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Siegman, Marion J.; Mooers, Susan U.; Li, Chenqing; Narayan, S.; Trinkle-Mulcahy, Laura; Watabe, Shugo; Hartshorne, David J.; Butler, Thomas M.

doi:10.1023/a:1018683823020

Cited by 60 publications

(29 citation statements)

References 43 publications

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“…We have shown that the catch state is regulated by the cAMP-dependent phosphorylation of a high molecular mass (Ϸ600 kDa) protein in the intact and permeabilized ABRM (15). Several lines of evidence support this conclusion.…”

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confidence: 56%

Phosphorylation of a twitchin-related protein controls catch and calcium sensitivity of force production in invertebrate smooth muscle

Siegman

Funabara

Kinoshita

et al. 1998

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

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ABSTRACT''Catch'' is a condition of prolonged, highforce maintenance at resting intracellular Ca 2؉ concentration ([Ca 2؉ ]) and very low energy usage, occurring in invertebrate smooth muscles, including the anterior byssus retractor muscle (ABRM) of Mytilus edulis. Relaxation from catch is rapid on serotonergic nerve stimulation in intact muscles and application of cAMP in permeabilized muscles. This release of catch occurs by protein kinase A-mediated phosphorylation of a high (Ϸ600 kDa) molecular mass protein, the regulator of catch. Here, we identify the catch-regulating protein as a homologue of the mini-titin, twitchin, based on (i) a partial cDNA of the purified isolated protein showing 77% amino acid sequence identity to the kinase domain of Aplysia californica twitchin; (ii) a polyclonal antibody to a synthetic peptide in this sequence reacting with the phosphorylated catchregulating protein band from permeabilized ABRM; and (iii) the similarity of the amino acid composition and molecular weight of the protein to twitchin. In permeabilized ABRM, at all but maximum [Ca 2؉ ], phosphorylation of twitchin results in a decreased calcium sensitivity of force production (halfmaximum at 2.5 vs. 1.3 M calcium). At a given submaximal force, with equal numbers of force generators, twitchin phosphorylation increased unloaded shortening velocity Ϸ2-fold. These data suggest that aspects of the catch state exist not only at resting [Ca 2؉ ], but also at higher submaximal [Ca 2؉ ]. The mechanism that gives rise to force maintenance in catch probably operates together, to some extent, with that of cycling myosin crossbridges.Isometric force production in the anterior byssus retractor muscle (ABRM) of Mytilus edulis, initiated by cholinergic nerve stimulation, is maintained for a prolonged period of time after cessation of stimulation. This condition, during which relaxation of force occurs at an extremely slow rate, lasting minutes, or even hours, has been termed ''catch.'' When catch occurs, the intracellular Ca 2ϩ concentration ([Ca 2ϩ ]), which was transiently elevated as a result of the stimulus, has declined to near-resting levels (1, 2). In this invertebrate smooth muscle, calcium activates contraction by direct binding to myosin (3, 4), and its subsequent removal establishes the catch state (5). The catch state is characterized by a marked slowing of crossbridge cycling rate, measured as energy usage (6-8) and mechanical behavior such as force-velocity relations and force redevelopment after a quick release (9-12). Catch exemplifies the high economy of smooth muscle: the ability to maintain force with a low expenditure of energy. In the ABRM, rapid relaxation, or release of catch, occurs on stimulation of serotonergic nerves, a response that is mediated by an increase in cellular cAMP and the activation of protein kinase A (13, 14).We have shown that the catch state is regulated by the cAMP-dependent phosphorylation of a high molecular mass (Ϸ600 kDa) protein in the intact and permeabilized ABRM (15). Seve...

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mentioning

confidence: 56%

Phosphorylation of a twitchin-related protein controls catch and calcium sensitivity of force production in invertebrate smooth muscle

Siegman

Funabara

Kinoshita

et al. 1998

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

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“…The supernatant and pellet were applied to SDS-PAGE. The in vivo molar ratio of twitchin/ myosin is 1:15 (6), and the higher levels of TWD2-S were used to facilitate its detection on gels.…”

Section: Methodsmentioning

confidence: 99%

“…Twitchin phosphorylation is required for relaxation of the muscle from catch. For this cycle to repeat, dephosphorylation of twitchin is necessary (6). Thus, in this scheme, twitchin is a major regulator of the catch state.…”

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confidence: 99%

Myosin Loop 2 Is Involved in the Formation of a Trimeric Complex of Twitchin, Actin, and Myosin

