal muscle myosin. By performing in vitro motility studies under nearly identical conditions, we show that the relatively slow actin velocity generated by minus-insert heavy meromyosin is significantly influenced, but not limited, by k ؊D . Our results support a model in which two separate intermediate steps in the actin-myosin catalyzed ATP hydrolysis reaction are energetically coupled through mechanical interactions, and we discuss this model in the context of the ability of tonic muscle to maintain high forces at low energetic cost (latch).Muscle shortening and force generation result from actinmyosin binding events that are coupled to the actin-myosin catalyzed ATP hydrolysis reaction illustrated in Fig. 1. Upon binding to an actin filament (A) and releasing inorganic phosphate (P i ), myosin undergoes a large and discrete rotation of its lever-like light chain domain, which is capable of generating both motion and force (1-5). With the subsequent release of ADP (at the rate k ϪD ) an additional rotation of the light chain domain of myosin has been observed in smooth muscle myosin (6, 7), but unlike the work generating rotation associated with actin binding/P i release, the rotation associated with ADP release is thought to be a strain-sensing biochemical step (8 -10). Following the release of ADP, ATP binding induces the detachment of myosin from the actin filament (at the rate k T ), after which ATP is hydrolyzed.Muscles differ significantly in their shortening speeds and force-generating capacities. For instance, smooth muscle produces a greater average force per myosin and slower speeds of shortening than skeletal muscle (11). To a large extent these mechanical differences are caused by kinetic differences among the different myosin isoforms that exist within different muscle types (12, 13). For example, phasic and tonic smooth muscle (found in the intestine and aorta respectively) express two myosin heavy chain isoforms that differ by a seven-amino acid insert in a surface loop spanning their nucleotide binding pocket (14 -16). Phasic smooth muscle contains primarily the plus-insert myosin, whereas tonic smooth muscle contains primarily the minus-insert myosin. In addition, two essential light chain isoforms are coordinately expressed with the heavy chain isoforms. The acidic isoform (LC 17a ) is coexpressed with the plus-insert heavy chain whereas the basic isoform (LC 17b ) coexpresses with the minus-insert heavy chain (17-19). Based on in vitro motility studies, the presence or absence of the sevenamino acid insert in the heavy chain is t...

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Section: Discussionmentioning

confidence: 99%

The Unique Properties of Tonic Smooth Muscle Emerge from Intrinsic as Well as Intermolecular Behaviors of Myosin Molecules

Baker

Brosseau

Fagnant

et al. 2003

Journal of Biological Chemistry

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“…Since myosin in tonic smooth muscles contains a greater proportion of LC 17b , has a lower K D for MgADP, and lacks the heavy chain insert, the present finding that the LC 17 isoforms can modulate the V us of nonphosphorylated, cycling cross-bridges may have implications concerning the molecular origin(s) of the latch or catchlike state. Furthermore, under most physiological conditions RLC are only partially phosphorylated, and cooperative cycling of nonphosphorylated cross-bridges under these conditions (20,41) could also be modulated by the LC 17 isoform. We propose that the coordinate expression of a particular LC 17 isoform and the presence or absence of the 7-amino acid insert loop near the catalytic site of the motor domain, leads to the different affinities for MgADP and underlies the tonic (slow) or phasic (fast) contractile phenotypes of smooth muscle (1).…”

Section: Discussionmentioning

confidence: 99%

Myosin Essential Light Chain Isoforms Modulate the Velocity of Shortening Propelled by Nonphosphorylated Cross-bridges