Funabara

Osawa

Ueda

et al. 2009

Journal of Biological Chemistry

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Molluscan smooth muscles, such as mussel anterior byssus retractor muscle (ABRM) 2 and adductor muscle, exhibit a low energy cost phase of tension maintenance termed catch. Catch muscle develops active tension following an increase of the intracellular [Ca 2ϩ ] induced by secretion of acetylcholine. Myosin is activated by direct binding of Ca 2ϩ to the regulatory myosin light chain and initiates a relative sliding between thick and thin filaments (1). After a decrease of intracellular [Ca 2ϩ ] to resting levels, the catch state is formed where tension is maintained over long periods of time with little energy consumption (2, 3). Catch tension is abolished by secretion of serotonin and an increase of intracellular [cAMP] with the resulting activation of cAMP-dependent protein kinase and phosphorylation of twitchin (4, 5). Twitchin phosphorylation is required for relaxation of the muscle from catch. For this cycle to repeat, dephosphorylation of twitchin is necessary (6). Thus, in this scheme, twitchin is a major regulator of the catch state.Molluscan twitchin is known as a myosin-binding protein belonging to the titin/connectin superfamily. It is a single polypeptide of 530 kDa containing multiple repeats of immunoglobulin (Ig) and fibronectin type 3-like motifs in addition to a single kinase domain homologous to the catalytic domain of myosin light chain kinase of vertebrate smooth muscle (7). There are several possible phosphorylation sites in molluscan twitchin recognized by cAMP-dependent protein kinase, and two, D1 and D2, have been identified. The D1 phosphorylation site (Ser-1075) is in the linker region between the 7th and 8th Ig motifs (numbering from the N terminus). The D2 site (Ser-4316) is in the linker region between the 21st and 22nd Ig motifs. Additional sites are found close to D1, but are thought not to be vital for catch regulation.The molecular mechanisms underlying development and maintenance of the catch state have been controversial for several years. One theory proposes that catch reflected attached frozen or slowly cycling cross-bridges (8, 9). What distinguished the attached cross-bridge from the detached relaxed state is not clear. Also it was suggested that interactions between thick filaments, other than cross-bridges, or between thin and thick filaments are responsible for the catch contraction (10). In either of the latter cases, the cross-bridge (myosin head) was not involved.Recently we found that a twitchin fragment including the D2 phosphorylation site and its flanking Ig motifs (TWD2-S) interacted with myosin and actin in a phosphorylation-sensitive manner, and it was suggested that this trimeric complex contributed to tension maintenance in catch (11). TWD2-S bound to a region of the actin molecule known also to interact with loop 2 of myosin that is involved in the ATP-driven movement of myosin with actin (12). In the present study, we show that the myosin loop 2 binds to TWD2-S using competitive cosedimentation assays and isothermal titration calorimetry (ITC). Th...

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“…Of course, force maintenance by these tethers does not require any energy input. Given that there is one twitchin for every 15 myosin molecules (9), there would be a twitchin-mediated link between filaments for every 7-8 myosin cross-bridges. Importantly, phosphorylation causes a decrease in binding of both links, allowing rapid relaxation of catch force and loss of resistance to lengthening.…”

Section: Discussionmentioning

confidence: 99%

“…The protein that is phosphorylated by PKA during relaxation of catch force is twitchin, and it is the phosphorylation state of this protein that determines the presence or absence of the catch state at basal [Ca 2ϩ ] (9,10). Twitchin is so named because Caenorhabditis elegans mutants that lack the protein show nearly constant twitching of body wall muscles (11).…”

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confidence: 99%

The N-terminal Region of Twitchin Binds Thick and Thin Contractile Filaments

Butler

Mooers

Narayan

et al. 2010

Journal of Biological Chemistry

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Catch is a mechanical state present in some invertebrate smooth muscles in which force output and high resistance to muscle lengthening are maintained for long periods of time with very low energy utilization. In the catch state, intracellular [Ca 2ϩ ] is close to basal concentration (1), and redevelopment of force following unloading of the muscle is absent (2). A hypothesis to explain the long term maintenance of catch force was based on a slowing of cycling of myosin attached to actin when activation waned (see Lowy and Millman (3)). On the other hand, Ruegg proposed that catch was maintained by an independent linkage among thick filaments, possibly mediated by paramyosin (4, 5). As will be discussed below, much of the recent evidence supports the idea that catch results from an independent linkage between thick and thin filaments.Catch force is rapidly relaxed by stimulation of serotonergic nerves (6) that results in an increase in cAMP (7) and subsequent activation of cyclic AMP-dependent protein kinase (PKA) 2 (8). The protein that is phosphorylated by PKA during relaxation of catch force is twitchin, and it is the phosphorylation state of this protein that determines the presence or absence of the catch state at basal [Ca 2ϩ ] (9, 10). Twitchin is so named because Caenorhabditis elegans mutants that lack the protein show nearly constant twitching of body wall muscles (11). Twitchin is a mini-titin that is composed of Ig and fibronectin domains as well as a kinase domain near the C terminus (11). The domain organization of twitchin from Mytilus (12) is shown in Fig. 1. The central portion of the molecule shares domain homology with part of the A band portion of titin, and twitchin is known to be associated with thick filaments in catch muscle (13,14). In vitro, twitchin is phosphorylated by PKA with a stoichiometry of about 3 mol of phosphate/mol of twitchin (15). The D1 phosphorylation site is located between the 7th and 8th Ig domain in the N-terminal part of the molecule, and the D2 site is near the C terminus between the 21st and 22nd Ig domain (12,15). Another site of phosphorylation shares a seven-amino acid sequence with the D1 site, is located in the same Ig linker region, and is referred to as DX (16). Also present in the same linker region as D1 and DX is a DFRXXL actin-binding motif (17) similar to those that mediate the binding of smooth muscle myosin light chain kinase to the thin filament (18) and myosin IIIA tail interactions with the thin filament (19).Using an in vitro motility assay, Yamada et al. (20) showed that mechanical aspects of the catch state can be demonstrated using thick filaments reconstituted from purified myosin, F-actin, and twitchin. They found, however, that little if any binding of twitchin to actin occurred in co-sedimentation experiments. On the other hand, Shelud'ko et al. (21) described significant twitchin-actin interactions that were greatly decreased when twitchin was phosphorylated. Their

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Cited by 60 publications

References 43 publications

Phosphorylation of a twitchin-related protein controls catch and calcium sensitivity of force production in invertebrate smooth muscle

Phosphorylation of a twitchin-related protein controls catch and calcium sensitivity of force production in invertebrate smooth muscle

Myosin Loop 2 Is Involved in the Formation of a Trimeric Complex of Twitchin, Actin, and Myosin

The N-terminal Region of Twitchin Binds Thick and Thin Contractile Filaments

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