Matthew

Khromov

Trybus

et al. 1998

Journal of Biological Chemistry

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The differential effects of essential light chain isoforms (LC 17a and LC 17b ) on the mechanical properties of smooth muscle were determined by exchanging recombinant for endogenous LC 17 in permeabilized smooth muscle treated with trifluoperazine (TFP). Co-precipitation with endogenous myosin heavy chain verified that 40 -60% of endogenous LC 17a could be exchanged for recombinant LC 17a or LC 17b . Upon addition of MgATP in Ca 2؉ -free solution, recombinant LC 17 exchange induced slow contractions unaccompanied by regulatory light chain (RLC) phosphorylation only in TFP-treated, but not in untreated, permeabilized smooth muscle; the shortening velocity and rate of force development were approximately 1.5 and 2 times faster, respectively, in response to LC 17a than LC 17b . Additional incubation with recombinant, thiophosphorylated RLC increased the shortening velocity, independent of the LC 17 isoform exchanged. The LC 17 -induced contractions of TFPtreated muscles were abolished by prior addition of nonphosphorylated RLC. We suggest that LC 17 stiffens the lever arm of myosin and, in the absence of regulation by RLC, permits cross-bridge cycling without requiring RLC phosphorylation. Our results are compatible with nonphosphorylated RLC acting as a repressor and with LC 17 isoforms modulating the MgADP affinity and, consequently, rate of cooperative cycling of nonphosphorylated cross-bridges.The markedly different rates of contraction and relaxation in fast, phasic and slow, tonic smooth muscles reflect differences not only at the level of the membrane potential, signal transduction, rates of myosin light chain phosphorylation and dephosphorylation, but also in the kinetic properties of the actomyosin motor. The muscle-specific differences in actomyosin kinetics have been ascribed to the existence of specific myosin isoforms, but their relative contributions to the mechanical properties of smooth muscles have not been unequivocally demonstrated (reviewed in Ref. 1). The expression of the two myosin heavy chain (MHC) 1 isoforms, SM-1 and SM-2 (2) that differ by, respectively, the presence or absence of a 34-amino acid extension of the carboxyl terminus (3), does not correlate with phasic or tonic kinetics of contraction. On the other hand, motility assays show different rates of movement propelled by MHC isoforms that are the products of an alternative splicing mechanism resulting in the presence or absence of a 7-amino acid insert near the nucleotide-binding region of the myosin head (4, 5). Disparate results have been obtained concerning the effects on contractile kinetics of the two LC 17 isoforms, acidic (LC 17a ) and basic (LC 17b ), that differ in 5 of the 9 COOHterminal amino acid residues and are products of a single gene generated by an alternative splicing mechanism (6). The faster kinetics of phasic, smooth muscle in situ (7-9) correlates with the expression of the LC 17a isoform (10, 11), albeit the proportion of myosin heavy chain containing the insert was also variant in these muscles, whereas ...

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“…Myosin rods have been expressed in a Dictyostelium myosin null cell line without any obvious signs of instability (T. Egelhoff A more attractive possibility suggested by our results is that an important cooperative aspect of myosin II function has been affected in vivo. In smooth muscle, for example, the activation of myosin II by light chain phosphorylation is a cooperative phenomenon along a myosin filament (24). It is possible that the phosphorylation of the RMLC in the single-headed myosin may not be as efficient as in a double-headed molecule.…”

Section: Discussionmentioning

confidence: 99%

Single-headed myosin II acts as a dominant negative mutation in Dictyostelium.

Burns

Larochelle

Erickson

et al. 1995

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

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Conventional myosin II is an essential protein for cytokinesis, capping of cell surface receptors, and development of Dictyostelium cells. Myosin II also plays an important role in the polarization and movement of cells. All conventional myosins are double-headed molecules but the significance of this structure is not understood since singleheaded myosin II can produce movement and force in vitro. We found that expression of the tail portion of myosin II in Dictyostelium led to the formation of single-headed myosin II in vivo. The resultant cells contain an approximately equal ratio of double-and single-headed myosin II molecules. Surprisingly, these cells were completely blocked in cytokinesis and capping of concanavalin A receptors although development into fruiting bodies was not impaired. We found that this phenotype is not due to defects in myosin light chain phosphorylation. These results show that single-headed myosin II cannot function properly in vivo and that it acts as a dominant negative mutation for myosin II function. These results suggest the possibility that cooperativity of myosin II heads is critical for force production in vivo.

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Cooperative activation of myosin by light chain phosphorylation in permeabilized smooth muscle

Cited by 113 publications

References 25 publications

The Unique Properties of Tonic Smooth Muscle Emerge from Intrinsic as Well as Intermolecular Behaviors of Myosin Molecules

The Unique Properties of Tonic Smooth Muscle Emerge from Intrinsic as Well as Intermolecular Behaviors of Myosin Molecules

Myosin Essential Light Chain Isoforms Modulate the Velocity of Shortening Propelled by Nonphosphorylated Cross-bridges

Single-headed myosin II acts as a dominant negative mutation in Dictyostelium.

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